690

between Hb level and increase in COHb after smoking,it is possible that the lower Hb levels of the women(mean 12-8 g. per 100 ml. compared with 14.4 g. per100 ml. for the men) together with their smallerblood volume accounts for their greater COHbincreases after smoking. This relation between sex,Hb level, and COHb uptake has not as far as we

know been reported previously. It is certainly notgenerally appreciated. It may have especial implica-tions for those women who smoke in pregnancy andwho are anaemic.

The findings indicate that besides variation in tarand nicotine yield, commercial brands of cigarettevary greatly in their CO yield. Direct measurementof the CO concentration of mainstream smoke

presents no more of a technical problem than themeasurement of its nicotine or tar content. Indeed,it is likely that the tobacco manufacturers alreadyknow the CO yield of their cigarettes and cigars.Because the health implications are probably as

important as is the case with tar and nicotine, theCO yield of all commercially available cigarettesshould be published together with their tar andnicotine yield. Furthermore, this should be donesoon.

We thank Upendra Patel of the Addiction Research Unitfor checking the statistical calculations, Pauline Beattie forsecretarial help, and the Medical Research Council, the

Department of Health and Social Security, and the JointResearch Board of St. Bartholomew’s Hospital for financialsupport.

Requests for reprints should be addressed to M. A. H. R.

REFERENCES

1. Lancet, 1973, i, 874.2. Wynder, E. L. in Second World Conference on Smoking and

Health (edited by R. G. Richardson); p. 197. London, 1971.3. Royal College of Physicians. Smoking and Health Now; p. 134.

London, 1971.4. Surgeon General. The Health Consequences of Smoking. U.S.

Department of Health, Education and Welfare, 1972.5. Astrup, P. Br. med. J. 1972, iv, 447.6. Astrup, P., Trolle, D., Olsen, H. M., Kjeldsen, K. Lancet, 1972, ii,

1220.

7. Wald, N, Howard, S., Smith, P. G., Kjeldsen, K. Br. med. J. 1973,i, 761.

8. Armstrong, H. E. ibid. 1922, i, 992.9. Russell, M. A. H., Cole, P. V., Brown, E. Lancet, 1973, i, 576.

" The Malthusian nightmare, that mass starvation will be theultimate population control, is only one, and that the mostdistant, of several specters. Beyond the immediate question ofwhether this year’s crops will produce enough food to avoidmajor price disturbances, political instabilities, and famines,there is concern that the present alarms and scarcities mayreflect not just last year’s bad weather, but a fundamentaldeterioration in the world food situation.. Already there arethose who foresee a period of food scarcity, in which those withfood to sell will have a useful political weapon in their hands.Governments of developing countries will find this year that thesoaring prices of food grains and freight rates have driven theirimported food bills up by 60 percent or roughly$2 billion, adrain on foreign reserves which, should it continue, threatensto retard economic development and make the gap between richnations and poor grow faster still. Much besides the threat offamine, therefore, hinges upon the ability of developing countriesto make crop yields grow faster than people."-NICHOLAS WADE,Science, 1973, 181, 634.

DETECTION OF HEPATITIS-B ANTIGEN

BY RADIOIMMUNOASSAY IN CHRONIC

LIVER DISEASE AND HEPATOCELLULAR

CARCINOMA IN GREAT BRITAIN

W. D. REED A. L. W. F. EDDLESTON

R. B. STERN ROGER WILLIAMS

Liver Unit, King’s College Hospital and Medical School,London SE5

A. J. ZUCKERMAN A. BOWES

PAMELA M. EARL

Hepatitis Research Unit, Department of Microbiology,London School of Hygiene and Tropical Medicine

