electron microscopic and peroxidase cytochemical analysis ... · in 1974 and termed the...

[CANCER RESEARCH 40, 4473-4481, December 1980]0008-5472/80/0040-4473S02.00

Electron Microscopic and Peroxidase Cytochemical Analysis of PinkPseudo-Chediak-Higashi Granules in Acute Myelogenous Leukemia1

William A. Dittman, Robert J. Kramer, and Dorothy F. Bainton2

Sacred Heart Medical Center, Spokane. Washington 99024 [W. A. D.]; Station 5. Pasco. Washington 99301 Â¡R.J. K.J; and Department of Pathology. University ofCalifornia School of Medicine. San Francisco. California 94143 Â¡D.F. B.Â¡

ABSTRACT

Giant round pink inclusions (=2 jam)were seen in neutrophilicmyeloblasts, promyelocytes, and myelocytes from three patients with acute myelogenous leukemia. On preliminary examination of the bone marrow smears, these inclusions lookedlike ingested red blood cells in that they were pink and notazurophilic. The bone marrow specimens were processed forthe electron microscopic demonstration of peroxidase with3,3'-diaminobenzidine and H2C>2at pH 7.6. In all three cases,the inclusions were determined to be large peroxidase-positive

granules since they were limited by a single unit membraneand, unlike endocytized red blood cells, were not containedwithin phagocytic vacuoles. The granules were homogeneouslydense for peroxidase and showed no obvious crystalline structure when examined stained or unstained on grid. We believethat they correspond to the giant pink round granules VanSlyck and Rebuck observed in immature leukemic granulocytesin 1974 and termed the pseudo-Chediak-Higashi anomaly. Like

the giant purple granules seen in leukemia with this anomaly,these granules also appear to be an abnormal variant of peroxidase-positive azurophil (primary) granules. Their lack ofazurophilia is due to the absence of sulfated glycoaminogly-

cans.

INTRODUCTION

In 1974, Van Slyck and Rebuck (40) observed, on Wright's-

stained smears, giant round granules in leukemic cells from 2patients with acute myelomonocytic leukemia. Since this acquired morphological abnormality closely mimicked the giantround granules seen in the Chediak-Higashi anomaly, a rareautosomal recessive disorder, they termed it the pseudo-Chediak-Higashi anomaly of acute leukemia. Since some of thegiant granules stained azurophilic while others were pink, VanSlyck and Rebuck hypothesized that they represented abnormalities of azurophilic (primary) and specific (secondary) granule populations, respectively. More cases of this anomaly associated with leukemia have now been reported (16, 18, 20,26, 36, 38), and three cases with purple-staining giant granules

have been thoroughly studied by electron microscopy andperoxidase cytochemistry (30, 39). These investigators demonstrated that these dark giant granules were indeed abnormalvariants of the peroxidase-positive azurophilic granule population. Interestingly, their clinical courses were complicated byDIG.3

' This investigation was supported by Grant CA-14264 from the National

Cancer Institute, Department of Health. Education and Welfare2 To whom requests for reprints should be addressed.3 The abbreviations used are: DIG, disseminated intravascular coagulation;

AML, acute myelogenous leukemia; RER. rough endoplasmic reticulum.Received May 13. 1980; accepted September 12. 1980.

We have had the opportunity to study 3 patients with AMLwhose blasts, promyelocytes, and myelocytes contained enormous round pink inclusions, initially thought to be ingestedRBC. Analysis by electron microscopy and peroxidase cytochemistry revealed that these pink structures were large peroxidase-positive granules and therefore also represented ab

normal variants of the azurophilic (primary) granule population.In addition, these 3 cases did not have DIC.

MATERIALS AND METHODS

Case Studies

Case 1. A 62-year-old white housewife with a history of

hypertension and arthritis was sent for a hematological consultation in June 1974 because of fatigue, anemia, and recentonset of bruising. Physical examination revealed pallor andmultiple ecchymoses but not organomegaly or adenopathy.Laboratory examination of the blood revealed pancytopenia(Table 1), but no abnormal circulating leukocytes were detected. A bone marrow aspiration and biopsy specimen werehypercellular, showing an increase in the granulocytic series,5% myeloblasts, and promyelocytes containing Auer rods andsalmon-pink inclusion bodies. The pink inclusions were usuallylarge (1 to 2 firn) and round, but they were occasionally tearshaped or tapered at both ends. Rarely, both forms werepresent in a single cell. The patient's condition was diagnosed

