Hepatitis b e antigen and antibody: detection by radioimmunoassay in chimpanzees during experimental hepatitis b

Hepatitis B e antigen (HBeAg) and its antibody (anti-HBe) were evaluated using a sensitive radioimmunoassay (RIA) in weekly serum samples obtained from nine chimpanzees experimentally infected with hepatitis B virus (HBV). In two chimpanzees with HBV infection with detectable hepatitis B surface antigen (HBsAg) for less than five weeks, and in one chimpanzee with documented HBV infection with no detectable HBsAg, HBeAg was not detected; in all three, anti-HBe became detectable early in the infection. In six chimpanzees in which HBsAg was detected for 16 weeks or longer, HBeAg was detected early in the infection; in five, anti-HBe became detectable and HBeAg unde-tectable prior to the clearance of HBsAg. The sixth remained HBsAg-positive and HBeAg-positive for more than two years and never developed anti-HBe. These results confirm the sensitivity of this RIA and its value in predicting the course of HBV infections.     

Evaluation of a semi-automatic radioimmunoassay for hepatitis B surface antigen (HBsAg)

The recently developed semi-automatic Hepatube system^® was evaluated in comparison to another radioimmunoassay for the detection of hepatitis B surface antigen (HBsAg), the manual Ausria II-125 test^®. After incubation of serum in anti-HBs coated tubes, the Hepatube system uses a machine to wash the tubes and to add tracer. After a second incubation, tubes are washed again in the machine and are manually transferred to the γ counter. Two machines were used. Machine 1 had an undefined defect. Of 1490 samples tested, 69 (4.6%) gave false-positive results versus 11 (0.7%) in the Ausria II-125 test. Machine 2 had one false-positive result among 920 samples versus 5 in the Ausria II-125 test. The sensitivity was measured with reference panels from Wellcome and Abbott as well as in titration series. The Hepatube system was found to be a factor three less sensitive than the Ausria II-125 test. The Hepatube processor is easy to handle; radioactive material can be held at a distance during the whole procedure; waste material is limited and less voluminous than in the Ausria II-125 test.     

Hepatitis B core antigen (HBcAg) in serum of patients with chronic hepatitis B detected by a modified radioimmunoassay

Serum hepatitis B core antigen (HBcAg) was investigated in 85 patients with chronic hepatitis B virus (HBV) infection using a modified radioimmunoassay technique, based on high molarity treatment of samples to avoid masking of the antigen by the excess homologous antibody. Eighty-eight percent of HBeAg-positive cases and 19% of anti-HBe-positive cases were HBcAg positive in serum, with a positive correlation with the presence of HBcAg in the liver. Although the sensitivity of the method for the presence of complete virions was not absolute, as shown by the comparison with serum HBV-DNA testing, this technique may be helpful for assessing virus synthesis in patients with HBV infection.     

Cryptic association of e antigen with different morphologic forms of hepatitis B surface antigen

When highly purified HBsAg particles, separated by rate zonal centrifugation into populations differing in predominant size, were tested for HBeAg, the e1 specificity was detected preferentially in association with particle fractions containing large filaments and Dane particles. These results were obtained both by agar gel diffusion and by radioimmunoassay for e antigen. The e antigen activity present in these fractions was potentiated by prior treatment of particles with Tween 80, suggesting cryptic localization of e1 specificity within or under the outer membrane. The HBeAg released by detergent treatment from a purified preparation composed predominantly of small-particle forms of HBsAg was separated by electrofocusing into a peak of nonparticulate e antigen in the pH range of 5.7--6.0. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed three major polypeptides in this preparation with approximate molecular weights of 25,000, 55,000, and 70,000. Furthermore, two additional peaks of e antigen activity were detected which migrated in association with HBsAg particles at isoelectric points of 4.4 and 5.5--5.6. The major portion of e antigen remained in association with particles after further purification by rate zonal centrifugation.     

Monoclonal radioimmunoassay for hepatitis B surface antigen in sera of children with acute leukemia

Hepatitis B surface antigen (HBsAg) was detected by a monoclonal antibody radioimmunoassay in sera from five of 43 children (11.6%) with acute leukemia, who were negative by conventional assay. None of the nine positive sera had evidence of reactivity for HBV-DNA or DNA-polymerase activity. No correlation was found between the presence of HBsAg in serum by monoclonal RIA and the behaviour of anti-viral antibodies. Twenty-two children could be studied for liver HBsAg by immunofluorescence, and nine of them (40.9%) were positive, including three patients having HBsAg reactivity in serum. These data indicate that monoclonal antibodies increase the sensitivity of RIA for the detection of serum HBsAg in children with acute leukemia, who previously have frequently been found to have an atypical hepatitis B virus (HBV) serology.     