Summary In a series of 264 patients with chronicliver disease or hepatocellular car-

cinoma, a sensitive radioimmunoassay techniqueshowed 37 cases to be positive for HBAg. 20 of these37 patients were HBAg negative by immunodiffusionand electrophoresis. In active chronic hepatitis 18%of the 94 patients were HBAg positive. Comparisonof these cases with HBAg-negative cases showed ahigher frequency of HBAg in males and those bornoverseas. There were no significant differencesbetween HBAg positive and negative patients forclinical manifestations, immunoglobulins, autoanti-bodies (after allowing for differences in sex ratio),cell-mediated immunity, or prognosis, and in somecases the progression of tissue damage in this con-dition may be unrelated to the continued presenceof circulating HBAg. HBAg was found in only 1

of 45 patients with primary biliary cirrhosis, andcoincidental infection following blood-transfusion wasa possibility in half of the HBAg-positive cases of

cryptogenic cirrhosis. However, blood-transfusionscould be implicated in only two of the six HBAg-positive patients with alcoholic cirrhosis. In primaryhepatocellular carcinoma, 23 % of the 38 patients werepositive, the frequency being higher in those bornoverseas (66% compared with 15%), showing that thereported geographical variation in this associationcannot be due entirely to differences in techniquefor detecting HBAg.

Introduction

AN association between viral hepatitis and activechronic hepatitis was recognised in one of the earliestreports of this syndrome, in which it was called " activechronic infectious hepatitis ".1 However, the sub-

sequent observation of a positive lupus-erythematosus(L.E.) cell phenomenon,2 2 hypergammaglobulin2emia 3and the presence of autoantibodies in the serum I

led to the concept of a primary autoimmune patho-genesis.5 More recently, the detection of hepatitis-Bantigen (HBAg) in the serum of some patients 1-1has renewed interest in a viral etiology. The pro-portion of HBAg-positive cases varies from 51 % in

Italy 9 to 3 % in Australia.1O In Great Britain, HBAgwas present in 8 % of patients with active chronichepatitis but all of the 6 positive cases had comefrom abroad." A pronounced geographical variationin the frequency of HBAg has also been reported inhepatocellular carcinoma.12,13

691

In the present study we have used a sensitive

radioimmunoassay technique to detect circulatingHBAg in a series of 264 patients with active chronichepatitis and certain other chronic liver diseases aswell as hepatocellular carcinoma. In many of thecases serial samples have been tested at intervalsfor up to four years. Since several reports havestressed differences in age, sex, prognosis, serology,and extrahepatic manifestations, between HBAgpositive and negative forms of active chronic hepa-titis,8-11 we have also investigated this aspect.

Methods

The radioimmunoassay used to detect HBAg 14 is basedon the solid-phase technique of Catt and Treager,15 usingpolypropylene tubes coated with hepatitis-B antibody(Abbott). The guineapig antibody possessed anti-a, anti-d, and anti-y specificities. Three sera known to be positivefor HBAg and seven sera known to be negative were testedas controls with each batch of a hundred tests. The

specificity of the present technique was confirmed by thedemonstration of HBAg particles by electron microscopyin several of the samples positive only by radioimmuno-assay. Furthermore, radioimmunoassay was negative ina patient whose serum gave a false-positive result on

immunodiffusion (using commercial antisera raised in

animals) due to the presence of antibodies induced byprevious immunisation with horse antitetanus serum. Inanother patient, who was repeatedly HBAg positive byradioimmunoassay alone, the serum became negativeafter infusion of a preparation of human immunoglobulincontaining specific antibody to HBAg.Routine testing for both antigen and antibody was also

performed using immunodiffusion and counterelectro-

phoresis.11 A total of 621 sera were tested under code,including 352 from patients with active chronic hepatitis.

Results

Active Chronic Hepatitis17 (18%) of the 94 patients had HBAg detected

in their serum by radioimmunoassay on at least oneoccasion. 7 of the 17 patients positive for HBAgby radioimmunoassay, were also positive by immuno-diffusion and electrophoresis, of the other 10 antigen-aemia was persistent in 3 and transient in 7. 2 others,in whom the serum was highly anticomplimentary,became positive for HBAg (by immunodiffusionand complement fixation) after heating at 85 °C. forthirty minutes, a procedure which is said to dissociateimmune complexesY

In 10 of the 17 positive patients, HBAg was presentin all of the 35 sera tested over intervals from one toeighteen months. In the other 7 patients, only onespecimen from each patient was positive (7 out of40 sera tested). In 5 of those with transient anti-genxmia, HBAg was first detected some eleven to

forty-one months after starting serial testing, andone of these patients had received a blood-transfusionin the preceding period. The remaining 2 patientswere positive when first tested and consistentlynegative thereafter; 1 had a fifteen-year history ofchronic liver disease, but in the other, detection ofHBAg coincided with the onset of the illness and

was followed by the appearance of hepatitis-B anti-body in the serum, which was still present at the

time of his death two years later. This was the

only patient in whom antibody was detected.