as AML.Over the next 2.5 years, the patient was given various

treatments for AML and had a smoldering course withoutcomplete remission. The Auer rods disappeared with partialremission, but the giant inclusions merely decreased in number. Initially, the patient was treated with daunomycin, vincris-tine, prednisone, and 1-/?-D-arabinofuranosylcytosine. Because she came near death twice and her marrow remainedabnormal, she was switched to a maintenance dose of dailycyclophosphamide and monthly vincristine. During the ensuingmonths, she had a number of complications, mainly infections.A bone marrow specimen was processed for fine-structural

examination in November 1975.In October 1976, the patient was found to have an endo-

metrial adenocarcinoma, which was treated with radiation therapy. At that time, her WBC count was 0.7 x 109/liter. She

developed bowel obstruction and died on January 8, 1977. Anautopsy revealed metastatic carcinoma to the peritoneum, liver,pleura, and bone. The marrow was hypercellular. During thecourse of her illness, there was no clinical or laboratory evidence of DIC.

Case 2. A white 16-year-old female was found to have AMLin May 1976. She had experienced dizziness on exertion, easybruising, and heavy menstrual flow for 6 weeks. Her WBC

DECEMBER 1980 4473

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W. A. Dittman et al.

count was 5.0 x 109/liter with a marked shift to the left in the

neutrophilic series, including 20% myeloblasts. Her blood dataare summarized in Table 1. Her bone marrow was hypercellularand showed granulocytic hyperplasia with a marked shift to theleft. The myeloblasts contained large salmon-pink cytoplasmic

inclusions and occasional typical azurophilic Auer rods. Culturefor Philadelphia chromosome was negative. Leukocyte alkalinephosphatase was normal. Bone marrow was obtained for electron microscopic examination in May 1976.

The patient was treated with thioguanine, vincristine, pred-nisone, 1-yS-D-arabinofuranosylcytosine, and cyclophospha-

mide and was in remission for 6 months. During this period, nogiant granules were seen in her granulocytic precursor cells. Abone marrow specimen taken in January 1977 showed 14%myeloblasts but no giant granules. The patient did not respondto reinduction therapy and died in April 1977 of a pulmonaryinfection. There was no evidence of DIG.

Case 3. In January 1976, the preoperative leukocyte differential for a white 52-year-old male scheduled to undergoelective surgery revealed abnormal circulating leukocytes withlarge round pink inclusions. About 12% of the cells were blast-

like and contained very large round pink granules. The physicalexamination revealed no abnormalities. The complete bloodcount is recorded in Table 1. An aspirate of sternal bonemarrow showed granulocytic hyperplasia and the same largepink granules in blasts and promyelocytes. Some blasts contained typical azurophilic Auer rods. A bone marrow specimenwas obtained for electron microscopy in March 1976. By May1976, the patient had become symptomatic with fatigue andbleeding and had a falling hematocrit and an increase in hiscirculating blasts to 50%. He received chemotherapy for thefirst time on March 9, 1976. Initially, he was given cyclophos-

phamide, thioguanine, prednisone, and vincristine every 4weeks, but, beginning in November 1977, he received thetreatment regimen every 6 weeks. His last hematology values,recorded on August 29, 1980, show a WBC count of 6.3 x109/liter, including 1% bands, 51% polymorphonuclear leu

kocytes, 32% lymphocytes, and 12% monocytes. His hematocrit was 45%, and his platelet count was 140 x 109/liter. A

recent marrow specimen still showed large pink inclusion bodies in many cells of the granulocytic series. There has been noevidence of DIG during the course of his illness, and he is stillalive as of October 1980.

Light and Electron Microscopy

Smears of bone marrow were stained for aldehyde fuchsin

(19, 30) and dialyzed iron (17, 35). Cells from bone marrowaspirates were fixed in 1.5% glutaraldehyde in 0.1 M sodiumcacodylate buffer (pH 7.4) with 1% sucrose for 10 min at 4Â°,

washed twice in the same buffer with 7% sucrose, and shippedon ice by air freight from Washington to California. They wereincubated for peroxidase with 3,3'-diaminobenzidine and H2O2

at pH 7.6 by the method of Graham and Karnovsky (21),postfixed in 1% OsO4, and subsequently processed as previously described (7).

