Quantitation of dengue precipitating antibody by inhibition countercurrent immunoelectrophoresis.

The inhibition countercurrent immunoelectrophoresis test was employed to detect dengue virus antibody in patients' sera. Anti-dengue type 2 titers determined by inhibition countercurrent immunoelectrophoresis correlated well with hemagglutination inhibition titers. In secondary cases, more than fourfold increases in precipitating antibodies were observed. The control sera were negative except for sera from a few patients with systemic lupus erythematosus, which showed low titers. Simultaneous detection of dengue virus antigen and antibody in sera collected during the acute phase could confirm at least 90% of cases. This method is recommended as a routine technique to quantitate antibody in sera from suspected cases of dengue hemorrhagic fever.     

Identification of measles virus-specific hemolysis-inihibiting antibodies separate from hemagglutination-inhibiting antibodies.

The occurrence of antibodies giving hemolysis inhibition (HLI) but not hemagglutination inhibition (HI) was examined in human convalescent and rabbit hyperimmune sera. HI antibodies, which through their interaction with hemagglutinin components display HLI activity, were removed by absorption with Tween 80-ether (TE)-treated measles virus material. This absorption did not change the titer of non-HI HLI antibodies. After removal of HI antibodies from 16 late measles convalescent sera and three batches of gamma globulin. HLI antibody titers showed a two- to eightfold reduction. The titers of neutralizing antibodies were reduced from 1/4 to 1/20 of the original titers. There was a good correlation between the titers of neutralizing and HLI antibodies both in sera from which HI antibodies had been removed by absorption and in sera spontaneously showing markedly higher HLI than HI antibody titers. HLI antibodies with these characteristics could be identified in HI tests when whole virus instead of TE-treated material was used an antigen and anti-antiserum was added to the tests. In contrast to the situation in human sera, antibodies remaining after removal of HI antibodies from rabbit hyperimmune sera against purified virus particles were detectable in neutralization and HLI tests only in the presence of anti-antiserum. However, virus particles from which the major fraction of all envelope projections had been removed by treatment with 0.004% trypsin induced the production of non-HI HLI antibodies active also in the absence of anti-antiserum. TE and formalin treatment destroyed the hemolytic activity of virus preparations and also their capacity to induce a production of non-HI HLI antibodies.     

Evaluation of an IgG enzyme-linked immunosorbent assay for dengue diagnosis.

BACKGROUND: The hemagglutination inhibition (HI) test has been one of the standards, with the IgM antibody capture ELISA (MAC-ELISA), for the diagnosis of dengue virus infections. The spread of dengue throughout the world and the increasing number of cases to be tested makes an ELISA-format test for IgG antibodies to replace the HI test highly desirable. OBJECTIVES: Evaluate the use of the IgG-ELISA as a substitute for the HI test in dengue diagnosis. STUDY DESIGN: Paired serum samples defined as being from primary or secondary dengue virus infections by HI, were tested by an ELISA that detects IgG antibodies. The correlations of titers and serologic interpretations between these two tests were examined. RESULTS: The IgG-ELISA showed a low correlation with the HI in primary infections, and a higher correlation in secondary infections because of the influence of IgM antibodies in the HI test. Nevertheless, IgG ELISA titers could be reliably associated with primary or secondary infections when analyzed by days after onset of symptoms, and can be used to characterize the immune response after flavivirus infections. CONCLUSION: The combination of the IgM and IgG ELISAs may be used to serologically diagnose dengue virus infections, since the IgG ELISA can substitute for the HI test in characterizing the immune response to dengue virus infections.     

Comparison of antibody titers determined by hemagglutination inhibition and enzyme immunoassay for JC virus and BK virus