TABLE I-COMPARISON OF CLINICAL, EPIDEMIOLOGICAL, AND

BIOCHEMICAL FEATURES IN HEPATITIS-B-ANTIGEN POSITIVE AND

NEGATIVE ACTIVE CHRONIC HEPATITIS

Statistical comparisons were done by the x2 test with Yates’ correctionor Fisher’s exact test where applicable.

Results are expressed as means.D.j except for serum biochemistrywhere the standard error is shown.

*, significantly different results (P < 001).

Statistical comparison of HBAg positive and

negative cases, showed that antigen was more oftenpresent in males (P<0&deg;O1) and in patients bornoutside Great Britain (P<0-01), most of the latter

coming from Mediterranean countries and the MiddleEast. There were no differences in age, type of

onset, history of exposure, or previous attacks of

hepatitis (table I). The numbers with jaundice (70%and 83 % in HBAg positive and negative cases,

respectively) and ascites (47 % and 49%, respectively)were similar. There was also no apparent differencein histological appearance of the liver in the two

groups, and cirrhosis had developed in 82 % and

84% of the positive and negative cases, respectively.Extrahepatic manifestations were more often presentin those with HBAg (59% as compared with 39%),although the difference was not statistically signifi-cant. There were no other significant differencesbetween the two groups for serum biochemistry,immunoglobulin levels, and the presence of auto-

antibodies (table 11). Antinuclear factor was detectedless often in HBAg-positive cases. However, theratio of males to females in the two groups was not

comparable, and an analysis of HBAg-negative cases

TABLE II-COMPARISON OF SERUM IMMUNOGLOBULINS, AUTOANTI-

BODIES, AND L.E. CELLS IN HEPATITIS-B-ANTIGEN POSITIVE ANDNEGATIVE ACTIVE CHRONIC HEPATITIS

Statistical techniques as for table i. None of the differences werestatistically significant. High titre=positive at dilutions > 1/80.

692

TABLE III-HEPATITIS-B-ANTIGEN DETECTED BY RADIOIMMUNOASSAY IN CHRONIC LIVER DISEASE AND HEPATOCELLULAR CARCINOMA

i I

* Included in this number are 2 patients with active chronic hepatitis, one with primary biliary cirrhosis, 8 with cryptogenic cirrhosis, and 8 withalcoholic cirrhosis, who are listed twice.

showed that the frequency of a high titre of anti-nuclear factor was significantly lower (p<0’002) inmales (9%) than in females (41%). When males

alone, who constituted a greater proportion of theHBAg-positive group, were examined the frequencyof antinuclear factor was the same for the two groups(table 11). Smooth-muscle antibody was different inthat high titres were found more often in the HBAg-positive cases. This difference was even more pro-nounced when males were considered separately(23% compared with 4%), but with the small numbersinvolved the difference was not statistically significant.Results of the leucocyte-migration test using liver-

specific lipoprotein as antigen 18 were the same in bothgroups, with 53 % and 51 % of the HBAg positiveand negative cases, respectively, showing inhibitionof migration.The difference in overall survival at six years-

62% for the HBAg-positive group and 80% for theHBAg-negative group-was not significant andneither was the difference in mean survival for the

patients who died (4-3 years, range 0.5-8.0 for theHBAg-positive cases; 5-2 years, range 05-18 forthe HBAg-negative group). The results of treatmentwith prednisone and/or azathioprine in the HBAg-positive group appeared no different from those inpatients without HBAg.The group containing patients who were trans-

iently positive included more females, fewer foreignpatients, had higher frequency of autoantibodies inhigh titre, and had higher frequency of extrahepaticmanifestation than the group with persistent HBAgin their sera. But only the difference in the frequencyof extrahepatic manifestation was significant (P<0&middot;05).