RESULTS

The question of whether the large round pink inclusions seenon smears in early neutrophilic precursor leukocytes (Fig. 1,inset) were exogenous particles (i.e., ingested RBC) or endogenous products of the leukocytes was easily resolved by electron microscopy. These peroxidase-positive inclusions (Fig. 1)

were each surrounded by a single unit membrane (Fig. 2). Ifthey were indeed ingested RBC, there would be 2 membranes,an inner one belonging to the RBC and an outer one belongingto the phagocytic vacuole.

The cells with the large pink inclusions which were up to 3mmin diameter also contained normal peroxidase-positive azur-

ophil (primary) granules (Fig. 3a), which in humans are usuallyno larger than 500 nm. Some cells contained many giantgranules (Fig. 1), whereas others contained only one (Fig. 3a).Although the content of these granules was not completelyhomogeneous, since there were both light and dark areas, wewere unable to resolve crystalline structures even on unstainedmaterial. This lack of crystalline structure differs from the casepreviously reported by Tulliez ef al. (39). When examined bylight microscopy, the giant granules showed no staining foraldehyde fuchsin (Fig. 3a, inset), nor dialyzed iron (not illustrated), whereas the adjacent normal azurophil granules werestained.

Having demonstrated that the giant inclusions were abnormalperoxidase-positive granules of endogenous origin, we wanted

to further delineate their formation. In previous studies ofnormal human bone marrow (5, 7), we had demonstrated thatthe peroxidase-positive azurophilic, or primary, granules are

formed by the usual pathway followed by secretory products(29) (RER â€”Â»vesicles -Â»Golgi complex â€”Â»storage granules);

i.e., in normal promyelocytes, the marker enzyme peroxidaseis synthesized and segregated within the RER and transportedto the Golgi region by small transitional vesicles. The enzymeis concentrated within the Golgi cisternae (see Ref. 4, diagram);then small Golgi-derived vesicles bud off and move to the

Table 1

Summary of initial blood data for 3 patients with AML

NeutrophilicseriesPatient123Age621652SexFFMHemato-crit

(%)251644Plateletcount(xlOVliter)3721Leukocytecount(xlOVliter)1.15.05.7Myeloblasts(%)2012Promyelocytes(%)151Myelo-cytes(%)101Meta-myelo-cytes(%)51Bands

+ poly-mor-

phonu-clearneutro

philicleuko

cytes10252Lymphocytes(%)904127Mono-cytes

Eosino-(%)

phils(%)73

3

4474 CANCER RESEARCH VOL. 40



Abnormal Granules in AML

peripheral cytoplasm where they aggregate to form the largermature azurophil granules. In the 3 bone marrow specimensdescribed here, PER and Golgi cisternae contained no reactionproduct for peroxidase (Fig. 3a). This lack of reaction productmay well reflect the procedures used, rather than a true absence of peroxidase, since 3 days elapsed between fixation ofthe specimens in Washington and incubation for the enzyme inCalifornia. However, it is interesting to note that other investigators have had difficulty demonstrating peroxidase in PERand Golgi cisternae in leukemic cells containing giant granules(30, 39). Clustered near the giant granules in our specimenswere large numbers of small vesicles containing high concentrations of peroxidase (Fig. 3b). Parmley ef al. (30) havesuggested that the giant granules in leukemic cells are formedby the fusion of these small dense vesicles as well as by thefusion of intact primary granules and concomitant entrapmentof cytoplasmic material. The 2 abnormal granule forms in Fig.2 (labeled ag) may exemplify the latter mechanism.

In addition to the prominent round inclusions, the cells from2 patients contained several other forms of abnormally largeperoxidase-positive granules (Fig. 4). These granules were

usually elongated, and some contained amorphous densities(Fig. 4a). Others contained a crystalloid structure (Fig. 4b),and yet others contained an obvious well-organized crystal

(Fig. 4c). The structure containing the crystal could be seen bylight microscopy on Wright's-stained smears and was azuro-

philic with a rounded middle and pointed ends (Fig. 4c, inset).All these forms seem to be variants of Auer bodies. The formillustrated in Fig. 4c was recently termed Phi body by Hankeret al. (22, 23). Since Auer originally described and illustratedorganelles of these shapes, we prefer to call them Auer bodies(see Ref. 3, Fig. 15).

