Reference List 2007, 2, 27

細胞の誤謬に関連する文献の一覧。細胞バンクで収録してきたものだが、メドラインか

らも多数収録した。最近 Science でも細胞の誤謬が取り上げられた(1)ことからもその重要

性が理解できる。誤った細胞を使って研究を行うと、その後で解釈の再考が要求されたり、

目的と異なる解釈をしなければならなくなったりと、とかく研究のコスト上昇を招くなど

の問題が発生するので要注意。文献リストは Reference Manager を利用して細胞バンク内

で独自のデータベースとして管理している。1960-70 年代は HeLa 細胞の混入が大きな話題

になったが、現在では、STR-PCR 法の普及によりそれ以外の細胞間でのクロスコンタミが

数多く検出されるようになってきた。(JCRB 細胞バンク)。 1) Chatterjee R. Cell biology. Cases of mistaken identity. Science 2007 Feb 16;315(5814):928-31. Ref ID: 6120 2) Ciftcioglu N, McKay DS, Mathew G, Kajander EO. Nanobacteria: fact or fiction?

Characteristics, detection, and medical importance of novel self-replicating, calcifying nanoparticles. J Investig Med 2006 Nov;54(7):385-94.

Ref ID: 6112 3) Mohamed IS, Wynn RJ, Cominsky K, Reynolds AM, Ryan RM, Kumar VH, et al. White blood

cell left shift in a neonate: a case of mistaken identity. J Perinatol 2006 Jun;26(6):378-80. Ref ID: 6122 Abstract: We present a full-term male infant who presented with tachypnea and an increased band

count on his complete blood count (CBC) with an immature to total neutrophil (I:T) ratio of 0.6 raising suspicion of early onset sepsis. A blood culture was drawn and he was started on appropriate antibiotics. The patient's clinical condition rapidly improved; however, the white cell count 'left shift' persisted. When a detailed family history was obtained, it was discovered that the father, paternal uncle and the grandfather had been diagnosed with Pelger-Huet anomaly (PHA). As the urine, blood and CSF cultures were all negative in this now well-appearing infant, the left shift on the CBC was believed to be due to inheritance of the PHA. We present this case to emphasize that even in this age of sophisticated laboratory evaluation, a good clinical history, including family history, and clinical evaluation, are essential for accurate diagnosis

Notes: Department of Pediatrics, Division of Neonatology, School of Medicine and Biomedical Sciences, State University of New York at Buffalo, Women and Children's Hospital of Buffalo, Buffalo NY 14222, USAFAU - Mohamed, I S I

4) Scotting PJ. Are cranial germ cell tumours really tumours of germ cells? Neuropathol Appl

Neurobiol 2006 Dec;32(6):569-74. Ref ID: 6121 Abstract: Germ cell tumours of the brain and those that occur in the gonads are believed to share a

common origin from germ cell progenitors. This 'germ cell theory' rests upon similar histopathology between these tumours in different locations and the belief that endogenous somatic cells of the brain could not give rise to the range of cell types seen in germ cell tumours. An alternative 'embryonic cell theory' has been proposed for some classes of cranial germ cell tumours, but this still relies on the misplacement of cells in the brain (in this case the earliest embryonic stem cells) during early embryonic development. Recent evidence has demonstrated that neural stem cells of the brain can also give rise to many of the cell types seen in germ cell tumours. These data suggest that endogenous progenitor cells of the brain are a plausible alternative origin for these tumours. This idea is of central importance for studies aiming to elucidate the mechanisms of tumour development. The application of modern

molecular analyses to reveal how tumour cells have altered with respect to their cell of origin relies on the certain identification of the cell from which the particular tumour arose. If the identity of this cell is mistaken, then studies to elucidate the mechanisms by which the progenitor cell has been subverted from its normal behaviour will not yield useful information. In addition, it will prove impossible to generate an appropriate animal model in which to study the underlying causes of those tumours. This article makes the case that current assumptions of the origins of cranial germ cell tumours are unreliable. It reviews the evidence in favour of the 'germ cell theory' and argues in favour of a 'brain cell theory' in which endogenous neural progenitor cells of the brain are the likely origin for these tumours. Thus, the case is made that cranial germ cell tumours, like other brain tumours, arise by the transformation of progenitor cells normally resident in the brain

Notes: Children's Brain Tumour Research Centre, Institute of Genetics, University of Nottingham, Nottingham, UK paulscotting@nottinghamacukFAU - Scotting, P J

5) Chan WL, Pejnovic N, Liew TV, Hamilton H. Predominance of Th2 response in human

abdominal aortic aneurysm: mistaken identity for IL-4-producing NK and NKT cells? Cell Immunol 2005 Feb;233(2):109-14.

Ref ID: 6124 Abstract: Abdominal aortic aneurysm (AAA) is a complex remodeling process that involves both

synthesis and degradation of extracellular matrix proteins in the aortic wall, leading to decreased tensile strength, progressive dilation and eventual rupture. Chronic inflammation, increased local production of elastin-degrading proteases by inflammatory cells and destruction of medial elastic lamellae play important roles in aneurysm progression. Neovascularization in all layers of the arterial wall is prominent and angiogenesis can facilitate chronic inflammation. It is still unclear what initiates aneurysmal dilation and what determines its progression. The complex nature of the process has defied elucidation. Apart from macrophages, the predominant immune cell infiltrates reported so far are CD3(+)T cells that express CD4 and CD8. Infiltrates of type 2 Th cells and their production of IL-4 and IL-5 have been implicated in AAA development. However, NKT and NK cells have a Th0 cytokine profile and can also produce type 2 as well as type 1 (IL-2 and IFNgamma) cytokines. We have demonstrated the presence of NK and NKT cells in AAA tissue. With their growing importance in autoimmunity and transplantation, they may play a role in AAA development. Therefore, there is a need to use a combination of T and NK markers to fully characterize both innate and adaptive lymphoid cell subsets in local inflammatory infiltrates in order to elucidate their roles in AAA progression

Notes: Biochemical Pharmacology, William Harvey Research Institute, John Vane Science Centre, Queen Mary, University of London, Charterhouse Square, London, EC1M 6BQ, UK wlchan@qmulacukFAU - Chan, Woon Ling

6) Dirks WG, Faehnrich S, Estella IA, Drexler HG. Short tandem repeat DNA typing provides an

international reference standard for authentication of human cell lines. ALTEX 2005;22(2):103-9.

Ref ID: 5921 Abstract: Cell lines have wide applications as model systems in the medical and pharmaceutical

industry. Much drug and chemical testing is now first carried out exhaustively on in vitro systems, reducing the need for complicated and invasive animal experiments. The basis for any research, development or production program involving cell lines is the choice of an authentic cell line. Microsatellites in the human genome that harbour short tandem repeat (STR) DNA markers allow individualisation of established cell lines at the DNA level. Fluorescence polymerase chain reaction amplification of eight highly polymorphic microsatellite STR loci plus gender determination was found to be the best tool to screen the uniqueness of DNA profiles in a fingerprint database. Our results demonstrate that cross-contamination and misidentification remain chronic problems in the use of human continuous cell lines. The combination of rapidly generated DNA types based on single-locus STR and their

authentication or individualisation by screening the fingerprint database constitutes a highly reliable and robust method for the identification and verification of cell lines

Notes: German Collection of Microorganisms and Cell Cultures, Department of Human and Animal Cell Cultures, D-Braunschweig WDI@DSMZdeFAU - Dirks, Wilhelm Gerhard

7) Kreymborg K, Bohlmann U, Becher B. IL-23: changing the verdict on IL-12 function in

inflammation and autoimmunity. Expert Opin Ther Targets 2005 Dec;9(6):1123-36. Ref ID: 6123 Abstract: IL-12 and IL-23 are molecules mainly produced by activated accessory and

antigen-presenting cells. The tools for studying the biology of IL-12 in man and laboratory rodents have greatly advanced our appreciation of the central role of this molecule in cell-mediated immunity and inflammation. In particular, IL-12 is thought to be the prime-regulator of TH1 development. Targeting what was thought to be IL-12 function in vivo, resulted in drastic amelioration of inflammation and autoimmunity firmly linking TH1 polarisation to autoimmune development. Upon discovery of IL-23 and the fact that the large subunit of IL-23 is shared by IL-12, the research community only begins to grasp that the features attributed to IL-12 and TH1 development in inflammation are, in fact, dependent on IL-23 and not on IL-12. Hence, the perception of IL-12 biology is, to a large extent, based on a mistaken identity. In this review, the authors provide an overview of their current understanding of IL-12 and IL-23 biology in inflammation and autoimmunity, and how this viewpoint has been readjusted over the past 15 years

Notes: Department of Neurology, Universitatsspital/University of Zurich, Frauenklinikstrasse 10, CH-8091 Zurich, SwitzerlandFAU - Kreymborg, Katharina

8) Lie AK, Risberg B, Borge B, Sandstad B, Delabie J, Rimala R, et al. DNA- versus RNA-based

methods for human papillomavirus detection in cervical neoplasia. Gynecol Oncol 2005 Jun;97(3):908-15.

Ref ID: 5922 Abstract: OBJECTIVE: To compare DNA-based and mRNA-based methods for detection of

high-grade cervical neoplasia in Norway. METHODS: HPV prevalence was analyzed in 383 women with positive index cytology, selected from gynecology clinics. All patients were investigated by a new PAP smear, histology, and two commercially available HPV tests: Hybrid Capture II (Digene, Gaithersburg, MD) and the Pre Tect HPV-Proofer (NorChip AS). Cases with positive DNA test and negative mRNA test and cases with high-grade histology and negative HPV tests were retested with PCR and sequencing. We regarded the infection as latent or transient if sequencing revealed an HPV type included in both assays. RESULTS: High-risk HPV was detected in 99.7% of the histological confirmed high-grade lesions (CIN2+) (290/291). The DNA test was positive in 95% (275/291), and the mRNA test was positive in 77% (225/291) of the histological confirmed high-grade lesions. All invasive carcinomas were mRNA positive. The DNA test was significantly more often positive in benign and low-grade lesions, some of which were found to be false positive due to cross-contamination with unrelated types. High-grade histology was detected in 83% of women with normal cytology and positive mRNA test. Latent or transient infections were detected in 11 low-grade and 12 high-grade preinvasive lesions. Sequencing revealed high-risk HPV types included only in the DNA test in 35 high-grade preinvasive lesions, HPV 52 and 58 were the most prevalent HPV types. CONCLUSIONS: These HPV tests have the potential to improve the detection rate of high-grade cervical neoplasia, with some limitations. The mRNA test seems to be more appropriate for risk-evaluation. Larger scale, population based studies are necessary to evaluate the predictive values of HPV testing in Norway

Notes: Department of Pathology, The Norwegian Radium Hospital, Oslo, Norway agneskathrinelie@ahusnoFAU - Lie, A K

9) Melcher R, Maisch S, Koehler S, Bauer M, Steinlein C, Schmid M, et al. SKY and genetic

fingerprinting reveal a cross-contamination of the putative normal colon epithelial cell line

NCOL-1. Cancer Genet Cytogenet 2005 Apr 1;158(1):84-7. Ref ID: 5923 Abstract: In vitro studies addressing the primary prevention of colon carcinoma are preferably

conducted using normal colonic cells, because these cells are more likely to represent the potential target for prevention in vivo. Established cell lines of normal colonic origin are mostly lacking; however, this is probably due to the difficulties associated with establishment of such cell lines. Cross-contamination with malignant cells is a frequent event, and so any successfully established cell line of normal origin should be scrutinized prior to further investigation. We performed a cytogenetic (spectral karyotyping) and genetic fingerprint (Promega PowerPlex ES multiplex system and Applied Biosystems AmpFlSTR SGM Plus multiplex system) analysis of the putative normal colon epithelial cell line NCOL-1, derived from two different sources (NCOL-1a and 1b). We show that NCOL-1a and 1b are probably derived from the colon carcinoma cell line LoVo, with a matching probability of 99.9995, most probably through cross-contamination. Karyotypes of LoVo and NCOL-1a were identical; NCOL-1b displayed additional marker chromosomes. Our findings highlight the importance of molecular and cytogenetic characterization of established cell lines to avoid drawing misleading conclusions from the original findings

Notes: Institute of Human Genetics, University of Wuerzburg, Gastrolabor/Bau 4, Joseph-Schneider-Str 2, 97074 Wuerzburg, Germany melcher_r@klinikuni-wuerzburgdeFAU - Melcher, Ralph

10) Puvanachandra N, Rene C. A unique case of mistaken identity. Eye 2005

Jan;19(1):108-10. Ref ID: 6125 11) Bassing CH, Alt FW. Molecular biology: case of mistaken identity. Nature 2004 Mar

4;428(6978):29-31. Ref ID: 6126 12) Buehring GC, Eby EA, Eby MJ. Cell line cross-contamination: how aware are

Mammalian cell culturists of the problem and how to monitor it? In Vitro Cell Dev Biol Anim 2004 Jul;40(7):211-5.

Ref ID: 5924 Abstract: HeLa was the first human cell line established (1952) and became one of the most

frequently used lines because of its hardiness and rapid growth rate. During the next two decades, the development of other human cell lines mushroomed. One reason for this became apparent during the 1970s, when it was demonstrated that many of these cell lines had been overgrown and replaced by fast-growing HeLa cells inadvertently introduced into the original cultures. Although the discovery of these "HeLa contaminants" prompted immediate alarm, how aware are cell culturists today of the threat of cell line cross-contamination? To answer this question, we performed a literature search and conducted a survey of 483 mammalian cell culturists to determine how many were using HeLa contaminants without being aware of their true identity and how many were not using available means to ensure correct identity. Survey respondents included scientists, staff, and graduate students in 48 countries. HeLa cells were used by 32% and HeLa contaminants by 9% of survey respondents. Most were also using other cell lines; yet, only about a third of respondents were testing their lines for cell identity. Of all the cell lines used, 35% had been obtained from another laboratory instead of from a repository, thus increasing the risk of false identity. Over 220 publications were found in the PubMed database (1969-2004) in which HeLa contaminants were used as a model for the tissue type of the original cell line. Overall, the results of this study indicate a lack of vigilance in cell acquisition and identity testing. Some researchers are still using HeLa contaminants without apparent awareness of their true identity. The consequences of cell line cross-contamination can be spurious scientific conclusions; its prevention can save time, resources, and scientific reputations

Notes: School of Public Health, University of California, Berkeley, California 94720, USA buehring@uclink4berkeleyeduFAU - Buehring, Gertrude Case

13) Buehring GC, Eby EA, Eby MJ. Cell line cross-contamination: how aware are

Mammalian cell culturists of the problem and how to monitor it? In Vitro Cell Dev Biol Anim 2004 Jul;40(7):211-5.