Primary Biliary Cirrhosis, Cryptogenic Cirrhosis, andAlcoholic CirrhosisHBAg was detected in only 1 of the 45 patients

with primary biliary cirrhosis, a fifty-nine-year-oldmale who was born and had lived most of his lifein India. Only the last of the 4 sera collected overa period of forty-eight months was positive, and thisfollowed transfusion with 11 units of blood duringthe previous three years for a series of hip operations.

10 (26%) of 39 patients with cryptogenic cirrhosiswere HBAg positive. All of these were male; noneof the 9 females tested gave positive results. 5 ofthe 10 positive cases had previously received a blood-transfusion. The diagnosis of cryptogenic cirrhosishad been made in these patients because there wasno known cause for the disease and the lesion

appeared inactive both biochemically and on histo-logical examination.

6 (9%) of the 67 patients with alcoholic cirrhosishad HBAg, all were male and born in Britain

(table ill). Several of these patients had presentedwith bleeding oesophageal varices, but only 2 of the6 positive cases had been transfused before the detec-tion of HBAg.

Hepatocellular CarcinomaThis diagnosis had been confirmed histologically

in each of the 38 patients, either by liver biopsy orat necropsy. In 9 (24%) HBAg was detected byradioimmunoassay. The antigen was found moreoften in patients born outside Great Britain

(P=<0.01). Males (32%) were positive more oftenthan females (8%), although this difference was notstatistically significant (table III).The frequency of an underlying cirrhosis was

similar in HBAg positive (78%) and negative cases(55%), but the type of cirrhosis differed in the twogroups. In the positive group there was a higherproportion of cryptogenic cirrhosis (71% comparedwith 25 % for the HBAg negative cases) and a smallernumber with alcoholic cirrhosis (14% compared with44% in the negative group). There were 2 patientsin the series with an underlying active chronic

hepatitis (and cirrhosis); one was persistently HBAgpositive and the other negative on repeated testing.The frequency of a-fetoprotein in the serum of

26 patients was considerably higher (P=<0.05) in

HBAg-positive (71%) than in HBAg-negative cases

(22%).

Discussion

The order of sensitivity for the radioimmunoassaytechnique is said to be 2000 to 10,000 times greaterthan for immunodiffusion and electrophoresis, andup to 500 times greater than for complementfixation 14 According to Ling 19 it will detect 3 to 8times more positive sera. In the present series thenumber detected by radioimmunoassay was 22 timesgreater than by the other techniques used. Althoughthe frequency of HBAg in the general populationof Great Britain as determined by radioimmunoassayis not known, it is unlikely to be higher than the 1 0,’, ’’’found by this method in volunteer blood-donors inChicago."The frequency of HBAg (13 %) in our British-born

patients with active chronic hepatitis is comparablewith figures reported from areas where the populationhas a similar genetic background, such as Australia 10

and the U.S.A.2O Higher figures have been reportedfrom some centres in America but this may be

related to an increased frequency of parenteral

693

exposure in such areas.8 In our series the 60%frequency of HBAg in those born outside Great

Britain is similar to the 51 % reported from Italy. 9

Thus, differences in the sensitivity of the techniqueused cannot be implicated as the sole cause of suchgeographical variations.Previous reports have emphasised the clinical andserological differences between HBAg positive andnegative cases of active chronic hepatitis. HBAg-positive patients have been described both as beingyounger 9 and older 11 than HBAg-negative patients.An acute onset and previous exposure to hepatitisare said to be more common,8.9 and extrahepaticmanifestations are said to be more unusual amongHBAg-positive cases. Gammaglobulin levels werelower,8,11 autoantibodies absent from the serum,8,10,11,21biochemical abnormalities less severe, and prognosisbetter 11 among HBAg-positive patients. However,apart from finding an increased number of malesamong HBAg-positive patients, we have been unableto confirm any of these observations. Antinuclearfactor was less common in HBAg-positive cases, butthis difference was due to an increased number ofmales in this group, the frequency of antinuclearfactor being significantly lower in males of the antigen-negative group.The reasons for the increased susceptibility of

males and certain racial groups to chronic infectionwith HBAg are not known. It applies to both

healthy carriers and patients with liver disease.22In a study of HBAg-positive patients with hepatomafrom Uganda no generalised defect in either cellularor humoral immune responses was found,13 but thepossibility of a specific immune defect remains. Sub-