The following observations in more mature neutrophils weremade. Although the giant granules were observed only inmyeloblasts, promyelocytes, and myelocytes, other abnormalities were seen in mature neutrophils. The most striking of thepathological abnormalities was the paucity and sometimescomplete absence of peroxidase-negative specific granules

(Fig. 5). This secondary granule population is usually formedduring the myelocyte stage and ordinarily constitutes two-thirds

of the total granule population of human neutrophils (7). Inaddition, there were prominent nuclear blebs, which have beenreported to be more common in leukemic cells (2), usuallyblasts, than in normal cells.

DISCUSSION

Abnormal forms of granules have been observed in the earlymyeloid cells of patients with acute leukemia by many investigators (1, 3, 5, 6, 8, 9, 11 -14, 24, 25, 36, 37, 41 ). The first

form recognized and the one best characterized to date is theAuer rod (3), which is present in the myeloblasts and promyelocytes of from 10 to 15% of patients with AML. This cytoplasmic inclusion is usually needle shaped, although Auer alsocommented on other forms, and is easily recognized on smearsprepared with Romanowsky-type stains because it is azuro-

philic (i.e., stains reddish purple); it also stains for peroxidaseand several other enzymes (6, 8, 12, 14, 36). Only 7 cases of"giant lysosome-like structures" (26), "oval cytosomes" (36),

or pseudo-Chediak-Higashi granules (18, 30, 39) in cells from

patients with acute leukemia have been previously studied.

One major difference between our 3 cases and others wasthe color of the giant granules as observed by light microscopy.In the earlier reports, the granules were described as purple(39), reddish purple (30), or azurophilic (18), whereas theinclusions that we observed were pink. Furthermore, theseinclusions did not stain for aldehyde-fuchsin, a stain for anionic

glycoconjugate, or dialyzed iron, a stain for carboxylated andsulfated acidic glycoconjugates. In contrast, the giant reddish-purple inclusions described by Parmley et al. (30) stainedstrongly for aldehyde-fuchsin. Since Dunn and Spicer (1 7) and

Parmley ef al. (31) have demonstrated that the azurophilia ofnormal azurophil (primary) granules is due to the presence ofsulfated glycoaminoglycans, we hypothesize that the giantnonazurophilic granules we observed lack acid mucosub-

stance(s). It should be mentioned that these giant round granules in acute leukemia stain for other constituents of normalazurophil granules, such as peroxidases (18, 26, 30, 39),naphthol AS-D chloroacetate esterase (26, 30), acid phospha-tase (18, 26), Sudan black (30), and periodic acid-Schiff (16,18, 26).

Schmalzl ef al. (36) have noted that Auer bodies also usuallystain for peroxidase, naphthol AS-D chloroacetate esterase,

acid phosphatase, and Sudan black and are thus closely related to normal azurophil granules. They point out, however,that not all Auer bodies in a particular specimen exhibit all ofthese staining properties. This variability of staining and presumably of granule content seems to apply to the pseudo-Chediak-Higashi anomaly as well.

The 2 investigations of giant granules in leukemic cells byelectron microscopy and peroxidase cytochemistry (30, 39)have both shown that: (a) the granules are bound by a singleunit of membrane, and the vast majority contain a matrixdensely reactive for peroxidase; (b) swarms of small peroxidase-positive vesicles are prominent in the cytoplasm of cells

containing such granules; and (c) peroxidase reaction productis not clearly demonstrable in RER or Golgi cisternae in thesecells. There are, however, some major discrepancies amongthe findings. In the study by Tulliez ef al. (39), no Auer bodieswere observed. The giant granules contained crystalline cores,seen best on unstained grids, and exhibited a periodicity different from that of the typical Auer bodies seen in acutepromyelocytic leukemia associated with DIG. The giant granules frequently contained heterogeneous membranous materialand amorphous nucleoids. In our specimens, the granule matrixwas relatively homogeneous and did not have a crystallinesubstructure. Furthermore, our patients showed no evidenceof DIG. Like Parmley ef al. (30), we observed Auer rods, butonly rarely were both abnormal granule types seen in the samecell.