A comparison of antibody titers to JC virus (JCV) or BK virus (BKV) was made by hemagglutination inhibition (HI) and enzyme immunoassay (EIA) with 114 human plasma samples. Antibody titers to JCV or BKV determined by HI were lower than those determined by EIA. Nevertheless, as HI titers increased so did EIA titers. When antibody data were compared by the Spearman rank correlation test, highly significant correlations were found between HI and EIA titers. Results obtained by plotting EIA antibody titers for JCV against those for BKV generally showed a reciprocal relationship, i.e., samples with high antibody titers to JCV had lower antibody titers to BKV and vice versa. Some samples, however, had antibody titers to both viruses. Of the samples tested, 25.4% (25 of 114) had HI and EIA antibody titers to JCV and BKV which were identical or closely related. This is not the scenario one would expect for cross-reactive epitopes shared by the two viruses, but one suggesting that these samples were from individuals who had experienced infections by both viruses. Adsorption with concentrated JCV or BKV antigen of sera with high antibody titers to both JCV and BKV and testing by JCV and BKV EIA gave results which support this conclusion. Although 52.6% (51 of 97) of the samples from the Japanese population tested had very high antibody titers (>/=40,960) to either JCV or BKV, none of the samples were found by a dot blot immunoassay to have antibodies which cross-reacted with simian virus 40. The results from this study, in agreement with those of others, suggest that humans infected by JCV or BKV produce antibodies to species-specific epitopes on their VP1 capsid protein, which is associated with hemagglutination and cellular binding.     

Enzygnost-Rubella: an enzyme immunoassay for screening and quantitation of antibodies to rubella virus

Calbiochem-Behring, Enzygnost-Rubella is an enzyme-linked immunosorbent assay (ELISA) for the determination of IgG antibodies to rubella virus. Two procedures, screening and quantitative, were used for the measurement of rubella antibodies in a characterized panel of sera and in random, premarital serum specimens. Using the screening procedure, no false-positive or false-negative results were observed in testing a panel of 40 sera previously characterized by human "O" cell and chick cell hemagglutination-inhibition (HI) tests. Additional testing of 323 random, premarital serum samples resulted in 99.4% agreement with HI tests in identifying positive and negative specimens. In the evaluation of the quantitative procedure, a coefficient of correlation (r) of 0.93 between ELISA titers and the geometric mean HI titers was obtained when testing the panel of 40 characterized sera. Furthermore, 100% agreement with HI test was obtained in the detection of titer rises in 20 pairs of acute and convalescent sera. The average within-run coefficients of variation for the screening and quantitative procedures were determined to be 11.2% and 15.6%, respectively. It is concluded that the Enzygnost-Rubella reagents are sensitive, specific, and provide objective means for assessing immunestatus and detecting rises in antibody titers to rubella virus.     

Serological evidence of dengue fever among refugees, Hargeysa, Somalia

Epidemics of a malaria-like illness affected several thousand residents of the Dam Camp, a refugee camp near Hargeysa in Somalia, during 1985, 1986, and 1987. The disease was characterized by fever, chills, sweats, headache, back and joint pains for as long as 10 days in some patients. Blood smears from acutely ill patients were negative for malaria. Of 28 acute and 10 convalescent sera tested by the indirect fluorescent antibody (IFA) and by the hemagglutination inhibition (HI) tests, all were negative for antibody to Rift Valley fever, Crimean-Congo hemorrhagic fever, Sindbis, Chikungunya, yellow fever, and Zika viruses. However, antibody reactive to dengue 2 virus was detected by the IFA test in 39% (15/38), and 11 of 29 (38%) of the same sera were antibody positive by the HI test. Also, IgG antibody reactive to dengue 2 was demonstrated in 60% (17/28) of the same sera by the enzyme immunoassay (EIA), and 14% (4/28) were positive for IgM antibody. Of ten patients for which acute and convalescent sera were available, two developed four fold or greater rises in antibody titer evidencing infection. These data suggested that dengue virus may have been the cause of the epidemic among the Dam Camp refugees.     

Cross-reactivity in flavivirus serology: new implications of an old finding?

To investigate the influence of pre-existing antibodies against tick-borne encephalitis (TBE) or yellow fever (YF) viruses on dengue virus antibody test results, we examined sera from vaccinees and from individuals with previous TBE virus infection. Distinct IgG antibody cross-reactivity was found in about 15.1% in the YF-vaccinated group and in about 9.5% in the TBE-vaccinated group. Altogether 15 out of a total of 80 samples tested (18.8%) had detectable dengue virus IgG antibody titres. The serum samples from patients with acute TBE virus infection not only had the highest anti-TBE virus antibodies but were also highly cross-reactive against dengue virus antigens. The high cross-reactivity rate of YF and TBE antibody-positive sera in dengue virus antibody assays should be taken into account in the interpretation of laboratory tests for the diagnosis of flavivirus infections and when undertaking seroepidemiological surveys.     