jects with persistent HBAg rarely have detectablehumoral antibody. The continued presence of cir-

culating HBAg in some cases of active chronic

hepatitis may be unrelated to any role the hepatitis-Bvirus had in initiating the disease, these cases differingonly in their capacity to eliminate the infecting virus.The demonstration of sensitisation to a liver-specificantigen in HBAg-positive as well as in HBAg-negative cases lends further support to the proposalthat a common autoimmune process is involved intheir pathogenesis.Why some patients with active chronic hepatitis

have transient HBAg in their serum and others havepersistent HBAg is uncertain. The altered sex ratioin those transiently positive was of interest. Althoughcoincidental infection is always a possibility inpatients liable to receive blood-transfusions, in onlyone of the transiently positive cases was there sucha history. Furthermore, in another’ transientlypositive case the detection of HBAg at the onset ofthe illness and the persistent presence of antibodysubsequently, strongly suggest that the disease wasxtiologically related to infection with hepatitis-Bvirus.The relation of hepatitis-B virus infection to

primary biliary cirrhosis has been the subject of

controversy. Krohn et a123 reported HBAg particleson electron micropsy in 11 of 12 patients with primarybiliary cirrhosis, sera from 9 of them also beingpositive by high-voltage electrophoresis. However,the specificity of high-voltage techniques has been

questioned 20 and recently false-positive particles onelectron microscopy have been described in primarybiliary cirrhosis.24 Using a radioimmunoassay tech-nique, Doniach et al.25 found HBAg in 13 % of 69cases of primary biliary cirrhosis; but the incidencein 60 patients with Hashimoto’s thyroiditis was

similar. Sama,24 who found that 5 of 24 cases of

primary biliary cirrhosis were positive by radio-immunoassay and electron microscopy, suggested thatimpaired immune mechanisms, known to be presentin the later stages of this condition, may predisposepatients to HBAg carriage. The one positive casein the present series, in which conversion followedmultiple blood-transfusions some four years afterthe onset, supports the view of Maddrey et a1.26that the acquisition of HBAg in primary biliarycirrhosis is unrelated to the disease process.

Coincidental infection via blood-tranfusions was

also the probable explanation for the presence of

HBAg in half of the patients with cryptogeniccirrhosis. However, only 2 of the 6 HBAg-positivecases with alcoholic cirrhosis had previously receiveda blood-transfusion. The demonstration of lympho-cyte transformation in response to HBAg-rich serumin all of 11 patients with alcoholic cirrhosis has alsobeen interpreted as evidence implicating hepatitis-Bvirus in the pathogenesis of this disease.27 None ofthe patients had circulating HBAg or anti-HBAg asmeasured by a sensitive haemagglutination-inhibitiontechnique. However, in a similar study in which theleucocyte-migration test was used to detect cell-mediated immunity to HBAg, no positive responseswere obtained.28There seems to be little doubt that the frequency

of HBAg in hepatocellular carcinoma varies accordingto locality. Prince 12 found 2 of 52 cases in New Yorkto be HBAg positive, compared with 80% of 55cases in Taiwan, where the carrier-rate in the normalpopulation is 14% .21 In a recent survey in Ugandausing radioimmunoassay, 55% of 49 cases were

HBAg positive compared with 3% of controls .13

Although part of these discrepancies may be relatedto variations in sensitivity of the techniques used/&deg;in the present series there was a significant differencein the frequency of HBAg between foreign andBritish-born patients. The positive correlation witha-fetoprotein in the serum and HBAg is similar tothat found in Uganda,31 but does not accord with thefindings in Taiwan.29 Finally, the fact that two ofour positive hepatoma cases did not have an under-lying cirrhosis shows that the association with

hepatitis-B virus cannot be simply explained by theoccurrence of malignant change in the nodules ofan HBAg-positive cirrhosis.We thank Dr Deborah Doniach and Dr D. Tee for serum

autoantibody and immunoglobulin determinations respectively;and the World Health Organisation, the Medical ResearchCouncil, the Wellcome Trust, and Pfizer Ltd. for their support.W. D. R. was in receipt of an F. A. Hadley travellingscholarship from the University of Western Australia.

Requests for reprints should be addressed to W. D. R.