None of the investigations thus far, including our own, hasprovided clear evidence of the mechanism by which the giantgranules are formed. Tulliez ef al. (39) have suggested thatthey are formed by fusion of azurophil granules. Parmley ef al.(30) have hypothesized that they are formed by 2 processes:(a) the fusion of small peroxidase-positive vesicles; and (b) the

fusion of intact primary granules and concomitant entrapmentof cytoplasmic material. It should be mentioned that thesenumerous small peroxidase-positive vesicles (Fig. 3b) were

also a prominent feature in cells forming Auer rods (Fig. 4) aspreviously observed (6, 12). Also, perhaps, the previouslymentioned absence of stainable glycoaminoglycans in these

DECEMBER 1980 4475




large granules may provide a clue to their formation. Eventhough the factors responsible for the concentration of enzymes and their packaging into normal azurophil granules arestill obscure, we believe that they may be similar to the factorssuggested by Reggio and Palade (34) to explain the concentration of enzymes in zymogen granules of the pancreaticexocrine cell. These investigators proposed that the concentration was not achieved by membrane ion pumps in the condensing vacuoles but possibly occurred as a result of theosmotic effects of a sulfated polyanion, a peptidoglycan. Hy-

pothetically, this polyanion together with secretory proteinsthat are predominantly cationic could cause a reduction in theosmotic activity within the condensing vacuoles; the resultantefflux of water would bring about concentration. Additionaldata on normal human leukocytes are needed before abnormalities in the condensation of azurophil granules in leukemiccells can be further analyzed.

The formation of the megagranules in the hereditary Chediak-

Higashi anomaly has been carefully studied by Davis et al. (15).Promyelocytes from persons and animals with this trait lacknormal azurophil (primary) granules and, instead, contain enlarged pleomorphic granules, which may resemble azurophilgranules to varying degrees. These unusual granules are notpresent in the blasts and become apparent in promyelocytesafter the appearance of the precursor vesicles derived fromGolgi cisternae. The precursor vesicles undergo a series offusions leading to the formation of the megagranules. In humans, the characteristic crystalloid does not seem to form, andthe granule matrix develops a reticulated array of membrane-

like material. These granules also contain peroxidase and stainwith Sudan black. On Wright's staining of smears, they may

stain reddish purple, blue, purple, or sometimes pink (10, 15,32, 39, 40). The second stage of granulopoiesis, which givesrise to the peroxidase-negative specific (secondary) granule,

is unaffected; and a full complement of these granules ispresent in mature neutrophils. No normal azurophil granulesare present. It was recently demonstrated that some form ofthe megagranules also contains lactoferrin, indicating somelimited fusion with specific granules (32). The defects underlying the abnormal granule fusion in Chediak-Higashi leuko

cytes have not yet been defined although 2 possibilities, alteration of intracellular cyclic nucleotides and dysfunction ofmicrotubules and granule membranes, have been investigated(see Ref. 28). It should also be remembered that, in hereditaryChediak-Higashi syndrome, other hematopoietic cell types

contain megagranules as do many other nonhematopoieticcells, such as melanocytes, renal tubular epithelial cells, andcells of gastric mucosa, pancreas, thyroid, and neural tissues(15).

The relative paucity of peroxidase-negative specific granules

in the more mature segmented neutrophils warrants comment.It appears that cytoplasmic development has ceased after thepromyelocytic stage, whereas nuclear maturation has progressed in a fairly normal fashion, a phenomenon termed"maturation anarchy" by Bessis (8). He attributes this anarchyto failure of the "program" of cell maturation, which is char

acteristic of acute leukemias. The absence of certain normalorganelles from mature neutrophils of patients with acute leukemia has been previously documented by electron microscopy and cytochemistry (4-6) as well as by immunofluores-cence methods. For example, we have demonstrated the ab

sence of specific (secondary) granules in neutrophils frompatients with AML (4, 6), and Odeburg ef al. (27) and Rauschet al. (33) have observed the absence or marked decrease oflactoferrin, a substance found exclusively in the specific granule population. These 2 characteristics may permit recognitionof mature neutrophils derived from leukemic clones. We haveobserved these abnormal neutrophils quite frequently in acutemyeloid leukemia with maturation, i.e., the M2 variety, as wellas in certain cases of hematopoietic dysplasia. We plan tofollow patients with the latter condition to see whether thepresence of such neutrophils correlates with the eventual appearance of acute leukemia.

Several of the patients in previously published case reportshad acute promyelocytic leukemia with clinical and laboratoryfindings suggestive of DIG (30, 36, 39). In this study, we havedescribed 3 more patients, all with well-differentiated AML,

whose neutrophilic promyelocytes and myelocytes containedgiant pink granules. Unlike the patients previously studied, theyshowed no definite evidence of DIG.