Antibody responses determined for Japanese dengue fever patients by neutralization and hemagglutination inhibition assays demonstrate cross-reactivity between dengue and Japanese encephalitis viruses

Titers of antibodies to infecting dengue virus serotypes determined by serum neutralization assay were higher than those of antibody to Japanese encephalitis (JE) virus in Japanese dengue patients after disease day 8. Titers of antibody to dengue virus antigens determined by hemagglutination inhibition (HI) assay were higher in only 1 of 23 serum specimens after disease day 11. Thus, the neutralization test is more reliable than the HI test for serological diagnosis of dengue in countries where JE vaccination is widely used or JE is endemic.     

Antibody Responses Determined for Japanese Dengue Fever Patients by Neutralization and Hemagglutination Inhibition Assays Demonstrate Cross-Reactivity between Dengue and Japanese Encephalitis Viruses

Titers of antibodies to infecting dengue virus serotypes determined by serum neutralization assay were higher than those of antibody to Japanese encephalitis (JE) virus in Japanese dengue patients after disease day 8. Titers of antibody to dengue virus antigens determined by hemagglutination inhibition (HI) assay were higher in only 1 of 23 serum specimens after disease day 11. Thus, the neutralization test is more reliable than the HI test for serological diagnosis of dengue in countries where JE vaccination is widely used or JE is endemic.     

Comparison of immunofluorescence and hemagglutination inhibition tests and enzyme-linked immunosorbent assay for detection of serum antibody in rats infected with hemorrhagic fever with renal syndrome virus.

Serum antibodies against the B-1 strain of hemorrhagic fever with renal syndrome (HFRS) virus in wild and experimental rats were investigated by the indirect immunofluorescent antibody (IF) test, the hemagglutination inhibition (HI) test, and an enzyme-linked immunosorbent assay (ELISA). In the newly developed ELISA, monoclonal antibodies to the B-1 strain were applied as coating antibody. In sera from wild rats, the results obtained by the ELISA agreed quite well with those obtained by the HI test, but some serum samples that gave a negative reaction in the HI test gave a positive reaction in the IF test, although their IF titers were very low. In serum samples from experimental rats that had been kept in an animal house infected with HFRS virus, a group with high titers and a group with low or negative titers were clearly differentiated by all the tests.     

Antibodies against measles virus polypeptides in different disease conditions.

The occurrence of antibodies to the nucleoprotein and matrix (M) antigens of measles virus was determined in early and late measles convalescent sera and in sera from patients with multiple sclerosis, subacute sclerosing panencephalitis, chronic active hepatitis, and atypical measles. Antibodies to the two components were identified separately in serially diluted samples both by radioimmune precipitation assays and by complement fixation tests employing purified nucleoprotein and M components as antigens. The antibody response to M antigen in connection with acute infections was weak, and with time titers of antibodies to M antigen were reduced below detectable levels in most cases. A different situation was seen in patients with atypical measles. A pronounced antibody response to M antigen was shown to be a part of the generally accentuated immune response in these patients. Confirming results of others, it was shown that in spite of the increased antibody titers against most measles components in sera from patients with subacute sclerosing panencephalitis, no or only low titers of antibodies to M antigen were present. However, a similar representation of antibodies to measles virus components was also seen in sera from patients with active chronic hepatitis. The significance of this finding for the interpretation of a weak antibody response to M antigen in the presence of a pronounced antibody response to other components is discussed.     