REFERENCES

1. Saint, E. G., King, W. E., Joske, R. A., Finckh, E. S. Australas.Ann. Med. 1953, 2, 113.

2. Joske, R. A., King, W. E. Lancet, 1955, ii, 477.

694

DR REED AND OTHERS: REFERENCES&mdash;continued

3. Waldenstr&ouml;m, J. Leber, Blutproteine und Nahrungseiweiss Stoff-wechsel. Sonderband xv, p. 8. Bad Kissingen, 1950.

4. Johnson, G. D., Holborow, E. J., Glynn, L. E. Lancet, 1965, ii,878.

5. Mackay, I. R., Taft, L. I., Cowling, D. C. ibid. 1956, ii, 1323.6. Wright, R., McCollum, R. W., Klatskin, G. ibid. 1969, ii, 117.7. Gitnick, G. L., Gleich, G. J., Schonfield, L. J., Baggenstoss, A. H.,

Sutnick, A. I., Blumberg, B. S., London, W. T., Summerskill,W. H. J. ibid. p. 285.

8. Bulkley, B. H., Heizer, W. D.; Goldfinger, S. E., Isselbacher, K. J.,Shulman, N. R. ibid. 1970, ii, 1323.

9. Bianchi, P., Bianchi Porro, C., Coltorti, M., Dardanoni, L., DelVecchio Blanco, C., Fagiolo, U., Farini, R., Menozzi, I., Naccarato,R., Pagliaro, L., Spano, C., Verme, G. Gastroenterology, 1972, 63,482.

10. Cooksley, W. G. E., Powell, L. W., Mistilis, S. P., Olsen, G.,Mathews, J. D., Mackay, I. R. Aust. N.Z. J. Med. 1972, 3, 261.

11. Sherlock, S., Fox, R. A., Niazi, S. P., Scheuer, P. J. Lancet, 1970, i,1243.

12. Prince, A. M., Leblanc, L., Krohn, K., Masseyeff, R., Alpert, M. E.ibid. 1970, ii, 717.

13. Primack, A., Vogel, C. L., Barker, L. F. Br. med. J. 1973, i, 16.14. Tech. Rep. Ser. Wld Hlth Org. 1973, no. 512.15. Catt, K., Treager, G. W. Science, 1967, 158, 1570.16. Memorandum. Bull. Wld Hlth Org. 1970, 42, 957.17. Ross, C. A. C., Pringle, R. C. Lancet, 1971, ii, 434.18. Miller, J., Smith, M. G. M., Mitchell, C. G., Reed, W. D., Eddleston,

A. L. W. F., Williams, R. ibid. 1972, ii, 296.19. Ling, C. M., Overby, L. R. J. Immun. 1972, 109, 834.20. Kaplan, M. M., Grady, G. Lancet, 1971, i, 159.21. Wright, R. Vox sang. 1970, 19, 320.22. Cossart, Y. E. Br. med. Bull. 1972, 28, 156.23. Krohn, K., Finlayson, N. D. C., Jokelainen, P. T., Anderson, K. E.,

Prince, A. M. Lancet, 1970, ii, 379.24. Sama, S., Benz, W., Aach, R., Hacker, E., Kaplan, M. ibid. 1973,

i, 14.25. Doniach, D., del Prete, S., Dane, D. S., Walsh, J. H. Can. med. Ass.

J. 1972, 106, 513.26. Maddrey, W. C., Saito, S., Shulman, N. R., Klatskin, G. Ann.

intern. Med. 1972, 76, 705.27. Pettigrew, N. M., Goudie, R. B., Russell, R. I., Chaudhuri, A. K. R.

Lancet, 1972, ii, 724.28. Gerber, M. J., Vittal, S. B., Clowdus, B. F. ibid. p. 1034.29. Tong, M. J., Sun, S. C. M., Schaefer, B. T., Chang, N. I., Lo, K. J.,

Peters, R. L. Ann. intern. Med. 1971, 75, 687.30. Simons, M. J. Lancet, 1972, ii, 1089.31. Vogel, C. L., Anthony, P. P., Mody, N., Barker, L. F. ibid. 1970, ii,

621.