Evaluation of a new radioimmunoassay for detecting hepatitis B e antigen and its antibody

A new commercially available radioimmunoassay (RIA) (Abbott-HBe^TM) was used for determination of hepatitis B e-antigen (HBeAg) and its antibody (anti-HBe). Serial serum samples from 20 transiently HBsAg-positive patients with acute hepatitis were tested. In nearly all patients HBeAg could be shown for a short period with subsequent development of antiHBe. From 24 chronic HBsAg carriers serial serum samples collected during several years were tested. In 18 of 19 initially HBeAg-positive patients the HBeAg was lost after 6 months to 6 years; in 14 anti-HBe developed. A correlation was seen between the seroconversion and normalization of elevated alanine transferase levels. From another 22 chronic HBsAg carriers single serum samples were assayed. These samples were selected because neither HBeAg nor anti-HBe could be detected by the immunodiffusion (ID) technique. They had previously been examined for HBV-associated DNA-polymerase activity. In 20 patients HBeAg or anti-HBe could be detected by RIA. Those who were DNA-polymerase negative had anti-HBe, and 3 of 4 who were DNA-polymerase positive had HBeAg. When compared to earlier results by ID in these materials a higher frequency of HBeAg and, in particular, anti-HBe was detected by the RIA. By this test, HBeAg or anti-HBe was found in nearly all patients. The usefulness of HBeAg/anti-HBe in the evaluation of infectivity and prognosis in hepatitis B has been limited by the low sensitivity of the earlier test systems. Thus, this new RIA is a valuable addition to the diagnostic tests for patients with hepatitis B.     

New bacterial absorption method for determination of hepatitis A IgM and IgA antibodies

Antibodies against hepatitis A virus (anti-HAV) can be determined by a commercially available radioimmunoassay (RIA) (HavabTM, Abbott). To discriminate between recent and past hepatitis A infection this RIA was used in combination with absorption with protein A-containing staphylococci. However, nonabsorbable anti-HAV was repeatedly detected in late-convalescent sera using this methods. The nature of these antibodies was studied in serum samples from 12 such patients. In all patients, the late-convalescent sera contained no IgM class anti-HAV as judged by sucrose density gradient centrifugation. The restricted specificity of staphylococcal protein A explains the lack of absorption. Some recently described streptococcal strains capable of binding all IgG subclasses (including IgG3) as well as both IgA subclasses were, therefore, added to the staphylococci. Absorption studies using these strains indicated that the previously nonabsorbable anti-HAV in these 12 patients was mainly of the IgA class. A bacterial mixture including IgA-binding streptococci seems preferable to routine determination of IgM anti-HAV in acute hepatitis A diagnosis. The results also indicate that IgA anti-HAV in serum can persist for more than two years after a hepatitis A infection.     

Detection of HBeAg and anti-HBe in acute hepatitis B by a sensitive radioimmunoassay

A solid-phase radioimmunoassay using anti-HBe-coated polysterene beads and iodine-¹²⁵-labeled anti-HBe of human origin was developed for the detection of HBeAg. Anti-HBe could be determined by a blocking test. Both assays were about 500-fold more sensitive than immunodiffusion. Few nonspecific positive results for HBeAg could be recognized in the anti-HBe test by increase in cpm over that of the negative control. HBeAg was not found in acute hepatitis A and non A-non B heptatis or in a control group of accident patients. On admission to the hospital 12 of 48 (25%) acute hepatitis B patients from Greece and 17 of 20 (85%) acute hepatitis B patients from Germany were HBeAg-positive. All 39 initially HBeAg negative sera were already anti-HBe positive. Tests of the acute stage and follow-up sera of the 20 German patients indicated that HBeAg is regularly present in the incubation period and early acute phase of hepatitis B. After onset of disease the antigen is cleared from the serum very rapidly in uncomplicated cases and is usually followed by the appearance of anti-HBe. Like anti-HBe, anti-HBe. can serve as a tool for the diagnosis of hepatitis B after the disappearance of HBsAg.     