Ref ID: 6110 Abstract: HeLa was the first human cell line established (1952) and became one of the most

frequently used lines because of its hardiness and rapid growth rate. During the next two decades, the development of other human cell lines mushroomed. One reason for this became apparent during the 1970s, when it was demonstrated that many of these cell lines had been overgrown and replaced by fast-growing HeLa cells inadvertently introduced into the original cultures. Although the discovery of these "HeLa contaminants" prompted immediate alarm, how aware are cell culturists today of the threat of cell line cross-contamination? To answer this question, we performed a literature search and conducted a survey of 483 mammalian cell culturists to determine how many were using HeLa contaminants without being aware of their true identity and how many were not using available means to ensure correct identity. Survey respondents included scientists, staff, and graduate students in 48 countries. HeLa cells were used by 32% and HeLa contaminants by 9% of survey respondents. Most were also using other cell lines; yet, only about a third of respondents were testing their lines for cell identity. Of all the cell lines used, 35% had been obtained from another laboratory instead of from a repository, thus increasing the risk of false identity. Over 220 publications were found in the PubMed database (1969-2004) in which HeLa contaminants were used as a model for the tissue type of the original cell line. Overall, the results of this study indicate a lack of vigilance in cell acquisition and identity testing. Some researchers are still using HeLa contaminants without apparent awareness of their true identity. The consequences of cell line cross-contamination can be spurious scientific conclusions; its prevention can save time, resources, and scientific reputations

Notes: School of Public Health, University of California, Berkeley, California 94720, USA buehring@uclink4berkeleyeduFAU - Buehring, Gertrude Case

14) Cheung ST, Allan RN. Mistaken identity: misclassification of TPMT phenotype

following blood transfusion. Eur J Gastroenterol Hepatol 2003 Nov;15(11):1245-7. Ref ID: 6128 Abstract: Azathioprine (AZA) is an effective treatment for inflammatory bowel disease. The

measurement of thiopurine methyltransferase (TPMT) activity levels helps to identify the one in 300 patients who are at risk of profound myelosuppression with standard doses of AZA. Thus, it is important that the measurement of TPMT activity is accurate. We report a case of misclassification of TPMT phenotype caused by prior blood transfusion. The patient appeared to have an intermediate level of TPMT enzyme activity as measured after the blood transfusion. However, following severe myelosuppression soon after starting AZA, her genotype was determined, which showed that she was homozygous for low TPMT activity. This report reviews some of the limitations in determining both the phenotype and the genotype of TPMT, especially following recent blood transfusion. The dilemma of whether or not to wait for the TPMT activity result before starting AZA is also discussed

Notes: Gastroenterology Unit, The Queen Elizabeth Hospital, Edgbaston, Birmingham, UK drstc88@hotmailcomFAU - Cheung, Seau-Tak

15) Drexler HG, Dirks WG, Matsuo Y, MacLeod RA. False leukemia-lymphoma cell lines: an

update on over 500 cell lines. Leukemia 2003 Feb;17(2):416-26. Ref ID: 5927 Abstract: Human leukemia-lymphoma (LL) cell lines represent an extremely important resource for

research in a variety of fields and disciplines. As the cell lines are used as in vitro model systems in lieu of primary cell material, it is crucial that the cells in the culture flasks faithfully

correspond to the purported objects of study. Obviously, proper authentication of cell line derivation and precise characterization are indispensable requirements to use as model systems. A number of studies has shown an unacceptable level of LL cell lines to be false. We present here the results of authenticating a comprehensively large sample (n = 550) of LL cell lines mainly by DNA fingerprinting and cytogenetic evaluation. Surprisingly, near-identical incidences (ca 15%) of false cell lines were observed among cell lines obtained directly from original investigators (59/395: 14.9%) and from secondary sources (23/155: 14.8%) implying that most cross-contamination is perpetrated by originators, presumably during establishment. By comparing our data with those published, we were further able to subclassify the false cell lines as (1) virtual: cross-contaminated with and unretrievably overgrown by other cell lines during initiation, never enjoying independent existence; (2) misidentified: cross-contaminated subsequent to establishment so that an original prototype may still exist; or (3) misclassified: unwittingly established from an unintended (often normal) cell type. Prolific classic leukemia cell lines were found to account for the majority of cross-contaminations, eg CCRF-CEM, HL-60, JURKAT, K-562 and U-937. We discuss the impact of cross-contaminations on scientific research, the reluctance of scientists to address the problem, and consider possible solutions. These findings provide a rationale for mandating the procurement of reputably sourced LL cell lines and their regular authentication thereafter

Notes: DSMZ-German Collection of Microorganisms and Cell Cultures, Department of Human and Animal Cell Cultures, Braunschweig, GermanyFAU - Drexler, H G

16) Liu MY, Lin SC, Liu H, Candal F, Vafai A. Identification and authentication of animal

cell culture by polymerase chain reaction amplification and DNA sequencing. In Vitro Cell Dev Biol Anim 2003 Nov;39(10):424-7.

Ref ID: 5925 Abstract: Polymerase chain reaction (PCR) amplification and deoxyribonucleic acid (DNA)

sequence analysis were used to identify the species origin of cell lines used in a cell culture facility where various cell lines of different species are routinely propagated. The aldolase gene family was selected for PCR amplification because the DNA sequences of this gene are highly conserved over a wide range of animals and humans. A total of 36 cell lines representing 13 different species were selected for this study. The DNA from each cell line was amplified, and PCR products were analyzed by agarose gel electrophoresis. The results showed unique profiles of amplified bands on agarose gels that allowed differentiation among non-closely related species. However, DNA amplification of closely related species, including rat and mouse or human and primate, resulted in similar and indistinguishable banding patterns that could be further differentiated by DNA sequence analysis. These results suggested that aldolase gene amplification coupled with DNA sequence analysis is a useful tool for identification of cell lines and has potential application for use in identification of interspecies cross-contamination

Notes: Biologics Branch, Scientific Resources Program, National Center for Infectious Diseases, Centers for Disease Control and Prevention, 1600 Clifton Road, MS-D43, Atlanta, Georgia 30333, USA mkl6@cdcgovFAU - Liu, Merry Y

17) Prakriya M, Lewis RS. CRAC channels: activation, permeation, and the search for a

molecular identity. Cell Calcium 2003 May;33(5-6):311-21. Ref ID: 6129 Abstract: The Ca2+ release-activated Ca2+ (CRAC) channel is a highly Ca2+-selective

store-operated channel that is expressed in T lymphocytes, mast cells, and other hematopoietic cells. In T cells, CRAC channels are essential for generating the prolonged intracellular Ca2+ ([Ca2+](i)) elevation required for the expression of T-cell activation genes. Here we review recent work addressing CRAC channel regulation, pore properties, and the search for CRAC channel genes. Of the current models for CRAC current (I(CRAC)) activation, several new studies argue against a conformational coupling mechanism in which IP(3) receptors communicate store depletion to CRAC channels through direct physical interaction. The study

of CRAC channels has been complicated by the fact that they lose activity in the absence of extracellular Ca2+. Attempts to maintain current size by removing intracellular Mg2+ have been found to unmask Mg2+-inhibited cation (MIC/MagNuM/TRPM7) channels, which have been mistaken in several studies for the CRAC channel. Recent studies under conditions that prevent MIC activation reveal that CRAC channels use high-affinity binding of Ca2+ in the pore to achieve high Ca2+ selectivity but have a surprisingly low conductance for both Ca2+ (approximately 10fS) and Na+ (approximately 0.2pS). Pore properties provide a unique fingerprint that provides a stringent test for potential CRAC channel genes and suggest models for the ion selectivity mechanism

Notes: Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Beckman Center B-111A, Stanford, CA 94305, USAFAU - Prakriya, Murali

18) Schmitt R, Weichert W. Immunostaining of small-cell carcinoma: mistaken identity?

Lancet 2003 Nov 8;362(9395):1583. Ref ID: 6127 19) van Pelt JF, Decorte R, Yap PS, Fevery J. Identification of HepG2 variant cell lines by

short tandem repeat (STR) analysis. Mol Cell Biochem 2003 Jan;243(1-2):49-54. Ref ID: 5926 Abstract: In the past years, in our laboratory, several cell lines have been generated starting from

a human liver (H7). Some of them have been used successfully in studies of the infection with and propagation of Hepatitis B and Hepatitis C viruses. Recently, several lines of evidence indicated that the origin of these cell lines was uncertain. Therefore, we now have determined the genetic characteristics of these cell lines in comparison to HepG2 cells received from ATCC and to HepG2 isolates grown at other laboratories. Quadruplex fluorescent short tandem repeat (STR) typing and karyotyping were performed. In addition, some biochemical characteristics of selected clones were studied. Genetically, all H7-derived cell lines were identical to HepG2 cells. However, some liver-specific functions varied between the different sub-cloned lines. The H7-derived cell lines that were generated proved to be sub-cloned lines of HepG2. The problem of cross-contamination during cloning of cell lines appears to be not uncommon. We found that two out of six HepG2 isolates obtained from other laboratories were not derived from the same individual as the original HepG2 cells. Therefore, STR typing should be applied as a rapid and sensitive technique to determine and monitor the origin of cell lines and to safeguard against contamination

Notes: Department of Liver and Pancreatic Diseases, University Hospital Gasthuisberg, Leuven, Belgium JosvanPelt@medkuleuvenacbeFAU - van Pelt, Jos F

20) Drexler HG, Quentmeier H, Dirks WG, Uphoff CC, MacLeod RA. DNA profiling and

cytogenetic analysis of cell line WSU-CLL reveal cross-contamination with cell line REH (pre B-ALL). Leukemia 2002 Sep;16(9):1868-70.

Ref ID: 5928 Abstract: a 21) Zhang Z, Harrison P, Gerstein M. Identification and analysis of over 2000 ribosomal

protein pseudogenes in the human genome. Genome Res 2002 Oct;12(10):1466-82. Ref ID: 6130 Abstract: Mammals have 79 ribosomal proteins (RP). Using a systematic procedure based on

sequence-homology, we have comprehensively identified pseudogenes of these proteins in the human genome. Our assignments are available at http://www.pseudogene.org or http://bioinfo.mbb.yale.edu/genome/pseudogene. In total, we found 2090 processed pseudogenes and 16 duplications of RP genes. In relation to the matching parent protein, each of the processed pseudogenes has an average relative sequence length of 97% and an average sequence identity of 76%. A small number (258) of them do not contain obvious disablements (stop codons or frameshifts) and, therefore, could be mistaken as functional genes, and 178 are

disrupted by one or more repetitive elements. On average, processed pseudogenes have a longer truncation at the 5' end than the 3' end, consistent with the target-primed-reverse-transcription (TPRT) mechanism. Interestingly, on chromosome 16, an RPL26 processed pseudogene was found in the intron region of a functional RPS2 gene. The large-scale distribution of RP pseudogenes throughout the genome appears to result, chiefly, from random insertions with the numbers on each chromosome, consequently, proportional to its size. In contrast to RP genes, the RP pseudogenes have the highest density in GC-intermediate regions (41%-46%) of the genome, with the density pattern being between that of LINEs and Alus. This can be explained by a negative selection theory as we observed that GC-rich RP pseudogenes decay faster in GC-poor regions. Also, we observed a correlation between the number of processed pseudogenes and the GC content of the associated functional gene, i.e., relatively GC-poor RPs have more processed pseudogenes. This ranges from 145 pseudogenes for RPL21 down to 3 pseudogenes for RPL14. We were able to date the RP pseudogenes based on their sequence divergence from present-day RP genes, finding an age distribution similar to that for Alus. The distribution is consistent with a decline in retrotransposition activity in the hominid lineage during the last 40 Myr. We discuss the implications for retrotransposon stability and genome dynamics based on these new findings

Notes: Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520, USAFAU - Zhang, Zhaolei

22) Drexler HG, MacLeod RA, Dirks WG. Cross-contamination: HS-Sultan is not a myeloma

but a Burkitt lymphoma cell line. Blood 2001 Dec 1;98(12):3495-6. Ref ID: 5929 Abstract: a 23) Luijt DS, Schirm J, Savelkoul PH, Hoekstra A. Risk of infection by reprocessed and

resterilized virus-contaminated catheters; an in-vitro study. Eur Heart J 2001 Mar;22(5):378-84.

Ref ID: 5095 Abstract: AIMS: In spite of increasing reuse of disposable catheters, there are few scientific data on

potential viral transmission and infection after reuse. To determine the theoretical risk of virus transmission during reuse of catheters an in vitro study was performed using an RNA virus (echovirus-11) and a DNA virus (adenovirus-2). METHODS AND RESULTS: After deliberate contamination of the catheters, reprocessing and reuse of the cleaned and glutaraldehyde sterilized catheters was simulated. The presence of residual virus was determined by cell culture and by polymerase chain reaction (PCR). After the sterilization step, infectious enterovirus was detectable in one (10%) of the samples, whereas two (20%) contained detectable enterovirus RNA. After simulated reuse, enterovirus was cultured from one (10%) of the catheters, but no less than six (60%) of the samples were enterovirus PCR positive and one (10%) contained detectable adenovirus DNA. After sonification of the catheter tips no infectious virus could be detected, but enterovirus RNA was detected in two (20%) and adenovirus DNA in three (30%) of the samples. CONCLUSIONS: It has been clearly demonstrated in this in vitro study that, even after rigorous cleaning and sterilization, virus was still present in the catheter. Reuse of catheters, labelled for single-use only, is dangerous and should be prevented. Copyright 2001 The European Society of Cardiology

Notes: Regional Public Health Laboratory, Groningen, The Netherlands 24) Meyer C, MacLeod RA, Quentmeier H, Janssen JW, Coignet LJ, Dyer MJ, et al.

Establishment of the B cell precursor acute lymphoblastic leukemia cell line MUTZ-5 carrying a (12:13) translocation. Leukemia 2001 Sep;15(9):1471-4.

Ref ID: 5931 Abstract: Continuous leukemia-lymphoma cell lines are important research tools, in particular as

starting material for the cloning of recurrent translocations. In 1998, we established the continuous leukemia cell line MUTZ-5 and its two simultaneous sister cell lines MUTZ-6 and

MUTZ-7. The primary specimen was obtained from the peripheral blood of a 26-year-old man with B cell precursor acute lymphoblastic leukemia at relapse carrying a t(12;13). The immunoprofile of MUTZ-5 corresponds to that of a precursor B cell. The immunoglobulin heavy chain gene was found to be rearranged. Despite receptor expression, none of the cytokines examined enhanced proliferation; several cytokines had significant inhibitory effects. Giemsa-banding cytogenetics showed the following karyotype which was identical in all three sister cell lines: 45<2n>X, -Y, t(12;13)(p12;q13-14). The karyotype and DNA fingerprinting confirmed the malignant nature and the authenticity of the cell line, excluding cross-contamination with other cells. MUTZ-5 represents a new unique leukemia B cell line; its scientific significance lies in the t(12;13)

Notes: DSMZ-German Collection of Microorganisms and Cell Cultures, Braunschweig, GermanyFAU - Meyer, C

25) Nelson-Rees WA. Responsibility for truth in research. Philos Trans R Soc Lond B Biol

Sci 2001 Jun;356(1410):849-51. Ref ID: 5140 Abstract: For over half a century, cell cultures derived from animals and humans have served

researchers in various fields. To this day, cross-contamination of cultures has plagued many researchers, often leading to mistaken results, retractions of results, cover-ups and some out-and-out falsification of data and results following inadvertent use of the wrong cells. Also, during years of examining cultures for purity we learned that many virologists were not too concerned about the specificity of the cultures they used to propagate the particular virus under study as long as the substrate (whatever it might have been) gave optimal virus yield. Polio virus propagates in primate cells, and much research has involved cells from man and various species of primates. In the 1950s a large number of chimpanzees were held in captivity in Africa for extensive studies of the efficacy of polio vaccine in production at the Wistar Institute in Philadelphia and elsewhere. Chimpanzee tissues, particularly kidneys, were thus readily available and could have also provided substrates for polio virus production, since little was known about the purity of substrates and little attention was paid to their specificity at that time

Notes: 999 Green Street, no 2302, San Francisco, CA 94133-5402, USA 26) Silva LM, Montes de OH, Diniz CR, Fortes-Dias CL. Fingerprinting of cell lines by

directed amplification of minisatellite-region DNA (DAMD). Braz J Med Biol Res 2001 Nov;34(11):1405-10.

Ref ID: 6131 Abstract: The development of in vitro propagation of cells has been an extraordinary technical

advance for several biological studies. The correct identification of the cell line used, however, is crucial, as a mistaken identity or the presence of another contaminating cell may lead to invalid and/or erroneous conclusions. We report here the application of a DNA fingerprinting procedure (directed amplification of minisatellite-region DNA), developed by Heath et al. [Nucleic Acids Research (1993) 21: 5782-5785], to the characterization of cell lines. Genomic DNA of cells in culture was extracted and amplified by PCR in the presence of VNTR core sequences, and the amplicons were separated by agarose gel electrophoresis. After image capture with a digital camera, the banding profiles obtained were analyzed using a software (AnaGel) specially developed for the storage and analysis of electrophoretic fingerprints. The fingerprints are useful for construction of a data base for identification of cell lines by comparison to reference profiles as well as comparison of similar lines from different sources and periodic follow-up of cells in culture

Notes: Centro de Pesquisa e Desenvolvimento, Fundacao Ezequiel Dias (FUNED), Rua Conde Pereira Carneiro, 80, 30510-010 Belo Horizonte, MG, BrazilFAU - Silva, L M

27) van BA, Varella-Garcia M, Korch C, Miller GJ. TSU-Pr1 and JCA-1 cells are derivatives

of T24 bladder carcinoma cells and are not of prostatic origin. Cancer Res 2001 Sep 1;61(17):6340-4.