Evaluation of the Single Radial Hemolysis Test for Measuring Hemagglutinin- and Neuraminidase-Specific Antibodies to H3N2 Influenza Strains and Antibodies to Influenza B

Antibodies to the H3 hemagglutinin of influenza A virus could be specifically measured by single radial hemolysis (SRH) when test antigens were recombinant viruses containing the relevant H3 hemagglutinin antigen and irrelevant Neq1 neuraminidase of A/equine/Prague/1/56 virus. Antibodies to influenza B virus could also be measured by the SRH technique. Antibody rises to influenza A or B virus measured by SRH agreed with results of hemagglutination inhibition (HI) tests for about 80% of the sera tested, including sera from volunteers receiving killed influenza vaccine and sera from patients naturally infected with influenza. Correlation between antibody titers measured by SRH and HI was also good. Antibodies to the N2 neuraminidase of influenza A virus could be specifically measured by SRH when test antigens were recombinant viruses containing the relevant N2 neuraminidase antigen and irrelevant Heq1 hemagglutinin of A/equine/Prague/1/56 virus. The SRH test for neuraminidase antibodies was more strain specific than was the SRH test for hemagglutinin antibodies. Probably for this reason, agreement between neuraminidase antibody determinations in human sera by the SRH test and by the neuraminidase inhibition test was poorer than agreement between the SRH test for hemagglutinin antibodies and the HI test.     

Laboratory diagnosis of primary and secondary dengue infection.

BACKGROUND: Dengue fever is routinely detected in many laboratories using commercial tests for the specific detection of dengue IgM antibodies. OBJECTIVES: We have studied the sensitivity of IgM antibody detection in paired serum samples of 43 patients with either with primary dengue (PD) or secondary dengue (SD). STUDY DESIGN: Two consecutive samples were drawn from 23 Vietnamese and 20 German patients. All patients were selected for a positive PCR and for the fact that consecutive serum samples were available. The diagnosis of PD was based on seroconversion to dengue antigen and in SD on the detection of virus RNA in the presence of anti-dengue IgG antibodies. RESULTS: In samples of patients with PD fever taken during days 1-3 of the disease no IgM antibody could be detected. During days 4-7 and after day 7, IgM antibody was detected in 55% and 94%, respectively. In patients with SD fever, even less positive IgM samples were found in samples taken during days 4-7 (47%) and after day 7 (78%). IgG titers were significantly higher in SD compared to PD patients, although high (>1280) titers were also found in some PD patients. CONCLUSION: In numerous acute dengue fever patients an early diagnosis will be obtained only by combining IgM antibody detection with detection of virus or virus RNA using RT-PCR.     

Human antibody to influenza C virus: its age-related distribution and distinction from receptor analogs.

Sera from persons of four age groups (1 to 2 years, 2 to 5 years, 20 to 30 years, and 65 to 85 years) were analyzed for hemagglutination inhibition (HI) activity for influenza C virus. Significant HI activity was found in 66% of the 237 sera tested, and titers ranged from 8 to 512. In the yoiung adult group, 96% had antibody and the highest mean titer (74.7) of any age group. Positive sera were far less common in young children (36 to 47%), and relatively low titers (18.3) were common among adults over 65. The high percentage of sera with antibody to influenza C virus suggests that infections with this virus occur at a rate greater than previously recognized. The high percentage of young adults with elevated levels of HI antibody suggested either that an immune response to influenza C infections is common or that the observed HI activity might be attributable, in part at least, to nonspecific inhibitors in the sera. We showed both directly and indirectly that most if not all the inhibitory activity in the human sera we examined was due to specific antibody, mostly immunoglobulin G. This conclusion is based on the finding that the single serum protein fraction with HI activity was found to have a molecular weight equivalent to that of 7S antibody (150,000) and that the HI activity was removed by absorption to staphyloccal protein A. Moreover, immunoglobulin from only HI-positive sera bound specifically to cells infected with influenza C virus, as shown by inhibition of hemadsorption and immunofluorescence. These findings were supported by similar results obtained with chicken antisera to C virus.     

New tests for characterization of mumps virus antibodies: hemolysis inhibition, single radial immunodiffusion with immobilized virions, and mixed hemadsorption.

Hemolysis inhibition (HLI), single radial immunodiffusion (SRID) with immobilized virions, and mixed hemadsorption tests were used for measuring antibodies against mumps virus. Rabbit hyperimmune sera against mumps and early and late human convalescent sera were analyzed. All three tests identified antibodies against both hemagglutinin and the second major envelope component, hemolysin (fusion factor). The sensitivity of the HLI test corresponded to that of the hemagglutination inhibition (HI) test, but in some sera HLI antibodies occurred in greater quantity than HI antibodies. The SRID test readily identified rises in antibody titers in connection with acute infection. Due to its simplicity and lack of sensitivity to nonspecific inhibitors, it is recommended for use in this context. The mixed hemadsorption test showed a high sensitivity for specific identification of mumps antibodies. It therefore may be suitable for use in screening for immunity to mumps.     