Hepatitis B surface antigen confirmatory testing for diagnosis of hepatitis B virus infection in Taiwan

This study aimed to examine the application of hepatitis B surface antigen (HBsAg) confirmatory testing when diagnosing hepatitis B infection among young persons in Taiwan with a low prevalence rate of hepatitis B infection. HBsAg status, the presence of antibodies against HBsAg (anti-HBs), and the presence of antibodies against hepatitis B core antigen (anti-HBc) were compared among 403 graduate students (mean age 22.8 ± 0.7 years) and 1,745 undergraduate students (18.6 ± 1.0 years) from one university, and 367 adult subjects (41.1 ± 15.8 years) in 2008. Any HBsAg-positive subjects were tested with an HBsAg confirmatory test. Chi-square tests for trend and predictive values of positivity (PVP) when using HBsAg-positive only for determining confirmed cases of hepatitis B infection were compared across the three cohorts. The prevalence of HBsAg positivity among subjects decreased from 16.3% in the adults to 5.2% in the graduate students and then to 2.8% for the undergraduate students (P = 0.0007). The PVP of HBsAg testing when determining cases of hepatitis B decreased from 0.97 for the adults to 0.81 for the graduate students and then to 0.56 for the undergraduate students (P < 0.0001). Thus, a significant decrease in the true-positive rate of HBsAg among the students born after the introduction of hepatitis B vaccination was observed only when HBsAg testing was applied. Additional neutralization tests may therefore become mandatory for persons with a positive HBsAg test result who were born after the commencement of the universal neonatal hepatitis B vaccination program in Taiwan.     

Serologic response in human hepatitis A: detection of antibody by radioimmunoassay and immune adherence hemagglutination.

An indirect solid-phase radioimmunoassay (RIA) for detection of antibody to the hepatitis A antigen (anti-HAV) was developed using polystyrene pearls as the solid phase and hepatitis A antigen (HAAg) extracted from marmoset livers. This RIA was compared to an immune adherence hemagglutination assay (IAHA) which employed HAAg derived from the stools of chimpanzees collected during acute hepatitis A. Anti-HAV was detected in the sera of 15 humans with naturally acquired hepatitis A infection. Sensitivity and specificity were greater using the RIA, permitting the detection of anti-HAV as early as the time of onset of jaundice. Either seroconversion or a significant increase in the titer of anti-HAV was demonstrated following hepatitis A exposure in paired sera from six patients by both techniques. No significant difference in anti-HAV responses was noted between patients with icteric compared to anicteric hepatitis A or between children and adults with hepatitis A.     

Detection and persistence of specific IgA antibodies in serum of patients with hepatitis A by capture radioimmunoassay

The serum immunoglobulin A (IgA) response to hepatitis A virus (HAV) was investigated with a sensitive capture radioimmunoassay. In serial serum samples drawn from 15 patients with viral hepatitis A, IgA anti-HAV antibodies reached their highest titer between 1-2 weeks after onset and peak titers ranged from 10,000-20,000. Serum samples were available from six patients 30-32 months after onset of illness. These samples were all positive for IgA anti-HAV and some had titers similar to peak titers during illness. However, the height of the titration curves, expressed as the binding ratio (BR) at a dilution of 1/1000, was in all cases significantly lower at 30-32 months than during acute illness and early convalescence. The significance of the persistence of the IgA anti-HAV and possible reasons for the change in the BR are discussed.     

The separation and analysis of hepatitis B e antigen.

A solid-phase radioimmunoassay has been used successfully for detecting hepatitis B e antigen in fractionated hepatitis B virus-containing serum. Ammonium sulphate precipitation followed by gel filtration through a column of Sepharose CL-6B resulted in two fractions of antigen-containing material with molecular weights of 220,000 and 130,000. The smaller of these two fractions was found to possess an average isoelectric point of 4.9 and consisted of two major polypeptide species with estimated molecular weights of 66,000 and 17,000 respectively. Affinity chromatography on Blue Sepharose showed that e antigen was not retained under conditions which bound serum albumin. These results are discussed in relation to the immunopathogenesis of hepatitis B.     

Prevalence of a non-A, non-B-associated antigen/antibody system detected by radioimmunoassay in acute and chronic liver disease

An antigen/antibody system associated with one form of parenterally transmitted non-A, non-B (NANB) hepatitis has been identified by solid-phase radioimmunoassay (RIA). The antigen is found in 3% of normal subjects and in increased frequency in patients with haemophilia, renal homograft recipients, homosexual men, prostitutes, drug addicts, and patients with chronic hepatitis B virus (HBV) infection. The antigen was found in normal frequency in patients with acute hepatitis A and B, sporadic NANB hepatitis (nondrug addicts), autoimmune and alcohol-induced chronic liver disease, and primary biliary cirrhosis.     