Ref ID: 5930 Abstract: We have shown previously that the putative prostate carcinoma cell lines TSU-Pr1 and

JCA-1 share a common origin. The observation that these cell lines have p53 and Ha-ras mutations identical to those in bladder carcinoma cell line T24 prompted us to investigate their possible interrelations. We used cytogenetics and DNA profiling to compare the genetic backgrounds of the three cell lines. At least 12 structural chromosomal abnormalities are shared between T24, TSU-Pr1, and JCA-1 cells. DNA profiles were identical for all three cell lines. These results clearly indicate that the cell lines TSU-Pr1 and JCA-1 are not of prostatic origin but are derivatives of the bladder carcinoma cell line T24. TSU-Pr1 and, to a lesser extent, JCA-1 are frequently used as models in prostate cancer research, and numerous publications have appeared based on these lines. Several other T24 cross-contaminants have been identified in the past, and some of these, such as ECV304, continue to be used under the wrong identity. Our findings highlight the insidious problem that can occur when information regarding cross-contamination does not reach individual researchers and/or the importance of the problem is not fully acknowledged

Notes: Department of Pathology, University of Colorado Health Sciences Center, Denver, Colorado 80262, USA adrievanbokhoven@uchsceduFAU - van Bokhoven, A

28) Albert LJ, Inman RD. Gram-negative pathogens and molecular mimicry: is there a case

for mistaken identity? Trends Microbiol 2000 Oct;8(10):444-5. Ref ID: 6133 Notes: Divn of Rheumatology, Dept of Medicine, Toronto Western Hospital, University Health

Network, FP 1-221, 399 Bathurst St, ON, M5T 2S8, Toronto, CanadaFAU - Albert, L J 29) Bubenik J. Cross-contamination of cell lines in culture. Folia Biol (Praha)

2000;46(5):163-4. Ref ID: 5097 30) Falkner E, Frick W, Kapeller B, Eberl H, Macfelda K, Losert UM. [Concerning cell

cultures for biocompatibility-testing: monitoring by DNA-fingerprinting]. ALTEX 2000;17(3):135-7.

Ref ID: 5096 Abstract: Cell cultures are innovative tools for e.g. biocompatibility testing of biomaterials in vitro.

In our studies we used fibroblast, endothelial cell and chondrocyte cultures of human origin and of the test animal species most common for this purpose in vivo. Verification of the identity of these cells is obligatory for reproducibility of the tests and valid interpretation of the results. Cultured cells have to be checked for identity, contaminations of various origins and also for genomic mutations occuring during prolonged cultivation in vitro or due to exposition to biomaterials. Furthermore, the risk of genetic cross-contamination with other cells increases with the number of cell cultures passaged parallel in the same laboratory. Therefore, we generated reference fingerprints of the cultures in varying passages for comparative monitoring of cells purposed for in vitro tests. Minisattelite DNA polymorhism resulting in reproducible individual DNA fingerprints is very discriminatory and can be used for cell culture monitoring. The patterns are stable over several passages, although sudden changes did happen in two cases, i.e. loss/gain of bands or changes in band-intensity, indicating massive genomic mutations of the cultures in vitro. Influences of biomaterials on the prints could not be detected. Several tasks can be followed at the same time: detection of contaminant cells, identification of these cells of primary culture origin used for in vitro testing and finally, monitoring for eventual genomic mutations due to prolonged cultivation or contact to biomaterials. Inconclusive results in just one of these aspects should lead to the disqualification of the monitored cultures from usage in vitro

Notes: Institut fur Biomedizinische Forschung, Universitat A-Wien 31) Masters JR. Human cancer cell lines: fact and fantasy. Nat Rev Mol Cell Biol 2000

Dec;1(3):233-6. Ref ID: 5304 Abstract: Cancer cell lines are used in many biomedical research laboratories. Why, then, are they

often described as unrepresentative of the cells from which they were derived? Here, I argue that they have been unjustly accused. Under the right conditions, and with appropriate controls, properly authenticated cancer cell lines retain the properties of the cancers of origin.

Notes: Institute of Urology, University College London, 67 Riding House Street, London W1W 7EY, UK jmasters@uclacuk

32) Soloski MJ, Lo WF, Metcalf ES. Gram-negative pathogens and molecular mimicry: is

there a case for mistaken identity? Response. Trends Microbiol 2000 Oct;8(10):446-7. Ref ID: 6132 Notes: Dept of Medicine, Johns Hopkins School of Medicine, Baltimore, MD 21205, USAFAU -

Soloski, M J 33) Barnes MR, Russell RB, Copley RR, Ponting CP, Bork P, Cumberledge S, et al. A

lipid-binding domain in Wnt: a case of mistaken identity? Curr Biol 1999 Oct 7;9(19):R717-R719.

Ref ID: 6135 34) Dirks WG, MacLeod RA, Drexler HG. ECV304 (endothelial) is really T24 (bladder

carcinoma): cell line cross- contamination at source. In Vitro Cell Dev Biol Anim 1999 Nov;35(10):558-9.

Ref ID: 5934 Abstract: a 35) Do H, Healey JF, Waller EK, Lollar P. Expression of factor VIII by murine liver

sinusoidal endothelial cells. J Biol Chem 1999 Jul 9;274(28):19587-92. Ref ID: 5101 Abstract: Factor VIII (fVIII) is the procoagulant plasma glycoprotein that is missing or decreased in

hemophilia A. The cellular origin of fVIII synthesis is controversial. Liver transplantation cures hemophilia A, demonstrating that the liver is a major site of fVIII synthesis. We detected fVIII mRNA in purified populations of murine liver sinusoidal endothelial cells (LSECs) and hepatocytes, but not Kupffer cells. LSECs and hepatocytes contained comparable numbers of fVIII mRNA (40 and 70 transcripts per cell, respectively) by quantitative competitive reverse transcriptase-polymerase chain reaction analysis. There was not detectable mRNA for factor IX, a hepatocyte marker, in the LSEC preparation, nor was there detectable mRNA for von Willebrand factor, an endothelial cell marker, in the hepatocyte preparation. This excludes the possibility that detectable fVIII mRNA is due to cross-contamination in the hepatocyte or LSEC preparations. Primary cultures of LSECs were established in which fVIII mRNA levels were indistinguishable from purified LSECs. LSECs secreted active fVIII into the culture medium. This finding represents the first demonstration of homologous expression of fVIII mRNA and protein in cell culture and should facilitate studies of fVIII gene regulation. Additionally, LSECs potentially are targets for a fVIII transgene during gene therapy of hemophilia A

Notes: Division of Hematology-Oncology, Department of Medicine, Emory University, Atlanta, Georgia 30322, USA

36) Drexler HG, Dirks WG, MacLeod RAF. False human hematopoietic cell lines:

cross-contaminations and misinterpretations. Leukemia 1999;13:1601-7. Ref ID: 5014 Notes: The risk of adventitious contamination and subsequent overgrowth of cell lines by unrelated

cells is a potential and often recurring problem where cells are grown and studied. This problem of intraspecies and interspecies cross-contamination among human cell lines has been

redognized for over 25 years; incidences of cell cross-contamination between 17 and 35% have been reported. The most useful methods to detect human cell cross-contamination are DNA fingerprinting and cytogenetic analysis, each complementing the other. Using this combination, we found that in total 14.8% of the human hematopoietic cell lines received either from the original investigator (n = 117 cetl lines) or from secondary sources (n = 72 cell lines) were cross-contaminated with another hematopoietic cell line and were thus false cell cultures. Another problem relates to the fact that not every cell line established from a patient with a hematopoietic malignancy is a malignant cell line; unintended immortalization of non-malignant B cells by 'passenger' Epstein-Barr virus (EBV) Ieads to the establishment of B-lymphoblastoid cell lines (termed EBV+ B-LCLs), an event which is much more frequent than the establishment of a 'true' Ieukemia-lymphoma-myeloma cell line. These EBV+ B-LCLs are most often (albeit not always) unrelated to the malignant clone. The misinterpretation of such EBV+ B-LCLs as true malignant hematopoietic cell lines (particularly in research areas investigating B cell-derived neoplasms such as myeloma) and the indiscriminate use of these cell lines may render some of the results of such studies irrelevant to the pathobiology of the disease concerned. However, a combination of markers commonly allows for an accurate determination of the nature of EBV+ B-LCLS: immunoprofile, cellular morphology, EBV status, and karyotype. In summary, the continuous need for vigilant quality and identity control procedures is emphasized by the high incidences of cross-contaminated cell lines. Most laboratories using cells cultured in vitro maintain multiple cell lines. Such cell lines should be monitored regularly for their identity and specific characteristics in order to prevent invalidation of research work due to incidents of cell line cross-contamination or misinterpretation.

Keywords: Ieukemia; Iymphoma; myeloma; cell lines; authentication. 37) Eisen JS. A case of mistaken identity. Neuron 1999 Aug;23(4):626-7. Ref ID: 6136 Notes: Institute of Neuroscience, University of Oregon, Eugene 97403, USAFAU - Eisen, J S 38) Jayme DW. An animal origin perspective of common constituents of serum-free medium

formulations. Dev Biol Stand 1999;99:181-7.:181-7. Ref ID: 5100 Abstract: Serum-free formulations may be << re-engineered >> to eliminate traditional protein

constituents and replace their biological function with non-protein substitutes. Non-protein additives may also be obtained from animal sources. Nutrient formulations totally free of exogenous protein and containing no materials of animal origin may be designed for high density cell culture and biological production. Cell-culture medium production requires (i) strict vendor qualification and raw material specifications; (ii) scrupulous maintenance of media kitchen facility and equipment; (iii) monitoring of process water; (iv) air-handling systems and technical personnel; (v) clearly-defined manufacturing protocols to ensure correct formulation and dispensing and (vi) validated sanitization processes to guard against cross-contamination within a multi-use facility

Notes: Industrial Cell Culture Applications, Life Technologies, Inc, Grand Island, NY 14072, USA 39) MacLeod RA, Dirks WG, Matsuo Y, Kaufmann M, Milch H, Drexler HG. Widespread

intraspecies cross-contamination of human tumor cell lines arising at source. Int J Cancer 1999 Nov 12;83(4):555-63.

Ref ID: 5936 Abstract: We present a panoptic survey of cell line cross-contamination (CLCC) among original

stocks of human cell lines, investigated using molecular genetic methods. The survey comprised 252 consecutive human cell lines, almost exclusively tumor-derived, submitted by their originators to the DSMZ and 5 additional cell repositories (CRs), using a combination of DNA profiling (4-locus minisatellite and multilocus microsatellite probes) and molecular cytogenetics, exploiting an interactive database (http://www.dsmz.de/). Widespread high levels

of cross-contaminants (CCs) were uncovered, affecting 45 cell lines (18%) supplied by 27 of 93 originators (29%). Unlike previous reports, most CCs (42/45) occurred intraspecies, a discrepancy attributable to improved detection of the more insidious intraspecies CCs afforded by molecular methods. The most prolific CCs were classic tumor cell lines, the numbers of CCs they caused being as follows: HeLa (n = 11), T-24 (n = 4), SK-HEP-1 (n = 4), U-937 (n = 4) and HT-29 (n = 3). All 5 supposed instances of spontaneous immortalization of normal cells were spurious, due to CLCC, including ECV304, the most cited human endothelial cell line. Although high, our figure for CCs at the source sets a lower limit only as (i) many older tumor cell lines were unavailable for comparison and (ii) circulating cell lines are often obtained indirectly, rather than via originators or CRs. The misidentified cell lines reported here have already been unwittingly used in several hundreds of potentially misleading reports, including use as inappropriate tumor models and subclones masquerading as independent replicates. We believe these findings indicate a grave and chronic problem demanding radical measures, to include extra controls over cell line authentication, provenance and availability

Notes: DSMZ-German Collection of Microorganisms and Cell Cultures, Braunschweig, Germany rml@dsmzdeFAU - MacLeod, R A

40) MacLeod RAF, Dirks WG, Matsuo Y, Kaufmann M, Milch H, Drexler HG. Widespread

intraspecies cross-contamination of human tumor cell lines arising at source. Int J Cancer 1999;In print.

Ref ID: 5015 Notes: We present the first panoptic survey of cell line cross contamination (CLCC) among original

stocks of human cell lines investigated using molecular genetic methods. The survey comprized 252 consecutive human cell lines, almost exclusively tumor-derived, submitted by their originators to the DSMZ and flve additional cell repositories (CR), using a combination of DNA-proflling (four-locus minisatellite-, and multilocus microsatelllte probes) and molecular cytogenetlcs, exploiting an interactive database (available at: http://www.dsmz.de/). ¥Videspread, high levels of cross-contaminants (CC) were uncovered, affecting 45 cell lines (18%) as supplied by 27 out of 93 originators (29%). Unlike previous reports, most CC (42/45) occurred intraspecies, a discrepancy attributable to improved detection of the more insidious intraspecies CC afforded by molecular methods. The most prolific CC were classic tumor cell lines, the numbers of CC they caused being as follows: HELA (n=11), T-24 (n=4), SK-HEP-1 (n=4), U-937 (n=4), and HT-29 (n=3). All flve supposed instances of spontaneous immortalization of normal cells were spurious, due to CLCC; including ECV304, the most cited human endothelial cell line. Although high, our flgure for CC at source sets a lower limit only as: 1) many older tumor cell lines were unavailable for comparison; and 2) circulating cell lines are often obtained indirectly, rather than via originators or CR. The misidentifled cell lines reported here have already been unwittingly used in several hundreds of potentially misleading reports, including use as inappropriate tumor models and subclones masquerading as independent replicates. We believe these flndings indicate a grave and chronic problem demanding radical measures, to include extra controls over cell line authentication, provenance and availability.

41) Matsuo Y, Nishizaki C, Drexler HG. Efficient DNA fingerprinting method for the

identification of cross-culture contamination of cell lines. Hum Cell 1999 Sep;12(3):149-54. Ref ID: 5933 Abstract: In order to identify cross-culture contamination of cell lines, we applied DNA

fingerprinting using variable number of tandem repeat (VNTR) loci and short tandem repeat (STR) loci amplified by polymerase chain reaction (PCR) instead of a radioisotope labeled multilocus probe. Eleven cell lines were used for the Apo B and D1S80 loci detection, and twelve cell lines were examined in the Y-chromosome analysis. The data obtained from the sister cell lines NALM-6 and B85, two MOLM-1 cultures from two cryopreserved tubes, and four subclones of BALM-9 and its sister cell line BALM-10, displayed clear and distinct bands of each PCR product for both Apo B and D1S80. Detection of a Y-chromosome DNA sequence

is another very informative marker for the identification of cell lines, if the Y-chromosome is present. We examined eight cell lines for the expression of four STR loci; the data thus generated were compared with the results previously reported from other laboratories. The resulting electrophoretic banding patterns showed that our "home-made" STR detection system is a useful and efficient tool for the authentication of cell lines. PCR detection of VNTR and STR loci represents a simple, rapid and powerful DNA fingerprinting technique to authenticate human cell lines and to detect cross-culture contamination. This PCR technique may be used in lieu of the more time-consuming, labor-intensive and radioactive Southern blot multilocus method

Notes: Fujisaki Cell Center, Hayashibara Biochemical Labs, Inc, Okayama, Japan yomatsuo@hayashibaracojpFAU - Matsuo, Y

42) Muir P, Ras A, Klapper PE, Cleator GM, Korn K, Aepinus C, et al. Multicenter quality

assessment of PCR methods for detection of enteroviruses. J Clin Microbiol 1999 May;37(5):1409-14.