POST-TRANSFUSION HEPATITIS IN

RECIPIENTS OF BLOOD SCREENED BY

NEWER ASSAYS

RONALD L. KORETZ DONALD R. KLAHS

SUSAN RITMAN KARLA H. DAMUS

GARY L. GITNICKDivision of Gastroenterology, University of California at

Los Angeles, Center for the Health Sciences,Los Angeles, California 90024, U.S.A.

Summary To assess the efficacy of screeningblood with sensitive hepatitis-B-

antigen (HBAg) assays to reduce the incidence of

post-transfusion hepatitis, 6399 units of blood,previously found to be negative for HBAg whentested by counterelectrophoresis (C.E.P.), were re-

screened for HBAg by red-blood-cell agglutination(R.C.A.) and radioimmunoassay (R.I.A.). Of these units,2&middot;2% were positive. The recipients of these HBAg-positive units were followed, as were 19 ageand sex matched controls who received the samenumber of units of HBAg-negative blood. Among63 recipients of HBAg-positive blood (C.E.P. negative,R.I.A. or R.C.A. positive), 24 (38%) developed hepatitis.In addition, 5 others developed hepatitis-B antibody(HBAb) (8%), and 1 other became an HBAg carrier

without evidence of hepatitis. 1 control patientdeveloped HBAb without hepatitis, and 9 (47%)control recipients developed hepatitis. The R.I.A.

test was frequently associated with false-positiveresults (no hepatitis, HBAg carrier state, or HBAbseroconversion resulting), and was not more sensitivethat R.c.A. in predicting the occurrence of hepatitisor seroconversion. When each test (R.C.A. or R.I.A.)was compared to the clinical outcome, a positiveR.C.A. test correlated more closely with the develop-ment of hepatitis or seroconversion (94%).

Introduction

POST-TRANSFUSION hepatitis (P.T.H.) continuesto be an important complication of the use of bloodproducts, and concern over its development maycause doctors to withhold this form of therapy. Elimi-nation of blood products positive for hepatitis Bantigen (HBAg) has resulted in an apparent de-creased frequency of P.T.H.1 The common screeningmethods used in blood-banks throughout the worldare agar-gel immunodiffusion and counterelectro-

phoresis (C.E.P.). Now more sensitive assays havebeen developed, including complement fixation, red-blood-cell agglutination (R.c.A.), and radioimmuno-

assay (R.I.A.).2-4We have found that of 21 patients with P.T.H. 5

had received blood which was HBAg positive bythe more sensitive techniques having been negativeby C.E.p.5 We have now done a prospective controlledstudy in an attempt to answer the following questions:what is the frequency of " low titre " HBAg-positive(HBAg negative by C.E.P., but HBAg positive byR.C.A. or R.I.A.) blood products; do recipients of suchblood develop hepatitis; and are more sensitive HBAgscreening techniques useful in preventing P.T.H. ?

Patients and Methods

Before transfusion all units of blood at the U.C.L.A.Center for the Health Sciences were screened by C.E.P.

and were found to be negative. Since August, 1972,a small sample of each blood unit has been retestedby R.C.A. and R.I.A. at least 2 weeks after the unit wastransfused. The recipients of positive units were askedto participate in the study, and those that agreed werefollowed up for 9 months with biweekly determinationof their serum-glutamic-oxaloacetic-transaminase (S.G.O.T.)and serum-glutamic-pyruvic-transaminase (S.G.P.T.) levelsas well as assays for HBAg and antibody to HBAg(HBAb).As each patient entered the study, a matched control

patient was sought who was of the same sex and raceand within five years in age. This control was approxi-mately matched as to number of units received.No patient or control was excluded because of under-

lying chronic liver disease. However, patients who hadactive hepatitis at the time of transfusion, who hadreceived transfusions within 6 months, or who had an

underlying disease that would clearly make themcandidates for further maintenance blood-transfusionswere followed without controls. No such patients wereaccepted as controls.

R.C.A. testing for HBAg was performed by a previouslydescribed method.6,7 The Abbott solid-state radioimmuno-assay was used for the R.I.A. testing.8 HBAb was deter-mined by hai-magglutination9 and counterelectrophoresis.1oThe criterion for P.T.H. was an increase of the S.G.O.T-