Detection by radioimmunoassay of igm class antibody to hepatitis b core antigen: a comparison of two methods

A solid-phase radioimmunoassay (RIA) for IgM anti-HBc is described. The assay (anti-m? RIA) is based on the adsorption of IgM to a tube coated with sheep antibody to the human IgM m?-chain. The adsorbed immunoglobulin is assayed for anti-HBc activity. Positive reactions in the test are shown to be due to IgM antibody, and a confirmation blocking test is described. Reactivity of test sera can be quantitated by comparison with standard sera containing known levels of IgM anti-HBc. Sera assayed for IgM anti-HBc by both the anti-m? RIA and a method employing serum fractionation and competitive RIA gave similar results. Rheumatoid factor (RF) alone did not react in the anti-m? RIA, but reactivity could be generated in the presence of RF and preformed aggregates of IgG anti-HBc.     

转铁蛋白放射免疫分析方法的建立

目的 :在样品血清不作稀释的前提下建立转铁蛋白的放射免疫分析方法。方法 :通过降低标记抗原的比放射性 ,提高抗血清的浓度 ,改进曲线拟合方式等方法 ,扩大测定范围。结果 :抗血清亲和率为 3 82×10 -10 mol/L ,标记抗原的比放射性为 15 3mg/mCi,批间误差CV =5 38%批内误差CV =3 5 % ,曲线形态良好。 30例正常人的血清Tf含量为 2 81± 0 972mg/ml。结论 :在样品血清不作稀释的前提下建立转铁蛋白的放射免疫分析法 ,具有较好的临床实用价值。     

Oyster-associated gastroenteritis in Australia: the detection of Norwalk virus and its antibody by immune electron microscopy and radioimmunoassay

Following widespread outbreaks of oyster-associated gastroenteritis in Australia during 1978 in which Norwalk virus was implicated as the causative agent, collaborative studies were undertaken between laboratories in Australia and the United States to confirm the etiology. Immune electron microscopy (IEM) techniques were used in Australia and radioimmunoassay (RIA) methods in the United States. Norwalk virus was detected by IEM in seven of 15 faecal samples, and four were positive by RIA. A much better correlation was found with antibody determinations. Both methods demonstrated significant increases in antibody to Norwalk virus in 22 of 30 sets (73%) of "acute" and "convalescent" sera, confirming that Norwalk virus was responsible for the majority of cases. It is significant that the RIA serology was determined using Norwalk antigen originating in the United States and the IEM serology was determined using 27--30-nm particles originating in Australia.     

HBeAg and anti-HBe detection by radioimmunoassay: correlation with vertical transmission of hepatitis B virus in Taiwan.

A sensitive radioimmunoassay technique was used to detect hepatitis B e antigen (HBeAg). A strong correlation was found between HBeAg positivity of the serum of hepatitis B surface antigen (HBsAg) carrier women in Taiwan and the subsequent development of surface antigenemia in their babies. All babies who became chronic HBsAg carriers were born to HBeAg positive women, maternal HBeAg positivity being a better prior indication of chronic antigenemia developing in the baby than the HBsAg titer in the mother's serum.     

Prevalence of antibodies to Hepatitis A antigen in sera from patients with haematological malignancies

The prevalence of antibodies to hepatitis A (anti-HA) was investigated in patients attending a clinic for haematological malignancies. Patients were divided into three age groups; the number with antibodies in each group was recorded. Of the 130 patients studied, 44% of those aged 4-26, 70% of those aged 27-50, and 83% of those aged 51-76, were positive. This study identifies patients attending a clinic for haematological malignancies (ie, patients with immunologic deficiencies) as a group of people having a high prevalence of anti-HA. In the oldest group (51-76), urban patients had a higher prevalence than rural patients - a pattern similar to that observed in elderly Red Cross blood donors in a previous study. Seroconversion was noted in six patients, three of whom seroconverted from anti-HA-negative to anti HA-positive within 12 months of the initial visit to the clinic.     

Immunofluorescent technique for the detection of monoclonal internal image anti-idiotypic antibodies of hepatitis B surface antigen

Monoclonal anti-idiotypic antibodies to HBsAg were screened by immunofluorescence for the presence of a subset behaving as the internal image of the original antigen. We describe the technique and the criteria fulfilled to establish that 2/6 monoclonals studied act as the internal images of the a determinant of hepatitis B surface antigen.