Ref ID: 5102 Abstract: We conducted a multicenter evaluation of commercial and in-house PCR methods for the

detection of enteroviruses. Three coded panels of test and control RNA samples, artificial clinical specimens, and representative enterovirus serotypes were used to assess amplification methods, RNA extraction methods, and reactivities with different enterovirus serotypes. Despite several differences between PCR methods, there was good agreement, although some variation in sensitivity was observed. Most PCR methods were able to detect enterovirus RNA derived from 0.01 50% tissue culture infective dose (TCID50) and were able to detect at least 1 TCID50 of enterovirus in cerebrospinal fluid, stool, or throat swab specimens. Most were also able to detect a wide range of enterovirus serotypes, although serotypic identification was not possible. Some laboratories experienced false-positive results due to PCR contamination, which appeared to result mainly from cross-contamination of specimens during RNA extraction. Provided that this problem is overcome, these PCR methods will prove to be a sensitive and rapid alternative to cell culture for the diagnosis of enterovirus infection

Notes: Department of Virology, Guy's, King's College & St Thomas' Hospitals' School of Medicine, London, United Kingdom pmuir@umdsacuk

43) Tumman J, Coggins R. A case of mistaken identity: primary cutaneous lymphoma

presenting as venous ulceration. Hosp Med 1999 Oct;60(10):761. Ref ID: 6134 Notes: Department of Surgery, Trafford General Hospital, ManchesterFAU - Tumman, J 44) Hay RJ. Cell line availability: where to get the cell lines you need. Methods Cell Biol

1998;57:31-47. Ref ID: 5105 Abstract: The availability and utility of cell lines with limited and continuous doubling potential are

summarized and documented. Reference to national cell banks is included with pertinent and current contact information. The continuing need for vigorous application of quality control procedures is emphasized with data illustrating frequencies of microbial infection in cultured cell lines as well as high incidences of cross-contamination of one cell line with another

Notes: American Type Culture Collection, Cell Culture Department, Rockville, Maryland 20852, USA

45) Kaplan J, Hukku B. Cell line characterization and authentication. Methods Cell Biol

1998;57:203-16.:203-16. Ref ID: 6137 Abstract: Research and development involving the use of cell lines require precise knowledge of

the purity and species of origin of the cell lines used. This can only be assured by periodic monitoring of cultured cell lines for possible contamination by other cells and for

characteristics that authenticate the cell line identity. In the absence of such monitoring, inter- and intraspecies cell line contaminations are likely to occur in the laboratories of unsuspecting investigators and can result in the generation of mistaken conclusions with an attendant loss of investigators' time, effort, and resources. This chapter provides a history and an overview of the methods that have been developed for cell line authentication, the type of information each of these different methods provides, and how synthesis of that information can be used to characterize a cell line and confirm its identity. An effective cell line monitoring strategy is described that involves testing for a combination of genetic markers, including cell membrane species antigens, isoenzymes, chromosomes, and DNA fingerprints, and use of databases for each marker system to compare the results obtained with a test cell culture with results from an extensive panel of previously tested cell lines

Notes: Children's Hospital of Michigan, Department of Pediatrics, Wayne State University School of Medicine, Detroit 48201, USAFAU - Kaplan, J

46) Lincoln CK, Gabridge MG. Cell culture contamination: sources, consequences,

prevention, and elimination. Methods Cell Biol 1998;57:49-65.:49-65. Ref ID: 5104 Abstract: The subject of the chapter is cell culture contamination. Contamination may enter the cell

culture system as a physical, chemical, and/or biological component of the environment. The potential sources and consequences of cell culture contamination are unique to the cell culture system and the contaminant. A basic understanding of cell culture contamination is necessary to appreciate the need to develop and practice standardized cell culture procedures. General sources, consequences, and preventative measures are discussed for physical and chemical contamination based on current technology. Mycoplasmal contamination is the focus of the discussion on biological contamination and its impact on cell cultures. The introduction of other biological contaminants should be controlled by the institution of cell culture management procedures needed to minimize the incidence of mycoplasmal contamination. The need to eliminate the routine use of antibiotics in cell culture systems and institute routine testing to detect contamination is emphasized. More rapid detection of contamination should reduce the incidence of cross-contamination and minimize the consequences of any contamination event

Notes: Bionique Testing Laboratories, Inc, Saranac Lake, New York 12983, USA 47) Markovic O, Markovic N. Cell cross-contamination in cell cultures: the silent and

neglected danger. In Vitro Cell Dev Biol Anim 1998 Jan;34(1):1-8. Ref ID: 5107 Abstract: Cell cross-contamination in cell cultures is a common problem during cell culturing and

use. Contamination invalidates research results, compromises the comparison of results between laboratories, reduces reproducibility required in industrial production of cell lines, and may lead to unusable therapeutic products. The problem can be solved by increasing the awareness of its seriousness and by introducing regular quality control of cell cross-contamination in every laboratory where cells are grown and used

Notes: BioSciCon, Inc, Rockville, MD 20852, USA 48) Nims RW, Shoemaker AP, Bauernschub MA, Rec LJ, Harbell JW. Sensitivity of

isoenzyme analysis for the detection of interspecies cell line cross-contamination. In Vitro Cell Dev Biol Anim 1998 Jan;34(1):35-9.

Ref ID: 5106 Abstract: The analysis of the gel electrophoresis banding patterns and relative migration distances

for the individual isoforms of intracellular enzymes, such as lactate dehydrogenase, purine nucleoside phosphorylase, glucose-6-phosphate dehydrogenase, and malate dehydrogenase, is used routinely in the biopharmaceutical industry for confirmation of cell line species of origin. In the present study, the sensitivity of the technique (AuthentiKit, Innovative Chemistry, Marshfield, MA) for determining interspecies cell line cross-contamination was examined.

Extracts were prepared from a CHO-K1 line (AA8, Chinese hamster), MRC-5 (human) cells, and L929 (mouse) cells and from several proportional mixtures of the various binary combinations of cells. The isoenzymes were analyzed according to standard procedures for the technique. Contamination of MRC-5 cells with CHO-K1 or with L929 cells was clearly detectable with each enzyme analyzed. Similarly, the contamination of L929 or CHO-K1 cells with MRC-5 cells was readily apparent with each enzyme. On the other hand, contamination of CHO-K1 cells with L929 cells was only detected with lactate dehydrogenase analysis, and contamination of L929 cells with CHO-K1 cells was not detected with any of the four enzymes examined. For the latter case, the analysis of an additional enzyme (peptidase B) was required. The results indicate that interspecies cross-contamination should be detectable with isoenzyme analysis if the contaminating cells represent at least 10% of the total cell population

Notes: MA BioServices, Inc, Rockville, Maryland 20850, USA 49) Price JE, Wolf JK, Pathak S. Distinctive karyotypes and growth patterns in nude mice

reveal cross-contamination in an established human cancer cell line. Oncol Rep 1998 Jan;5(1):261-6.

Ref ID: 5108 Abstract: A human cancer cell line was found to be heterogeneous for expression of the epidermal

growth factor receptor (EGFR). Clones and variants of this cell line were separated on the basis of EGFR expression level, and those expressing high EGFR had different growth characteristics, in vitro and in vivo, than variants expressing low levels of EGFR. Karyotype analysis revealed that the heterogeneity was the result of mixing of two lines, the 2774 ovarian cancer cell line, and the SW620 colon cancer cell line. Our results reinforce the necessity for accurate identification of cell lines. Also, that measurement of gene expression on a single cell level, for example by flow cytometric analysis, can be more informative than measurements of cell lysates, since the initial indication of heterogeneity would not have been detected by northern or western blotting. The different cell types retained characteristic growth patterns when injected i.p. in nude mice, i.e. peritoneal carcinomatosis and ascites formation by the 2774 ovarian cancer cells, and liver metastasis and growth of discrete abdominal tumors by the SW620 colon cancer

Notes: Department of Cell Biology, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA

50) Wagner SJ, Robinette D. Evaluation of an automated microbiologic blood culture device

for detection of bacteria in platelet components. Transfusion 1998 Jul;38(7):674-9. Ref ID: 5103 Abstract: BACKGROUND: Automated culture methods have been used by several investigators to

detect bacterial contamination of cellular blood components. We investigated several factors affecting detection by automated culture of bacteria in platelet concentrates (PCs).These factors included the initial contamination level in PCs, the PC sample volume, the PC sample time, and the white cell level in relation to bacteria levels in the PCs. STUDY DESIGN AND METHODS: Staphylococcus epidermidis or Escherichia coli was inoculated into freshly prepared PCs or white cell-reduced PCs to yield colony-forming unit (CFU) levels of 10, 1, or 0.1 per mL. At the time of inoculation (t=0) and at t=6, t=24, and t=48 hours, 0.5, 1.0, and 2.0 mL samples of the contaminated PCs were transferred into culture bottles. The presence of bacteria in the culture bottles was subsequently monitored by an automated blood culturing instrument. Bacteria levels in the PC at the time of first automated culture detection were determined by quantitative plating. RESULTS: E. coli was detected in 92 percent of experiments when 1.0- or 2.0-mL samples were taken at t=6 hours. At t=24 hours, 100-percent detection was observed with all tested inoculation volumes; however, by that time, >10(7) CFU per mL of bacteria were present in every PC. For S. epidermidis, 89 percent and 83 percent of contaminated PCs were detected with a t=24 hour sampling time and 2.0- or 1.0-mL sampling volume. Seven of 36 PCs with a 2.0-mL sampling volume and 10 of 36 PCs with a 1.0-mL sampling volume contained >10(6) CFU per mL of S. epidermidis at the time of first detection.

CONCLUSION: Data from this preliminary evaluation suggest that sampling times of 24 hours or more would be necessary to provide confidence in detection of E. coli or S. epidermidis in PCs using this culture method

Notes: Jerome H Holland Laboratory for the Biomedical Sciences, American Red Cross Biomedical Services, Rockville, MD 20855, USA

51) Boyce JM, Potter-Bynoe G, Chenevert C, King T. Environmental contamination due to

methicillin-resistant Staphylococcus aureus: possible infection control implications. Infect Control Hosp Epidemiol 1997 Sep;18(9):622-7.

Ref ID: 5110 Abstract: OBJECTIVE: To study the possible role of contaminated environmental surfaces as a

reservoir of methicillin-resistant Staphylococcus aureus (MRSA) in hospitals. DESIGN: A prospective culture survey of inanimate objects in the rooms of patients with MRSA. SETTING: A 200-bed university-affiliated teaching hospital. PATIENTS: Thirty-eight consecutive patients colonized or infected with MRSA. Patients represented endemic MRSA cases. RESULTS: Ninety-six (27%) of 350 surfaces sampled in the rooms of affected patients were contaminated with MRSA. When patients had MRSA in a wound or urine, 36% of surfaces were contaminated. In contrast, when MRSA was isolated from other body sites, only 6% of surfaces were contaminated (odds ratio, 8.8; 95% confidence interval, 3.7-25.5; P < .0001). Environmental contamination occurred in the rooms of 73% of infected patients and 69% of colonized patients. Frequently contaminated objects included the floor, bed linens, the patient's gown, overbed tables, and blood pressure cuffs. Sixty-five percent of nurses who had performed morning patient-care activities on patients with MRSA in a wound or urine contaminated their nursing uniforms or gowns with MRSA. Forty-two percent of personnel who had no direct contact with such patients, but had touched contaminated surfaces, contaminated their gloves with MRSA. CONCLUSIONS: We concluded that inanimate surfaces near affected patients commonly become contaminated with MRSA and that the frequency of contamination is affected by the body site at which patients are colonized or infected. That personnel may contaminate their gloves (or possibly their hands) by touching such surfaces suggests that contaminated environmental surfaces may serve as a reservoir of MRSA in hospitals

Notes: Division of Infectious Diseases, Miriam Hospital, Providence, RI 02906, USA 52) Kshirsagar SG, Patole MS, Shouche YS. Characterization of insect cell lines:

heteroduplex analysis employing a mitochondrial 16S ribosomal RNA gene fragment. Anal Biochem 1997 Nov 1;253(1):65-9.

Ref ID: 5109 Abstract: Routine cell line characterization procedures are not adequate for characterizing the cell

lines of insect origin. Ribosomal RNA (rRNA) gene sequences and their comparisons have been used successfully for delineating species and phylogenetic analysis. Using similar principles, we have standardized a protocol for the confirmation of species identity of insect cell lines. The procedure includes PCR amplification of the mitochondrial 16S rRNA gene fragment from the cell line and larvae of known insect species and heteroduplex analysis to detect the sequence variation in the PCR-amplified rRNA gene fragments. If the PCR fragment of the cell lines yields a homoduplex with the larvae of known species, then the cell line is conspecific with the larvae. If the larvae and cell line are of two different species, then the analysis exhibits multiple bands of heteroduplexes. The technique also allows detection of cross-contamination of culture having two insect cell lines belonging to two different species

Notes: National Center for Cell Science, University of Pune Campus, Pune, Ganeshkhind, 411007, India

53) Gierow JP, Yang T, Bekmezian A, Liu N, Norian JM, Kim SA, et al. Na-K-ATPase in

lacrimal gland acinar cell endosomal system: correcting a case of mistaken identity. Am J Physiol 1996 Nov;271(5 Pt 1):C1685-C1698.

Ref ID: 6138 Abstract: Na-K-ATPase is associated with a variety of membrane populations in lacrimal acinar

cells. Acinus-like structures formed by rabbit acinar cells in primary culture were incubated with horseradish peroxidase (HRP) to label basolateral and endosomal membranes and then analyzed by electron microscopy cytochemistry with the 3-3'-diaminobenzidine reaction or by fractionation and measurement of marker catalytic activities or immunoreactivities. HRP adsorbed to basolateral membranes at 4 degrees C. Fractionation showed it associated with low-density membranes enriched in acid phosphatase and TGN38 but containing only minor amounts of Na-K-ATPase. Cells internalized HRP to cytoplasmic vesicles, Golgi structures, and lysosomes at 37 degrees C. The major endosomal compartment revealed by fractionation coincided with major peaks of Na-K-ATPase and Rab6 and secondary peaks of galactosyltransferase and gamma-adaptin. Carbachol (10 microM) increased lysosomal and Golgi labeling. Thus most of the Na-K-ATPase is located in the basolateral membrane-oriented endosomal system, concentrated in a compartment possibly related to the trans-Golgi network. Constitutive and stimulation-accelerated traffic to and from this compartment may serve several exocrine cell functions

Notes: Department of Physiology and Biophysics, University of Southern California School of Medicine, Los Angeles 90033, USAFAU - Gierow, J P

54) Honma M, Mizusawa H, Hayashi M, Kaoru S, Ohno Y, Sofuni T. Heterogeneity of the Y

chromosome following long-term culture of the human lung cancer cell line A549. In Vitro Cell Dev Biol 1996;Animal 32(May):262-4.

Ref ID: 4778 Notes: A549, a human lung cancer cell line, has been widely used in various kinds of studies. It was

initiated in 1972 by D.J.Giard et al. through explant culture of lung carcinomatous tissue from a 58 yr-old Caucasian male, and the culture in passage 68 was deposited by M.Lieber into American Type Culture Collection (ATCC) in 1976. Two Japanese cell banks, JCRB and RCB obtained the cell line from ATCC, and have maintained and distributed it to many Japanese researchers. Karyotype analysis, which is one of the routine tests used to guarantee the identity and quality of cell lines deposited in both Japanese cell banks, revealed that the A549 cell line maintained in RCB (RCB0098) did not have Y chromosome, even though it was derived from a male donor. We first suspected contamination by another cell cuture; cross-culture contamination leading to the replacement of one cell type with another has become a serious problem as the number of cell lines maintained in laboratories ahs increased. Loss of the Y chromosome during cell culture was another possibility, because karyotypes of established cell lines are genrerally unstable. We, therefore, compared the A549 cell sublines maintained in these three cell repositories at molecular and cytogenetic level to clarify this trouble.