We have no explanation at present for these different clinicalfindings. In addition, the survival of more than 3 years in Case3 is atypical of AML and deserves further comment. Despitepersistent bone marrow morphology consistent with the diagnosis of AML and the presence of megagranules in manyneutrophilic precursor cells, the peripheral blood remains relatively normal. This case may therefore be similar to the onepreviously described by Branda et al. (11) in which there wasan indolent course as well as a marked dichotomy between theblastic morphology of the bone marrow and a relatively normalblood picture. In addition, the blasts of their patient containedprominent vacuoles and frequent Auer rods. Furthermore,when the bone marrow was cultured, the leukemic cells differentiated to form abnormal primary granules which appeared torupture and cause cytolysis of these cells. On the basis of suchobservations, they suggested that the indolent clinical coursemight be related to blast destruction within the marrow andrecommended less intensive chemotherapy. It is reasonable topostulate that our Case 3 has intramedullary destruction ofleukemic cells.

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Fig. 1. Electron micrograph of a neutrophilic promyelocyte from bone marrow, stained for peroxidase. The cytoplasm contains at least 6 abnormally large roundgranules (arrows), the pseudo-Chediak-Higashi granules, in addition to normal-size azurophil granules (ag). Very small peroxidase-positive vesicles (ve) are presentnear some of the large granules, nu. nucleolus: m, mitochondria: er, endoplasmic reticulum. x 17.500. Inset, light micrograph of a Wright-Giemsa-stained bonemarrow preparation showing 2 large pink inclusions in a promyelocyte. N, nucleus, x 3,000.

Fig. 2. Higher magnification of part of a neutrophilic promyelocyte from bone marrow, stained for peroxidase. Pseudo-Chediak-Higashi granule (p-CH) is limitedby a single unit membrane (me), similar in thickness to the membranes of the adjacent azurophil granules (ag). Although the matrix of the pseudo-Chediak-Higashigranule is not entirely uniform, no crystalline structure could be detected even on unstained specimens, er, endoplasmic reticulum. x 67.000.

Fig. 3. Electron micrographs of neutrophilic promyelocytes from bone marrow, stained for peroxidase. In a, note the abnormally large (about 1 firn) roundperoxidase-positive granule (p-CH) in addition to many smailer normal-size (about 500 nm) azurophil granules (ag). The perinuclear cisterna (pn), endoplasmicreticulum (er), and Golgi cisternae (Gc) are not reactive for peroxidase. x 16.000. Inset, light micrograph of an aldehyde fuchsin-stained preparation showing normalreactive granules (a-0 and 2 large unstained megagranules (arrows), x 1.300. In b, the cluster of small reactive vesicles (ve) can be seen to better advantage. Thesevesicles probably fuse (ve') to form the larger granules (p-CH). Other azurophil granules (ag) are of normal size, m, mitochondria; N. nucleus, x 40.000.

Fig. 4. Portions of the cytoplasm of myeloblasts and promyelocytes, illustrating several other abnormal forms of peroxidase-positive granules. The granules in aand o may represent immature forms present before full crystallization of granule contents has occurred, as in the elongated form in c. Numerous small vesicles (ve)are present near these forming granules, ag. normal azurophil granules; m. mitochondria; er, endoplasmic reticulum; pn, perinuclear cisterna, a, x 22.000; b.x 39.000; c, x 27.000. Inset, light micrograph showing an elongated granule (arrow) like the one in c. Such granules were azurophiHc. x 1,600.

Fig. 5. Electron micrograph of a more mature neutrophil from bone marrow, stained for peroxidase. Although the nuclear chromatin is condensed and segmented(s) and the cell is presumably a mature polymorphonuclear neutrophilic leukocyte, its cytoplasm contains very few peroxidase-negative specific granules (sq). In anormal mature polymorphonuclear neutrophilic leukocyte, about two-thirds of the granules would be peroxidase-negative specific granules, and about one-thirdwould be peroxidase-positive azurophils (ag). Some of these peroxidase-positive granules are also abnormal in that they tend to be larger and less homogeneous incontent than normal azurophilic granules. A nuclear bleb (D) is also present. A/I and N?. nuclear segments; m, mitochondria, x 20.000.

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1980;40:4473-4481. Cancer Res William A. Dittman, Robert J. Kramer and Dorothy F. Bainton LeukemiaPink Pseudo-Chediak-Higashi Granules in Acute Myelogenous Electron Microscopic and Peroxidase Cytochemical Analysis of

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