55) Buehring GC, Valesco M, Pan CY. Cell culture contamination by mycobacteria. In Vitro

Cell Dev Biol Anim 1995 Nov;31(10):735-7. Ref ID: 6111 56) Iurkov SG. [Cross contamination of continuous cell cultures]. Vopr Virusol 1995

Sep;40(5):225-7. Ref ID: 5111 Abstract: The possibility of air-droplet cell transmission during cell transfer was experimentally

confirmed, this appearing to be the most probable route of cross contamination of cell cultures. The consequences of such contamination are assessed and measures reducing the risk of intercellular contamination proposed

57) Morita T, Shinohara N, Honma M, Tokue A. Establishment and characterization of a new

cell line from human bladder cancer (JMSU1). Urol Res 1995;23(3):143-9. Ref ID: 5658

Abstract: A new human bladder cancer cell line designated JMSU1 has been established from malignant ascitic fluid of a 75-year-old Japanese man with bladder cancer, and maintained in culture for more than 7 years and over 240 passages. Inverted phase-contrast microscopy revealed that JMSU1 was composed of morphologically distinct cells (polygonal to spindle-shaped cells), showing morphological heterogeneity in vitro. Histological examination of xenografts showed poorly differentiated transitional cell carcinoma, resembling the original tumor. Immunohistochemical staining for cytokeratin and electron microscopic examination suggested that JMSU1 was of epithelial origin. Chromosome analysis gave a modal number of 69 with no Y chromosome. Isozyme analysis (LDH, G6PD, and NP) showed the mobility pattern of human type B. DNA fingerprint analysis demonstrated that there was no cross-culture contamination of JMSU1 during the passages. In conclusion, a newly established and well-characterized cell line, JMSU1, offers promising material for the investigation of the biological properties of bladder cancer

Notes: Department of Urology, Jichi Medical School, Tochigi, JapanFAU - Morita, T 58) Loew ER. A third, ultraviolet-sensitive, visual pigment in the Tokay gecko (Gekko

gekko). Vision Res 1994 Jun;34(11):1427-31. Ref ID: 6139 Abstract: Numerous extraction and microspectrophotometric studies have shown that the nocturnal

Tokay gecko (Gekko gekko), has two visual pigments: a "green" with lambda max at 521 nm and a "blue" at 467 nm. In addition, similar studies on other nocturnal gecko species have found only the same two classes of visual pigment. With the finding that some diurnal species of gecko have a third visual pigment class with lambda max peaking in the UV, doubts were raised concerning the presence of only two visual pigment classes in nocturnal forms. Therefore, a microspectrophotometric re-examination of the Tokay gecko was undertaken to look specifically for a UV visual pigment. A UV-absorbing pigment (364 nm lambda max) was found in approx. 20% of the thin outer segments of type C double rods, thought previously to contain only the 467 nm pigment. That this UV-absorbing pigment was truly a visual pigment was confirmed by its dichroism, behaviour following exposure to UV radiation and "nomogram" fit. It is suggested that this visual pigment had been seen in previous microspectrophotometric studies, but its similarity to known photoproducts peaking in the same spectral region resulted in a case of mistaken identity

Notes: Department of Physiology, Cornell University, Ithaca, NY 14853FAU - Loew, E R 59) Gignac SM, Steube K, Schleithoff L, Janssen JW, MacLeod RA, Quentmeier H, et al.

Multiparameter approach in the identification of cross-contaminated leukemia cell lines. Leuk Lymphoma 1993 Jul;10(4-5):359-68.

Ref ID: 5112 Abstract: A common problem in cell culturing is cross-contamination with other cells or

misidentification of cells. An effective cell culture quality and identity control is required in order to avoid inter- and intraspecies contamination of cell lines and their further propagation and dissemination. We present evidence that supposedly unrelated cell lines that we received from the original investigators are in fact related to the chronic myeloid leukemia cell line K-562. The sister cell lines SPI-801 and SPI-802 were originally established from a patient with T-cell acute lymphoblastic leukemia and displayed T-cell associated features. However, data from morphological evaluation, immunophenotyping, bcr-abl gene rearrangement analysis, DNA fingerprinting, Northern blot analysis of globin gene expression and esterase isoenzyme analysis clearly established that the three cell lines are related. Cytogenetic examination while not proving the common identity of the cells provided further evidence for the suspected common origin of all three cell lines. Chromosome banding, DNA fingerprinting and bcr-abl genotyping suggested further evolution of these clones during long-term cultivation. Quality and identity control is an essential feature of cell culture technique. Only regular monitoring for purity and integrity of cell lines will significantly reduce the incidence of cell line contamination and misidentification

Notes: German Collection of Microorganisms and Cell Cultures, Department of Human and Animal Cell Cultures Braunschweig

60) Honma M, Stacey G, Mizusawa H. DNA profiling with polymorphic DNA markers. In:

Doyle A, Griffiths JB, Newell DG, editors. Cell & Tissue Culture: Laboratory Procedures. 1 ed. Chichester: John Wiley & Sons; 1993. p. 9A:5.1-9A:5.13.

Ref ID: 3704 Notes: Very details of the experimental procedures for DNA profiling are described. 61) Johnson A, Olofsson T. Flow cytometric clonal excess analysis of peripheral blood,

routine handling, and pitfalls in interpretation. Cytometry 1993;14(2):188-95. Ref ID: 6140 Abstract: Clonal excess (CE) analysis by flow cytometry is a convenient method to detect minimal

involvement of peripheral blood and bone marrow by B-cell lymphoma. The method is based on evaluation of the congruity between kappa and lambda light chain distributions within a normal B-cell population. By using the Kolmogorov-Smirnov method for evaluation of histogram identity, the maximum difference (D value) between distributions is calculated. However, variable adsorption of cytophilic plasma immunoglobulin (Ig) to CD16 positive cells, T-cell subsets, and the B cells themselves may cause incongruity between the light chain distributions that might simulate or disguise a true clonal excess and thus create major pitfalls in the interpretation of the CE analysis. These phenomena are observed both in normal blood donors and in patients with non-Hodgkin's lymphoma. In the present report we describe: (1) how freezing of the isolated mononuclear cells before immunostaining effectively removes most of the adsorbed cytophilic Ig, (2) how by exclusion of the CD16 positive population further resolution of the relevant histogram shape is obtained to avoid false interpretations of incongruity, and (3) how adsorption of Ig to B-cells creates a typical pattern of "reciprocal labeling" that can be mistaken for a clonal excess. Based on our observations we argue against an uncritical use of normal D values for definition of clonal excess and advocate an analytic evaluation of the kappa and lambda overlay distribution by dual immunofluorescence, either by excluding CD16 positive cells or by including only B cells, to reveal the nature of the different deviations

Notes: Department of Oncology, University Hospital of Lund, SwedenFAU - Johnson, A 62) Mizusawa H, Honma M, Nomura N. [Precaution of a cross-culture contamination and

DNA profiling system]. [Soshiki Baiyo] 1993;19:494-8. Ref ID: 3850 Notes: [Language in Japanese] Technical information for DNA profiling system and cross-culture

contamination. 63) Satoh M, Takeuchi M. Cross-contamination of cell lines as revealed by DNA

fingerprinting in the IFO animal cell bank. IFO Res Commun 1993;16:18-23. Ref ID: 3559 Notes: For quality control of cell lines, the institue for fermentation, Osaka (IFO) animal cell bank

recently introduced DNA fingerprinting analysis, which eenables verification of cell lines at the indivicual level, to detect cross-culture contamination. By using this analysis, we found two ccases of cross contamination of cell lines.

64) Embleton MJ, Gorochov G, Jones PT, Winter G. In-cell PCR from mRNA: amplifying

and linking the rearranged immunoglobulin heavy and light chain V-genes within single cells. Nucleic Acids Res 1992;20:3831-7.

Ref ID: 3566 Notes: We describe a process for the identification of mRNAs within single cells, as demonstrated

with the immunoglobulin (Ig) variable region (V) genes of two mouse hybridoma cell lines and the bcr-abl fusion gene of the human K562 myeloid leukaemia line. The cells were fixed and

permeabilised, the mRNA reverse transcribed to cDNA and the cDNA amplified by the polymerase chain reaction (PCR). After using fluorescent PCR primers, the amplified DNA could be detected within the cells as demonstrated by confocal fluorescence microscopy and flow cytometry. Furthermore the amplified Ig VH and VL DNA could be assembled within the same cell using suitable PCR primers. We detected no cross-contamination of amplified DNA between cells: the DNA isolated from mixtures of two hybridoma cell lines (B1-8 and NQ10/12.5) treated to in-cell PCR and assembly, was shown by cloning to correspond to the combinations of VH and VL genes of the parent hybridomas. We forsee diverse applications of in-cell assembly by PCR, especially for the analysis of the combinations of chains of rearranged Ig or T cell receptor (TCR) V-genes in a population of cells, and the construction of human antibodies from the V-genes of immune B-lymphocytes

65) Hampe J, Nurnberg P, Epplen C, Jahn S, Grunow R, Epplen JT. Oligonucleotide

fingerprinting as a means to identify and survey long-term cultured B cell hybridomas and T cell lines. Hum Antibodies Hybridomas 1992 Oct;3(4):186-90.

Ref ID: 5113 Abstract: Common problems encountered during cell culture are cross-contamination, instability,

and inadvertent exchange of cells. Here we report on the application of oligonucleotide fingerprinting as a simple and efficient method to screen hybridomas and T cell lines. Among the fingerprint probes tested, the simple repetitive oligonucleotide (CAC)5/(GTG)5 proved to be most useful for obtaining many fragments specific for each cell line. Because of variable loss of chromosomes, cloned hybridoma cells from one fusion exhibit different fingerprint patterns. Thus, antibody-secreting B cell hybridomas can be distinguished easily even when they originate from the same fusion. Furthermore, we were able to monitor the genomic integrity of myelomas and hybridomas over a period of more than 2 years, thereby highlighting long-term stability. Another application of the method is the control of T cell lines requiring irradiated or mitomycin-treated feeder cells for continuous growth or cloning. There cell lines are always threatened to be overgrown by feeder cells that have escaped from the lethal pretreatment. T cell clones of one individual, known to display differently rearranged T cell receptors, did not show differences in their fingerprint patterns. However, in EBV-transformed cloned B cells, slight differences between clones from the same donor were identified

Notes: Institute for Medical Immunology, Faculty of Medicine (Charite), Humboldt University, Berlin, Germany

66) Hay RJ. ATCC quality control methods for cell lines (2nd Ed.). 2 ed. Rockville: American

Type Culture Collection; 1992. Ref ID: 625 67) Hay RJ. Methods for authenticating cell lines. Dev Biol Stand 1992;76:25-37. Ref ID: 5114 Abstract: Methods for authentication of cell lines include tests for microbial contamination,

verification of the species of origin, documentation of particular characteristics or functions and the absolute identification of individual cell lines. Experience indicates that mycoplasma contamination is still a serious problem in the cell culture field, with estimates on frequencies of infection varying from 10% upwards. The utilization of pre-screened reagents and antibiotic-free cultivation, plus the application of improved procedures, such as fluorescent dyes and molecular probes for detection, provide effective means to avoid mycoplasma infection and facilitate control. Viral contamination is perhaps more problematic, especially where no overt cytopathic effect results. The potential exists for the introduction of viruses from the cell culture technician or reagents used. Representative problem viruses are listed with standard procedures for screening. Detailed studies on animal cell cross contaminations have been performed and published. The frequency of detection of problem culture varied from 17-36% in studies performed in the USA. Both interspecies and intraspecies contaminations have been involved. Awareness of the potential for this problem plus the application of several

characterization procedures are key factors for control. For example, fluorescent antibody staining, iso-enzyme analysis, cytogenetic evaluation and DNA finger-printing using molecular probes are needed for quality assurance on seed stocks. The critical importance of generating well-characterized reference cell stocks for use over the years is emphasized

Notes: American Type Culture Collection, Rockville, MD 20852 68) Honma M, Kataoka E, Ohnishi K, Ohno T, Takeuchi M, Nomura N, et al. A new profiling

system for cell line identification for usen in cell banks in Japan. In Vitro Cell Dev Biol 1992;28A:24-8.

Ref ID: 3008 Notes: Using the polymorphic DNA probes, ChdTC-15, ChdTC-114, pYNH24 and lambdaTM18, a

DNA profiling system was developed that verified identities of individual cultured cell lines collected in the Japanese cell banks, JCRB, RCB and IFO. These highly polymorphic DNA probes include both VNTR sequences and substantial lengths of unique regions. In the mixed probe system, several distinct bands from four to eight can be used for cell line identification. These bands were widely spread in a range of molecular sizes, and were stable and reproducible under stringent conditions of Southern-blot hybridization. Because the DNA profile was specific for each individual human cell line, it is useful not only to authenticate many existing cultured cell lines but also to monitor their identity during propagation in a laboratory, and to confirm newly established line as unique. [Manuscript accepted Aug.13, 1991. - in printing] [Manuscript number = #901109RR]

69) Honma M, Kataoka E, Mizusawa H. Identification of cell lines by DNA

fingerprinting/profiling system. Soshiki Baiyo 1992;18:118-24. Ref ID: 3312 Notes: Language in Japanese. DNA fingerprinting and DNA profiling systems are introduced. 70) Kataoka E, Honma M, Ohnishi K, Sofuni T, Mizusawa H. Application of highly

polymorphic DNA markers to the identificaion of Hela cell sublines. In Vitro Cell Dev Biol 1992;28A:553-6.

Ref ID: 3481 Notes: Polymorphic DNA markers were used to identify eight sublines derived from HeLa. Using

five highly polymorphic minisatellite DNA probes, these cell lines were deistinguished and classified into six groups by Southern blot analysis. Polymorphic DNA markers, therefore, can provide a useful tool for monitoring genetic changes of a cell line during culture and for distinguishing sublines derived from the same origin.

71) Stacey G, Bolton B, Doyle A, Griffiths B. DNA fingerprinting--a valuable new technique

for the characterisation of cell lines. Cytotechnology 1992;9(1-3):211-6. Ref ID: 5115 Abstract: DNA fingerprinting is an important new development for the authentication of cell lines.

Multilocus methods such as those developed by Alec Jeffreys provide information on a wide range of genetic loci throughout the human genome and thus give a useful genetic "snap-shot" of a cell culture. Our work has shown that Jeffreys multilocus fingerprinting method can be applied to cell lines from a wide range of animals including reptiles, birds, fish and diverse mammals. It can also differentiate very closely related cell lines including those from the same mouse strain. Routine fingerprint analysis has enabled an unprecedented level of confidence in the consistency of cell stocks. Our results demonstrate that this straightforward method represents a powerful and readily interpreted system for cell authentication and exclusion of cross-contamination

Notes: European Collection of Animal Cell Cultures and Animal Cell Technology, Biologics Division, PHLS-CAMR, Porton Down, Wiltshire, UK

72) Budowle B, Giusti AM, Waye JS, Baechtel FS, Fourney RM, Adams DE, et al. Fixed-bin

analysis for statistical evaluation of continuous distributions of allelic data from VNTR loci, for use in forensic comparisons. Am J Hum Genet 1991 May;48(5):841-55.

Ref ID: 6113 Abstract: The detection of DNA polymorphisms by RFLP analysis is having a major impact on

identity testing in forensic science. At present, this approach is the best effort a forensic scientist can make to exclude an individual who has been falsely associated with an evidentiary sample found at a crime scene. When an analysis fails to exclude a suspect as a potential contributor of an evidentiary sample, a means should be provided to assess suitable weight to the putative match. Most important, the statistical analysis should not place undue weight on a genetic profile derived from an unknown sample that is attributed to an accused individual. The method must allow for limitations in conventional agarose-submarine-gel electrophoresis and Southern blotting procedure, limited sample population data, possible subpopulation differences, and potential sampling error. A conservative statistical method was developed based on arbitrarily defined fixed bins. This approach permits classification of continuous allelic data, provides for a simple and portable data-base system, and is unlikely to underestimate the frequency of occurrence of a set of alleles. This will help ensure that undue weight is not placed on a sample attributed to an accused individual

Notes: Forensic Science Research and Training Center, Federal Bureau of Investigation Academy, Quantico, VA 22135FAU - Budowle, B

73) Fehr J. [Eosinophilic granulocytes: on the way to knowledge of their functional

significance]. Hautarzt 1991 Sep;42(9):541-4. Ref ID: 6141 Abstract: Although the eosinophilic granulocyte has been recognized as a blood cell type for more

than 100 years, its functional significance has long remained an enigma. The introduction of successful isolation procedures resulted in rapidly progressive research efforts, but the concept of the functional role of this cell type made a full about turn between the 1970s and 1980s. In the 1970s, the role of the eosinophil was thought to be a homeostatic one, with its main task being to repair mast-cell-dependent tissue damage in parasitic and allergic disease. Further structural analysis and improved biological and clinical integration of such knowledge led to the completely revised concept of the 1980s: the eosinophil is now seen as the main culprit in the damage that accompanies allergic disease, as a result of mistaken identity between the parasite (where cytotoxic power against the aggressor is desirable) and the allergen (where the eosinophil's cytotoxic power results in self-damage). The latest research news about cytokine-dependent regulatory mechanisms governing the eosinophilic reaction supports our hope that by specific blocking of tissue hormones, such as the lymphocyte-derived IL-5, elegant ways of manipulating hypereosinophilic reactions will be found in the near future

Notes: Abteilung Hamatologie, Departement Innere Medizin, Universitatsspital ZurichFAU - Fehr, J

74) Ottesen SS, Kieler J. Expression of polymorphic HLA-A,B epitopes in human urothelial

cell lines. Anticancer Res 1991 Jan;11(1):217-23. Ref ID: 6142 Abstract: Malignant human urothelial cell lines propagated in vitro have previously been

demonstrated to express low amounts of monomorphic HLA-A,B,C as compared to premalignant urothelial cells. In this study the expression of polymorphic HLA-A,B epitopes in human urothelial cell lines have been investigated in greater detail. The expression of HLA-B locus coded epitopes in malignant TGrIII cells was demonstrated to be low or absent as compared to pre-malignant TGrII or slightly transformed TGrI cells, suggesting a mechanism by which malignant cells could escape from the host immune response. The extreme polymorphism of HLA-A,B,C antigens suggests that HLA typing could be used as a method to identify the origin of cell lines which is essential in the study of the process of malignant transformation in vitro, or when correlating in vitro data with clinical observations of the patient. Two urothelial cell lines classified as slightly transformed (TGrI) and two as

pre-malignant (TGrII) could, according to their expression of polymorphic HLA-A,B epitopes, be identified as genuine independent cell lines. One TGrII cell line previously designated Hu1734 shared the same HLA-A,B phenotype as the genuine HCV29 (TGrII) cell line and is therefore suspected to be a subline of the latter. Out of 18 cell lines and sublines classified as TGrIII the fidelity of four (Hu1922 and three sublines of T24) was proved by their HLA-A,B phenotype. Mistaken identity either by contamination or false labelling of the cultures was suspected in samples of six TGrIII cell lines and sublines. In four of these the HLA-A,B type characteristic for the T24 cell line could be demonstrated, but in general HLA typing of TGrIII cell lines as a method to identify the origin of the individual cell lines failed, primarily due to the decreased expression of HLA-A,B,C antigens and the apparent selective loss of HLA-B locus coded antigens

Notes: Fibiger Institute, Danish Cancer Society, CopenhagenFAU - Ottesen, S S 75) Penttila IA, Devery JM, Gibson CE, LaBrooy JT, Skerritt JH. Cellular and humoral

responses in coeliac disease. 1. Wheat protein fractions. Clin Chim Acta 1991;204:95-107. Ref ID: 3567 Notes: The humoral and cellular immune response of coeliac individuals to various wheat protein

fractions was studied using serum antibody ELISA assays and the indirect leucocyte migration inhibition factor (LMIF) assays. Greater migration inhibition factor activity was seen in coeliacs on a gluten-free-diet having low serum antibody titres, and using purified T-cells instead of total peripheral blood mononucleocytes. Gliadin was the most active fraction in both assays. Raised antibodies to low- molecular weight and high-molecular weight glutenin polypeptides was observed, though these proteins had little migration inhibition factor activity. No cellular or humoral response was seen to albumins or globulins. Proteins associated with the granules of well-washed wheat starch are distinct from gluten proteins and had little T-cell activity, correlating with clinical observations that properly prepared wheat starch is devoid of coeliac toxicity. The greater specificity of the humoral response for individual wheat protein fractions in this study, compared with the earlier reports, likely results from cross-contamination in the earlier work of each fraction with gliadin

76) Stacey GN, Bolton BJ, Doyle A. The quality control of cell banks using DNA

fingerprinting. EXS 1991;58:361-70.:361-70. Ref ID: 5116 Abstract: Reproducibility in animal cell culture technology requires careful preparation and

characterisation of banks of cell cultures. The two standard techniques used in the quality control of such banks are isoenzyme analysis and cytogenetics which require complex and time-consuming procedures to enable cell line identification. However, DNA fingerprinting is potentially a more powerful method of analysis which can detect mutation and intra-species cross-contamination. At the European Collection of Animal Cell Cultures (ECACC) multilocus fingerprint analysis using probes 33.6 and 33.15 has been assessed in the quality control of cell banks. This method has confirmed consistency between master and working banks, has proven useful over a wide species range and can differentiate closely related cell lines. The key advantage of this method is its ability to detect cross-contamination by cell lines from a wide range of species using a straightforward and economical test. In addition the reproducibility of DNA fingerprints indicates their possible role in cell line authentication procedures which are important for patent and product licence applications

Notes: European Collection of Animal Cell Cultures, PHLS Centre for Applied Microbiology and Research, Salisbury, Wiltshire, Great Britain

77) Green JB, Howes G, Symes K, Cooke J, Smith JC. The biological effects of XTC-MIF:

quantitative comparison with Xenopus bFGF. Development 1990 Jan;108(1):173-83. Ref ID: 6143 Abstract: Mesoderm in Xenopus and other amphibian embryos is induced by signals from the

vegetal hemisphere acting on equatorial or animal hemisphere cells. These signals are

diffusible and two classes of candidate signal molecule have been identified: the fibroblast growth factor (FGF) and transforming growth factor beta (TGF-beta) types. In this paper, we compare the effects of cloned Xenopus basic FGF (XbFGF) and electophoretically homogeneous XTC-MIF (a TGF-beta-like factor obtained from a Xenopus cell line) on animal pole explants. We find that they have a similar minimum active concentration (0.1-0.2 ng ml-1) but that, nonetheless, XTC-MIF is at least 40 times more active in inducing muscle. In general, we find that the two factors cause inductions of significantly different characters in terms of tissue type, morphology, gene expression and timing. At low concentrations (0.1-1.0 ng ml-1) both factors induce the differentiation of 'mesenchyme' and 'mesothelium' as well as blood-like cells. These latter cells do not, however, react with an antibody to Xenopus globin. This raised the possibility that the identification of red blood cells in other studies on mesoderm induction might have been mistaken, but combinations of animal pole regions with ventral vegetal pole regions confirmed that genuine erythrocytes are formed. The identity of the blood-like cells formed in response to the inducing factors remains unknown. At higher concentrations XTC-MIF induces neural tissue, notochord, pronephros and substantial and often segmented muscle. By contrast, XbFGF only induces significant amounts of muscle above 24 ng ml-1 and even then this is much less than that induced by XTC-MIF. For both factors an exposure of less than 30 min is effective. Competence of animal pole cells to respond to XbFGF is completely lost by the beginning of gastrulation (stage 10) while competence to XTC-MIF is detectable until somewhat later (stage 11). Since animal pole tissue is known to be able to respond to the natural inducer at least until stage 10, and perhaps until stage 10.5, this suggests that bFGF cannot be the sole inducer of mesoderm in vivo. Taken together, these results are consistent with XTC-MIF being a dorsoanterior inducer and XbFGF a ventroposterior inducer, suggesting that body pattern is established by the interaction of two types of inducing signal. This model is discussed in view of the qualitative and quantitative differences between the factors

Notes: Laboratory of Embryogenesis, National Institute for Medical Research, London, UKFAU - Green, J B

78) Johansson KE, Johansson I, Gobel UB. Evaluation of different hybridization procedures

for the detection of mycoplasma contamination in cell cultures. Mol Cell Probes 1990 Feb;4(1):33-42.

Ref ID: 5118 Abstract: Cell culture samples were analysed for mycoplasma contaminations with two different

DNA probes which have been described earlier. One probe (the H900 probe), derived from the 23S rRNA gene of Mycoplasma hyorhinis, cross-hybridized with virtually all mycoplasmas (including the acholeplasmas). The other probe (the T2 probe), derived from a protein gene of Acholeplasma laidlawii, cross-hybridized with most acholeplasmas. The two probes were compared in three different direct filter hybridization procedures without previous isolation of DNA or RNA. One of the procedures, developed in the present study, gave the highest sensitivity in DNA-RNA hybridization but also worked satisfactorily in DNA-DNA hybridization. The sensitivity of the H900 probe in filter hybridization experiments was compared with the sensitivity of a commercial probe for detection of mycoplasma contaminations in cell cultures. The H900 probe was found to be at least 25 times more sensitive for all cell culture mycoplasmas except for A. laidlawii, for which they were equally sensitive

Notes: National Veterinary Institute, Uppsala, Sweden 79) Azuma M, Kawamata H, Kasai Y, Yanagawa T, Yoshida H, Sato M. Induction of cells

with a chondrocyte-like phenotype by treatment with 1 alpha,25-dihydroxyvitamin D3 in a human salivary acinar cell line. Cancer Res 1989 Oct 1;49(19):5435-42.

Ref ID: 5566 Abstract: A clonal cell with an acinar cell phenotype, which was induced by 5-azacytidine

treatment of a neoplastic human salivary intercalated duct cell line, was cultivated in the presence of 1 alpha,25-dihydroxyvitamin D3. Morphological changes occurred; large cells that

were polygonal or round in shape and had numerous vacuoles in their cytoplasm appeared in the treated cells, whereas the same concentration of 1 alpha,25-dihydroxyvitamin D3 did not affect the morphology of the parental cells. Major alterations, such as expression of type II collagen, alpha and beta chains of S-100 protein, and sulfated proteoglycans, were observed in these cells with a phenotype similar to chondrocytes. After the removal of 1 alpha,25-dihydroxyvitamin D3 from the culture, the treated cells returned rapidly to the phenotype of the untreated cells. These findings indicate that the reversible differentiation into chondrocyte-like cells of a human salivary acinar cell line occurs in growth medium containing 1 alpha,25-dihydroxyvitamin D3

Notes: Second Department of Oral and Maxillofacial Surgery, Tokushima University School of Dentistry, JapanFAU - Azuma, M

80) Iga H, Azuma M, Nagamine S, Kaji R, Takase M, Yanagawa T, et al. Expression of

neurofilaments in a neoplastic human salivary intercalated duct cell line or its derivatives and effect of nerve growth factor on the cellular proliferation and phenotype. Cancer Res 1989 Dec 1;49(23):6708-19.

Ref ID: 5567 Abstract: A neoplastic salivary cell line with an ultrastructure similar to that of an intercalated duct

cell of the salivary gland, established from a human submandibular salivary gland, has been used in our laboratory as a model for studying mechanisms regulating cytodifferentiation in salivary glands. The expression of neurofilaments (Mr 200,000, 160,000, and 68,000) in the neoplastic human salivary intercalated duct cell line and its derivatives was found by the immunofluorescence staining technique, immunoblotting, or immunoelectron microscopy. In addition, these cells stained with Bodian impregnation and expressed specific antigens such as tubulin alpha and beta chain, HNK-1 antigen, and laminin. When these cells were cultured in the presence of nerve growth factor, only the cells with a myoepithelial cell phenotype formed the long cytoplasmic processes which were densely packed with ample microfibrils in addition to microtubule bundles, and they exhibited marked suppression of anchorage-independent and anchorage-dependent growths. These findings indicate that the characteristics of neoplastic human salivary intercalated duct cell line and its derivatives are similar to those of neuronal cells

Notes: Second Department of Oral and Maxillofacial Surgery, Tokushima University School of Dentistry, JapanFAU - Iga, H

81) Azuma M, Kawamata H, Kasai Y, Nagamine S, Yoshida H, Yanagawa T, et al. Effects of

retinoic acid on morphological features and biological markers of a neoplastic human salivary intercalated duct cell line in culture. Cancer Res 1988 Dec 15;48(24 Pt 1):7219-25.

Ref ID: 5565 Abstract: Retinoic acid has marked effects on the growth, morphological features, and biological

markers of a neoplastic human salivary intercalated duct cell clone in culture, whereas the cell clone was not affected by other retinoids such as retinol and retinal. A cell clone with ultrastructure and biological markers specific to the intercalated duct cells of human salivary glands was cultivated in the presence of retinoic acid. Major alterations, such as expression of tonofilaments, Mr 68,000 cytokeratin, and involucrin, were observed in those cells with a phenotype similar to that of keratinizing squamous cells. In addition, the coexpression of Mr 68,000 cytokeratin and carcinoembryonic antigen in these altered cells was found. Both the anchorage-independent and anchorage-dependent growths were markedly suppressed in the presence of retinoic acid. After the removal of retinoic acid from the culture, the treated cells returned rapidly to the phenotype of the untreated cells. These findings indicate that reversible differentiation into the keratinizing squamous cells of a neoplastic human salivary intercalated duct cell clone occurs in growth medium containing retinoic acid

Notes: Second Department of Oral and Maxillofacial Surgery, Tokushima University School of Dentistry, JapanFAU - Azuma, M

82) Kuritzkes DR, Rein M, Horowitz S, Droege G, Waldron MA, Bell DA, et al. Detached ciliary tufts mistaken for peritoneal parasites: a warning. Rev Infect Dis 1988 Sep;10(5):1044-7.

Ref ID: 6145 Abstract: Detached ciliary tufts of columnar epithelial cells from the female genital tract may be

mistakenly identified as protozoa when examined in wet mounts of fluid specimens in the laboratory because of their appearance and motility, although they are generally identified correctly in fixed specimens prepared for cytologic examination. A case of such mistaken identity in specimens from a gynecologic patient was documented, and the literature on ciliary tufts was reviewed. Infectious disease and gynecology consultants should be alert to the potential confusion arising from the presence of ciliary tufts in body fluids

Notes: Infectious Disease Unit, Massachusetts General Hospital, Boston 02114FAU - Kuritzkes, D R

83) Lucas C, Nelson C, Peterson ML, Frie S, Vetterlein D, Gregory T, et al. Enzyme-linked

immunosorbent assays (ELISAs) for the determination of contaminants resulting from the immunoaffinity purification of recombinant proteins. J Immunol Methods 1988 Oct 4;113(1):113-22.

Ref ID: 5119 Abstract: Two immunoassays have been developed for the quantitation of part-per-million levels of

contaminants likely to co-purify with monoclonal antibodies produced in tissue culture and purified by protein A affinity chromatography. These contaminants are bovine IgG originating from the fetal bovine serum used in cell culture, and protein A. Mouse IgG was shown not to interfere in the bovine IgG assay, where contamination levels of 0.2-0.7% bovine IgG were measured in the lots of monoclonal antibody tested. The protein A ELISA was developed with monoclonal antibody included in the standard, and in the preparations of monoclonal antibody tested, 64 parts per million (ppm) or less of protein A were demonstrated. An additional immunoassay was developed to quantitate monoclonal antibody contamination of two recombinant proteins, rHBsAg and rgp 120 from HIV, purified by affinity chromatography with such antibodies. Possible interference of monoclonal antibody quantitation by the respective antigens was examined in this ELISA, and contamination levels of less than 56 ppm of antibody were determined in the purified recombinant proteins. The three immunoassays were shown to be specific for the major protein contaminants in either monoclonal antibodies or the recombinant proteins and were necessary in demonstrating their purity

Notes: Department of Medicinal and Analytical Chemistry, Genentech, Inc, South San Francisco, CA 94080

84) Masters JRW, Bedford P, Kearney A, Povey S, Franks LM. Bladder cancer cell line

cross-contamination: Identification using a locus-specific minisatellite probe. Br J Cancer 1988;57:284-6.

Ref ID: 779 Notes: Cross-contamination of cells in culture is a common occurrence. Because mammalian cells

in monolayer culture can be difficult to distinguish morphologically culture can be difficult to distinguish morphologically, cross-contamination can pass undertected......

85) Matsuba I, Lernmark, Madsen O, Michelsen B, Nielsen JH, Scholler J, et al. Gene probes

to detect cross-culture contamination in hormone producing cell lines. In Vitro Cell Dev Biol 1988;24:1071-6.

Ref ID: 260 86) McCormick JJ, Yang DJ, Maher VM, Farber RA, Neuman W, Peterson WDJ, et al. The

HuT series of 'carcinogen-transformed' human fibroblast cell lines are derived from the human fibrosarcoma cell line 8387. Carcinogenesis 1988;9:2073-9.

Ref ID: 3919

Notes: In 1977 Kakunaga reported the carcinogen-induced transformation of the diploid human fibroblast cell line KD into focus-forming, morphologically altered cells. Cell lines were developed from 15 individual foci. These exhibited an infinite lifespan in culture and all those that were tested (7/7) formed malignant tumors (sarcomas) in athymic mice. The existing cell lines, designated HuT-11 to HuT-14, have been studied intensively during the past decade as examples of human fibroblasts malignantly transformed by treatment with a chemical carcinogen, 4-nitroquinoline-1- oxide. Recently, in comparing the HuT-11, HuT-12 and HuT-14 cell lines with KD cells, McCormick and Maher (Mutat. Res., 199, 273- 291, 1988) found evidence that the malignant cells could not have been derived from the latter. But, this did not rule out the possibility that as the target cells for his original study of carcinogen-induced transformation, Kakunaga had inadvertently used cells from some other, unidentified normal individual. Since the donor of such cells would not be known and the original cell line was not available, it would be impossible to determine the degree of identity between such a target cell line and the HuT cell lines. However, in the course of examining methods for such testing, we recently became aware that the isozyme pattern of these HuT cell lines was identical to that of the human fibrosarcoma-derived cell line 8387 established in 1966. We here report that the HuT cell lines and the 8387 cell line also exhibit an identical series of HLA determinants and identical restriction fragment length polymorphisms (RFLPs). Assuming that each of these three assays measures independently inherited characteristics, the chance that an unrelated donor of the fibroblasts that gave rise to the HuT cell lines happened to possess characteristics identical to those of the patient whose fibrosarcoma gave rise to the 8387 cell line is 1 x 10(-8). Therefore, we conclude that 8387 cells are the source of the malignant cells designated HuT from Kakunaga's original transformation experiment. Additional RFLP analysis, using a probe made from M13 bacteriophage DNA which detects a hyperpolymorphic 'minisatellite' pattern in human DNA, also showed that DNA from HuT-14 cells and from 8387 cells exhibit identical banding patterns, indicating that the cell lines were taken from the same individual. The latter banding patterns differed from that observed with DNA from KD cells.(ABSTRACT TRUNCATED AT 400 WORDS)

87) Thornley AL, Veale RB, Scott E, Reinecke L. Cross-contamination of human cell lines.

Br J Cancer 1988;58:687. Ref ID: 277 Notes: The problem highlighted by this letter is a serirous ongoing one. 88) Van Helden PD, Wiid IJF, Albrecht CF, Theron E, Thornley AL, Hoal-van Helden EG.

Cross-contamination of human esophageal squamous carcinoma cell lines detected by DNA fingerprint analysis. Cancer Res 1988;48:5660-2.

Ref ID: 47 89) Van Helden PD, Wiid IJ, Albrecht CF, Theron E, Thornley AL, Hoal-van Helden EG.

Cross-contamination of human esophageal squamous carcinoma cell lines detected by DNA fingerprint analysis. Cancer Res 1988 Oct 15;48(20):5660-2.

Ref ID: 6144 Abstract: DNA "fingerprint" analysis has recently become known as a valuable technique for

positive identification of any given individual. The chances for mistaken identity have been estimated to be 10(-6) for close siblings or as little as 10(-23) for randomly selected individuals. This methodology thus represents a significant improvement over previously established identification tests using protein or enzyme analysis techniques and has already found application in forensic medicine. One of the chief problems in tissue culture studies is the question of the unequivocal identity of the cultured cells used and the very real possibility of their being contaminated by cells of a similar morphological appearance. We report here the application of the DNA "fingerprint" technique to the genotypic analysis of cultured human squamous carcinoma cells. The results show that a number of lines, designation HCu, have become cross-contaminated. Lines SNO, HCu 10, and HCu 13 are genetically distinct,

however lines HCu 10, 18, 33, 37, and 39 are genetically identical and are in fact subcultures of the same cells. In addition, a myocardial line known as Girardi is shown to be identical to HeLa cells. The introduction of this technique to tissue culture laboratories could therefore prevent contaminated cultures from being disseminated or used in research studies

Notes: Department of Medical Biochemistry, University of Stellenbosch Medical School, Tygerberg, South AfricaFAU - van Helden, P D

90) Sato M, Azuma M, Hayashi Y, Yoshida H, Yanagawa T, Yura Y. 5-Azacytidine induction

of stable myoepithelial and acinar cells from a human salivary intercalated duct cell clone. Cancer Res 1987 Aug 15;47(16):4453-9.

Ref ID: 5568 Abstract: A neoplastic human salivary intercalated duct cell clone was cultured in 5 microM

5-azacytidine for 5 days at 37 degrees C; then the cells were trypsinized and subcultured in growth medium without 5-azacytidine. Thereafter, subclones were cloned from the subculture. Of 12 subclones isolated, 7 clonal cell lines were established and characterized. The two subclones composed of cells which were spindle shaped or stellate exhibited phenotypes similar to those of myoepithelial cells such as microfibrils and myosin and formed a myoepithelioma upon transplantation of the cells into nude mice. The other five subclones were composed of polygonal cells with numerous secretory granules in their cytoplasm and containing amylase that seems to be specific to acinar cells; transplantation of these cells into nude mice resulted in production of acinic cell carcinoma. These findings indicate that a neoplastic human salivary intercalated duct cell is capable of at least bidirectional differentiation

91) Azuma M, Hayashi Y, Yoshida H, Yanagawa T, Yura Y, Ueno A, et al. Emergence of

differentiated subclones from a human salivary adenocarcinoma cell clone in culture after treatment with sodium butyrate. Cancer Res 1986 Feb;46(2):770-7.

Ref ID: 5563 Abstract: A human salivary gland adenocarcinoma cell line, which has ultrastructure and biological

markers specific to the intercalated duct cells of human salivary glands, was cultured in 5 mM sodium butyrate for 12 days; then the cells were trypsinized, subcultured for an additional 16 days, and then transferred to growth medium without sodium butyrate. Morphological changes appeared about 1 wk after return to growth medium without sodium butyrate; cells being spindle or stellate in shape appeared in the treated cells, whereas the untreated cells were polygonal in shape. This morphologically altered phenotype persists after more than 14 mo of culture in growth medium without sodium butyrate. Of 40 subclones isolated, 2 clonal cell lines were established from the subculture and characterized. The other 38 subclones were accompanied by cell death during the subcultures. The clonal lines exhibited a phenotype similar to myoepithelial cells such as myosin, beta-chain of S-100 protein, myofilaments, and oxytocin receptor in addition to decreased tumorigenicity and anchorage-independent growth. These findings indicate that commitment to differentiation into myoepithelial cells and conversion from malignant to normal phenotype occur in a human salivary gland adenocarcinoma cell line following the sodium butyrate treatment

92) White MC, Newland P, Daniels M, Turner SJ, Mathias D, Teasdale G, et al. Growth

hormone secreting pituitary adenomas are heterogeneous in cell culture and commonly secrete glycoprotein hormone alpha-subunit. Clin Endocrinol (Oxf) 1986 Aug;25(2):173-9.

Ref ID: 5120 Abstract: Cell culture methods were used to assess whether human pituitary adenomas secreting

GH and associated with clinical acromegaly also secreted the structurally unrelated glycoprotein hormone alpha-subunit. Thirty-two tumours, together with peri-adenomatous tissue from two of them and three normal pituitaries were studied. Anterior pituitary hormones were measured by radioimmunoassay and included PRL, TSH, LH, FSH, and ACTH, as well as GH and alpha-subunit. Normal pituitary tissues secreted all hormones assayed. All 32

tumours secreted GH ranging from 241 to 5556 ng/2 X 10(5) cells/24 h and 12 (37.5%) secreted alpha-subunit in amounts which could not be accounted for by cross-reaction of other hormones or contamination by normal pituitary tissue, and which ranged from 10.3 to 73.5 ng/2 X 10(5) cells/24 h. Ten other tumours also secreted alpha-subunit but in very small amounts, not exceeding 1.8 ng/2 X 10(5) cells/24 h. PRL was secreted from 21 tumours (66%), and small amounts of other hormones, chiefly LH and TSH, were occasionally secreted from tumours. These cell culture studies would suggest that pituitary adenomas causing acromegaly are hormonally heterogeneous and that PRL and glycoprotein alpha-subunit are commonly detected in addition to GH

93) Yoshida H, Azuma M, Yanagawa T, Yura Y, Hayashi Y, Sato M. Effect of dibutyryl cyclic

AMP on morphologic features and biologic markers of a human salivary gland adenocarcinoma cell line in culture. Cancer 1986 Mar 1;57(5):1011-8.

Ref ID: 5564 Abstract: Dibutyryl cyclic AMP (dB-cAMP) has marked effects on the growth, morphologic

features, and biologic markers of a human salivary gland adenocarcinoma cell line in culture. A cell line with ultrastructure and biologic markers specific to the intercalated duct cells of human salivary glands was cultivated in the presence of dB-cAMP. Major alterations, such as process formation and expression of myofilaments and oxytocin receptor in addition to myosin and S-100 protein, were observed in those cells with a phenotype similar to myoepithelial cells. Both the anchorage-independent and anchorage-dependent growths were markedly suppressed in the presence of dB-cAMP. After the removal of dB-cAMP from the culture, the treated cells returned rapidly to the phenotype of the untreated cells. These findings indicate that reversible differentiation into the myoepithelial cells of a human salivary gland adenocarcinoma cell line occurs in growth medium containing dB-cAMP

94) Sato M, Hayashi Y, Yanagawa T, Yoshida H, Yura Y, Azuma M, et al. Intermediate-sized

filaments and specific markers in a human salivary gland adenocarcinoma cell line and its nude mouse tumors. Cancer Res 1985 Aug;45(8):3878-90.

Ref ID: 5561 Abstract: The adenocarcinoma cell line HSG from human salivary gland, which proliferates in vitro

or in nude mice, was examined by the immunoperoxidase method for the expression of three different types of intermediate-sized filaments (IFs) and of specific antigens such as carcinoembryonic antigen, S-100 protein, secretory component, lactoferrin, myosin, tropomyosin, and actin. The cultured HSG cells were found to express three different types of IFs defined by antibodies to keratin, vimentin, and desmin. In HSG cells proliferating in vitro at 34 degrees C and 37 degrees C but not at 39 degrees C, the expression of tropomyosin and carcinoembryonic antigen was observed, although myosin and S-100 protein were not detected. The expressions of actin, lactoferrin, and secretory component were restricted to cultured HSG cells at 39 degrees C and 37 degrees C, respectively. Transplantation of HSG cells into nude mice resulted in the establishment of a nude mouse system with malignant characteristics such as invasion and metastasis. The expression of IFs in the primary tumors was restricted to keratin and desmin IFs, whereas coexpression of keratin, vimentin, and desmin IFs was observed in some neoplastic cells present in the metastatic tumors in regional lymph nodes and lung. In addition, expression of actin, myosin, tropomyosin, and S-100 protein was found in the metastatic tumors, whereas myosin and S-100 protein were not detected in the primary tumors. Moreover, the metastatic tumors were almost occupied by the neoplastic cells with oncocytic changes, although oncocytic change was not found in the cultured HSG cells and their primary tumors

95) Sato M, Yoshida H, Hayashi Y, Miyakami K, Bando T, Yanagawa T, et al. Expression of

epidermal growth factor and transforming growth factor-beta in human salivary gland adenocarcinoma cell line. Cancer Res 1985 Dec;45(12 Pt 1):6160-7.

Ref ID: 5562

Abstract: Neoplastic intercalated duct cells that originated from a human submandibular salivary gland release transforming growth factor and human epidermal growth factor into serum-free medium. The transforming growth factor with the soft agar colony-forming activity when assayed in the presence of mouse epidermal growth factor on normal rat kidney clone 49F indicator cells, but without mitogenic action to quiescent 3T3 cells, does not compete with epidermal growth factor for receptor binding, is heat and acid resistant but trypsin and dithiothreitol sensitive, and therefore is of the transforming growth factor-beta class. The immunoreactive human epidermal growth factor is detected by radioimmunoassay on certain fractions obtained by the Bio-Gel P-60 chromatography of neoplastic epithelial duct cells originated from a human submandibular salivary gland-conditioned medium and is biologically very similar to mouse epidermal growth factor. Moreover, the presence of human epidermal growth factor in cultured neoplastic epithelial duct cells originated from a human submandibular salivary gland is demonstrated by the peroxidase:antiperoxidase method

96) Nelson-Rees WA, Daniels DW, Flandermeyer RR. Cross-contamination of cell culture.

Science 1984;212:446-52. Ref ID: 1004 97) Reyes A, Cardenas ML. All hexokinase isoenzymes coexist in rat hepatocytes. Biochem J

1984 Jul 15;221(2):303-9. Ref ID: 5121 Abstract: The cellular distribution of hexokinase isoenzymes, N-acetylglucosamine Kinase and

pyruvate kinases in rat liver was studied. Hepatocytes and non-parenchymal cells with high viability and almost no cross-contamination were obtained by perfusion in situ of the liver with collagenase, with the use of an enriched cell-culture medium in all steps of cell isolation. Separation of hexokinase isoenzymes was done by DEAE-cellulose chromatography, and enzyme activities were measured by a specific radioassay. Cytosol from isolated hepatocytes contained high-affinity hexokinases A, B and C, in addition to hexokinase D. The last-mentioned represented about 95% of total glucose-phosphorylating activity. Only hexokinase A was found associated t the particulate fraction. Isolated non-parenchymal cells contained only hexokinases A, B and C. N-Acetylglucosamine kinase was measured with a specific radioassay and was found as a single enzyme form in both hepatocytes and non-parenchymal cells, with higher activities in the former. Pyruvate kinase isoenzyme L was present only in the hepatocytes and isoenzyme K only in the non-parenchymal liver cells, confirming that they are good cellular markers

98) Sato M, Hayashi Y, Yoshida H, Yanagawa T, Yura Y, Nitta T. Search for specific markers

of neoplastic epithelial duct and myoepithelial cell lines established from human salivary gland and characterization of their growth in vitro. Cancer 1984 Dec 15;54(12):2959-67.

Ref ID: 5560 Abstract: The neoplastic epithelial duct cells human salivary gland (HSG) and myoepithelial cells

human pleomorphic adenoma (HPA) established from human salivary gland were examined by the immunoperoxidase method for the presence of specific antigens such as carcinoembryonic antigen (CEA), S-100 protein, secretory component (SC), lactoferrin (LF), and myosin. Isolation of the cells and their morphologic features were reported previously. Consequently, the presence of CEA, SC, and LF in the HSG cells was demonstrated. The HPA cells were identified to express the specific antigens reactive to anti-S-100 protein, anti-myosin and anti-CEA sera in addition to the presence of oxytocin receptor. When the two cell lines were co-cultured in monolayer culture or within the sponge matrix, a large number of ductlike or tubular structures were formed in an optimal ratio of 1:2 in HSG and HPA cells, whereas the cultures of HSG cells only grew with occasional formation of ductlike structure. In addition, in HSG and HPA cells in an area with their contact in the mixed cultures, CEA staining was intensified as compared with the culture of HSG or HPA cells only and further S-100 protein was detected in HSG cells, whereas S-100 protein was not detected in the culture of HSG cells

only. These findings strongly suggest that the intercalated duct and myoepithelial cells from human salivary gland propagate with their interaction together in the expression of specific antigens such as CEA and S-100 protein or in the morphogenesis of salivary gland epithelial cells

99) Holliday R, Kirkwood TB. Theories of cell ageing: a case of mistaken identity. J Theor

Biol 1983 Jul 21;103(2):329-30. Ref ID: 6147 100) Olsen EG. Myocarditis--a case of mistaken identity? Br Heart J 1983 Oct;50(4):303-11. Ref ID: 6146 101) [Nise "Junkei-mouse" Demawaru]. [Shizen] 1982;21-2. Ref ID: 3885 Notes: [In Japanese] in "Mini Scope Section" Document introduces trouble of the BALB/c mice

distributed through private company. References scited in this issue are Science{3884} and New Scientist written by Kahan,B. et al.

102) Bell RS, Goodman SB, Fornasier VL. Coccygeal glomus tumors: a case of mistaken

identity? J Bone Joint Surg Am 1982 Apr;64(4):595-7. Ref ID: 6148 Abstract: We undertook an anatomical and histological study to differentiate glomus-cell tumors of

the pericoccygeal tissues from the normal coccygeal body. Removal of the coccyx was performed on five consecutive autopsy specimens from patients with no history of coccygeal symptoms. In each specimen, the coccygeal body (glomus coccygeum) was identified grossly and histologically. The histological appearance was indistinguishable from that of photomicrographs published in case reports of patients with glomus tumors of the coccyx. It is likely that the so-called tumors reported previously were in actuality normal glomus bodies

103) Hunter A, Brierley K, Tomkins D. 46,XX/46XY chromosome complement in amniotic

fluid cell culture followed by the birth of a normal female child. Prenat Diagn 1982 Apr;2(2):127-31.

Ref ID: 5122 Abstract: Experience indicates that the most likely explanation for a mixture of 46,XX/46,XY cells

in an amniotic fluid sample is that of maternal cell contamination and that a normal male child is to be expected at birth. We report the bith of a normal female child following prenatal diagnosis of such a mixture. Extensive postnatal studies failed to reveal an XY cell line. The possible sources of the XY cell line are discussed, as are the various techniques that were applied in an effort to discover it's origin. Cross-contamination of samples could be ruled out and there was no evidence of an unsuspected twin pregnancy. It is clear from this case that not all 46,XX/46,XY results obtained in amniotic fluid can be assumed to represent maternal cell contamination and some effort should be made to eliminate other potential sources for such a mixture

104) Kahan B, Auerbach R, Alter BJ, Bach FH. Histocompatibility and isoenzyme differences

in commercially supplied "BALB/c" mice. Science 1982;217:379-81. Ref ID: 3884 Notes: BALB/c mice obtained commercially were found to differ significantly from the standard

phenotype of BALB/c strain mice. Isoenzyme tests and H-2 haplotype analyses indicated that the majority of mice from two of the three sources tested appeared mixed, frequently heterozygous, and did not consistently express either the expected H-2 or glucose phosphate isomerase type

105) Dickson D. Contaminated cell lines. Nature 1981;289:227.

Ref ID: 1762 Notes: Yet another corruption of the scientific literature has been uncovered with the discovery that

several cell lines widely quoted in the scientific literature as originating from patients with Hodgkin's disease have now been shown not to be human cells at all, but to come from a strain of owl monkdy ( see the accompanying article, pages 228-230). The cells were grown in culture by Dr.John C. Long, who resigned a year ago as assistant pathologidst at the Massachusetts General Hospital and associate professor of pathology at Harvard Medical School, after admitting to having faked data in a research paper published in the Journal of the National Cancer Institute in 1979.

106) Harris NL, Gang DL, Quay SC, Poppema S, Zamecnik PC, Nelson-Rees WA, et al.

Contamination of Hodgkin's desease cell culture. Nature 1981;289(5795):228-30. Ref ID: 1002 Notes: Several laboratories have recently reported the establishment and characterization of

long-term cell lines thought to be related to the neoplastic cell of Hodgkin's disease. Here, Harris et al. discuss evidence that some of these lines are, in fact, not related to Hodgkin's disease but are non-human contaminants

107) Nelson-Rees WA, Daniels DW, Flandermeyer RR. Cross-contamination of cells in culture.

Science 1981;212(4493):446-52. Ref ID: 1763 Notes: Lists are presented of references to all known publications descrebing cell properties that

serve to characterize (i) known strains of HeLa and purported human cell lines indicated as HeLa contaminants,(ii) strains of human cell lines by cells from one or more other species. Frequencies of cell cross-contaminations are cited and references are presented to relatively simple techniques that could serve to detect such contamination.

108) O'Brien SJ, Shannon JE, Gail MH. A molecular approach to the identification and

individualization of human and animal cells in culture: Isozyme and allozyme genetic signatures. In Vitro Cell Dev Biol 1981;16:119-35.

Ref ID: 1006 109) Nelson-Rees WA, Hunter L, Darlington GJ, O'Brien SJ. Characteristics of HeLa strains:

permanent vs. variable features. Cytogenet Cell Genet 1980;27(4):216-31. Ref ID: 5135 Notes: Characteristic rearranged human chromosome markers have been observed in a variety of

HeLa cell sublines and in five suspected HeLa contaminant lines originally thought to be derived from differentiated tissues of different individual patients. The allozyme genetic signatures, representing the composite enzyme phenotype at eight polymorphic loci, of each of the studied contaminant lines were identical to each other and to those of HeLa cells. The probability that each of these lines would have an identical genetic signature (since the frequency of the HeLa genotype is 0.0017) is 4.2 X 10(-15). Differences between cell lines, however, could be detected by isoelectric focussing of the isoenzymes for glucose-6-phosphate dehydrogenase. The cell lines, including CLL 74, the older Chang liver line, failed to express five liver-specific proteins. One protein was detected in a new liver cell culture. Variations in cytogenetic, biochemical, and differentiated functions during continuous cell culture are discussed.

Publication Types: Review 110) Wright WC. Two relative efficiencies of polymorphic enzymes for characterizing cell

lines, detecting contaminations, and monitoring transplants. In Vitro Cell Dev Biol 1980;16:875-83.

Ref ID: 38 Notes: A new calculation of the relative efficiency of polymorphic enzyme markers, called the REB,

was determined and compared with one of Fisher's determinations of the relative efficiency called REA here.

111) Peterson WDJr, Simpson WF, Hukku B. Cell culture characterization: Monitoring for cell

identification. Method Enzymol 1979;58:164-78. Ref ID: 1005 112) Lavappa KS. Survey of ATCC stocks of human cell lines for HeLa contamination. In

Vitro Cell Dev Biol 1978;14:469-75. Ref ID: 371 Notes: Seed stocks of human cell lines deposited at the American Type Culture Collection (ATCC)

have been examined for cross-contamination with HeLa cells using Giemsa banded marker chromosomes. Sixteen additional cell lines investigated have been found to exhibit marker chromosomes typical of HeLa cells. Quinacrine fluorescence studies further revealed the absence of Y chromosome in these lines. These observations indicate that the lines are HeLa derivatives.

113) Lavappa KS, Verma RS, Dosik H. Cyto genetic investigations of long-term human hetero

ploid cell lines J-111 detroit-98-CCL18.2 and detroit-98-CCL18.3 with HeLa characteristics. Am J Hum Genet 1977;29:67A.

Ref ID: 69 Notes: Three different human heteroploid cell lines were cytogenetically examined, using

sequential QFQ (Q-bands by fluorescence using quinacrine) and RFA(R-bands by fluorescence using acridine orange) techniques. At least 20 metaphases were randomly selected from each line. Each cell line had all the human chromosomes except the Y. Marker chromosomes typical of HeLa were also found in these lines. In J111 there was one compy of HeLa marker #1 (HM1), 2 copies of HM2,3-4 copies of HM3 and 2 copies of HM4. In addition .........

114) Becker SN, Pepin DW, Rosenthal DL. Mesothelial papilloma: a case of mistaken identity

in a pericardial effusion. Acta Cytol 1976 May;20(3):266-8. Ref ID: 6149 115) Grimwade S. HeLa takes over. Nature 1976;259:172. Ref ID: 1603 Notes: Although it was suggested in 1967 by Gartler (Natn.Cancer Inst. Monogr.,26,167) on the

basis of isoenzyme patterns that not all cell lines in common use were what they purported to be, it was not until last year that cytogenetic studies by Nelson-Ress et al. (Science, 184, 1093; 1974) demonstrated conclusively the extent of contamination. Gartler showed that many commonly used cell lines, supposedly of Caucasian origin, contained the A-type isoenzyme of glucose-6-phosphate dehydrogenase (G6PD), which is in fact found only in approximately 30% of the Negro rade.

116) Haneen WK. HeLa cells and their possible contamination of other cell lines. Hereditas

1976;82:217-48. Ref ID: 379 117) Lavappa KS, Macy ML, Shannon JE. Examination of ATCC stocks for HeLa marker

chromosomes in human cell lines. Nature 1976;259:211-3. Ref ID: 376 118) Nelson-Rees WA, Flandermeyer RR. HeLa cultures defined. Science 1976;191:96-8. Ref ID: 381 Notes: A list is presented of references to all known publications on properties which have served to

relate strains of HeLa cells to each other as well as to indict other purported human cell lines as

HeLa cell contaminants. Eleven additional cell lines not previously indicted are described. When they exhibit (i) type A (fast) mobility for glucose-6-phosphate dehydrogenase, (ii) phosphoglucomutase type 1 at locus 1 and locus 3, (iii) absence of a Y chromosome by fluorescent staining, and (iv) possession of a complex of trypsin-Giemsa banded marker chromosomes present in known HeLa cells, then cell substrates regardless of designation should be considered de facto strains of HeLa.

119) Nelson-Rees WA, Flandermeyer RR. Inter- and intraspecies contamination of human

breast tumor cell lines HBC and BeCa5 and other cell culture. Science 1976;195:1343-4. Ref ID: 1001 120) Hough AJ, Sokoloff L. Tissue sampling as a potential source of error in experimental

studies of cartilage. Connect Tissue Res 1975;3(1):27-31. Ref ID: 5123 Abstract: Although it might seem trite to point out that tissue sampling is a potential source of

experimental error, this survey disclosed that even experienced investigators in fact often work with cartilage that is contaminated by non-cartilaginous tissue of which they were unaware. Twenty-two specimens ranging from chick embryo sternum to bovine nasal septum were studied by serial sectioning. Eighteen of the 22 contained extraneous tissue comprising from 3 to 50% of the cross-sectional area. The impact of the contamination depends on the use being made of the material and probably is greatest in cell culture studies because chondrocytes and fibroblasts have large differences in population doubling time. Several approaches for minimizing the error are suggested by the findings. Histological examination of specimen material is thus a desirable quality control procedure in the design and interpretation of experiments on cartilage as well as other tissues

121) Culliton BJ. HeLa cells: contaminating culture around the world. Science

1974;184:1058-9. Ref ID: 1003 122) Nelson-Rees WA, Flandermeyer RR, Hawthorne PK. Banded marker chromosomes as

indicators of intraspecies cellular contamination. Science 1974;184:1093-6. Ref ID: 374 Notes: Chromosome banding revealed marker chromosomes characteristic of HeLa cells in cultures

designated HEK. HEK/HRV, HBT-3, HBT-39B, MA160, and a strain of SA-4TxS-HuSa. Other HeLa cell characteristics found were glucose-6-phosphate dehydrogenase type A mobility and lack of the Y chromosome.

123) Francke U, Hammond DS, Schneider JA. The band patterns of twelve D98/AH 2 marker

chromosomes and their use for identification of intraspecies cell hybrids. Chromosoma 1973;41:111-21.

Ref ID: 380 124) Ludovici PP, Holmgren NB. Cell culture contaminants. Methods Cell Biol

1973;6:143-208. Ref ID: 3584 125) Walker JR. Identification and misidentification of the chromosomes of heteroploid cell

lines. J Natl Cancer Inst 1973;51:1113-7. Ref ID: 378 126) Fogh J, Holmgren NB, Ludovici PP. A review of cell culture contaminations. In Vitro

1971;7:26-41. Ref ID: 3585

Notes: [Abstruct] It is apparent that most laboratories employing tissue and cell cultures for experimental or diagnostic purposes encounter contaminations. If not detected and properly controlled or eliminated, contaminants may severely affect such investigations, and be the cause of incorrect interpretations. The Committee on Contaminations in Cell and Tissue Cultures of the Tissue Culture Association considered it as one of its functions to review the most pertinent literature related to contaminations with cells, bacteria, yeasts, molds, parasites, Mycoplasma, and viruses. the present review article is not intended to cover the subject completely but will, hopefully, be helpful to workers in the area of cell and tissue cultures who are concerned about the problem of contamination. Apparently many are not thoroughly familiar with the proper procedures for prevention, detection, and, perhaps, elimination of contaminants from the experimental systems that they employ.

127) Cross GF, Goodman MR, Chatterji J, Beswick TS, Chapman JA. Another case of

mistaken identity: rubella and mycoplasma. J Gen Virol 1970 Jul;8(1):77-81. Ref ID: 6150 128) Matsuya Y, Green H. Somatic cell hybrid between the established junam line D98

(presumptive HeLa) and 3T3. Science 1969;163:697-8. Ref ID: 70 Notes: Somatic cell hybrids have been made between an established human cell line with a long

culture history and an established mouse fibroblast line. 129) Gartler SM. Apparent HeLa cell contamination of human heteroploid cell lines. Nature

1968;217:750-1. Ref ID: 365 Notes: Interspecific cell culture contamination has been detected several times by karyotypic and

immunological procedures. These same measurements are of little value as detectors of intraspecific contamination, but polymorphic variants detectable at the cell culture level can be very useful for this purpose.

130) Gartler SM. Genetic markers as tracers in cell culture. Natl Cancer Inst Monogr

1967;26:167-95. Ref ID: 1811 Notes: Same as the refer ID# 383.This is the first paper representing the tissue culture

cross-contamination of HeLa cells. This paper deals with the use of various levels of genic expression to distinguish between cell cultures of different origin. DNA replication patterns, variations in the types of RNA molecules synthesized by different tissues, and the persistence of certain genetic markers in cell culture are considered. It is suggested that DNA-RNA hybridization as a means of comparing tissue and cell culture RNA molecules offers a promising tool for answers to questions of tissue origin-cell culture relationships. A survey of certain genetic markers in established human cell lines revealed the possibility of a serious problem of intraspecific contamination. Eighteen established human cell lines of supposed independent origin were shown to be identical with regard to their glucose-6-phosphate dehydrogenase (G6PD) (A) and phosphoglucomutase (1) types.

131) Gartler SM. Genetic markers as tracers in cell culture. Natl Cancer Inst Monogr

1967;26:167-95. Ref ID: 383 Notes: Same as the refer ID# 1811. This is the first paper representing the tissue culture

cross-contamination of HeLa cells. This paper deals with the use of various levels of genic expression to distinguish between cell cultures of different origin. DNA replication patterns, variations in the types of RNA molecules synthesized by different tissues, and the persistence of certain genetic markers in cell culture are considered. It is suggested that DNA-RNA hybridization as a means of comparing tissue and cell culture RNA molecules offers a

promising tool for answers to questions of tissue origin-cell culture relationships. A survey of certain genetic markers in established human cell lines revealed the possibility of a serious problem of intraspecific contamination. Eighteen established human cell lines of supposed independent origin were shown to be identical with regard to their glucose-6-phosphate dehydrogenase (G6PD) (A) and phosphoglucomutase (1) types.