Radioimmunoassay for detection of antibodies to Epstein-Barr virus in human infectious mononucleosis serum specimens.

A rapid microradioimmunoassay (RIA) technique was adapted for quantitatively measuring antibody titers to antigens occurring in Epstein-Barr virus (EBV)-infected lymphoid cells. In these experiments two EBV-infected cell lines, HR1K and EB-3, were used as antigen-positive cells and Molt-4 was used as the negative control cells. The antibody titers of sera from suspected infectious mononucleosis patients were compared by RIA and indirect fluorescent antibody (IFA) methods. As determined by each of the methods, 14 of 19 sera had positive antibody titers and the remainder of the sera had negative antibody titers. Thus, the two methods agreed completely in differentiating sera with antibodies to EBV antigens. To further evaluate the antibody specificity of the RIA, the antibody titers of paired sera, pre- or early infection and postinfection, from five confirmed infectious mononucleosis patients were determined by RIA and IFA. Seroconversion was demonstrated by both RIA and IFA for each of the patients. Thus, the sensitivity and specificity of the two procedures are about the same.     

A/PR/8/34(H1N1)病毒株血清流行病学调查

目的了解当今人群中是否仍有Ａ／ＰＲ／８／３４（Ｈ１Ｎ１）这类毒株的活动。方法从全国８个不同地区采集１９７５年之后出生人的血清，用常量半加敏ＨＩ测定Ａ／ＰＲ／８／３４（Ｈ１Ｎ１）毒株的抗体，然后凡ＨＩ效价≥１０的血清，一律按同法对Ａ／Ｓｗｉｎｅ／Ｉｏｗａ／１５／３０（Ｈ１Ｎ１）、Ａ／桂富／１０／１９４（Ｈ１Ｎ１）和Ａ／京防／５３／９７（Ｈ１Ｎ１）毒株进行复核测定。结果有２４份血清ＨＩ效价≥２０，分布于８个不同地区。结论当今我国人群中存在有Ａ／ＰＲ／８／３４（Ｈ１Ｎ１）毒株的活动     

BK virus: I. Seroepidemiologic studies and serologic response to viral infection

Sera of 656 patients in seven age groups were tested by hemagglutination=inhibition (HI) and complement=fixation (CF) tests for antibodies to BK virus. Both tests revealed that BK virus antibodies are common in the population of southern Germany, and that primary infection is apparently acquired during childhood. The highest rate of positive sera, 71% in the HI test and 56% in the CF test, was found in the 15–30 year old group. The antibody titers correlated in both serologic tests, but HI tests were more sensitive and reliable than CF tests. An indirect immunofluorescence assay for the detection of BK virus-specific IgM antibodies was also developed. Studying umbilical cord blood sera from 846 healthy donors, BK virus IgM antibodies were detected in 77 sera (9.1%).     

Identification of Different Measles Virus-Specific Antibodies in the Serum and Cerebrospinal Fluid from Patients with Subacute Sclerosing Panencephalitis and Multiple Sclerosis

Matched serum and cerebrospinal fluid (CSF) samples from eight cases of subacute sclerosing panencephalitis (SSPE) and 15 cases of multiple sclerosis (MS) were characterized in neutralization, hemolysis-inhibition (HLI), hemagglutination-inhibition (HI) with Tween 80—ether-treated antigen, complement-fixation (CF), and immunodiffusion tests. CF tests were carried out with crude virus material, purified nucleocapsids, and small particle hemagglutinin as antigens. A certain diversity in the relative content of antibodies against different virus products in various sera was found. There was a high degree of correlation between titers of neutralizing and HLI antibodies, but a less strict correlation between titers of HLI and HI antibodies. Serum samples from two cases of MS and one case of SSPE contained high titers of HLI and neutralizing antibodies in the presence of only low titers of HI antibodies demonstrable with Tween 80—ether-treated antigen. The major fraction of antibodies detected in CF and immunodiffusion tests reacted with nucleocapsids. There was a tendency of nucleocapsid CF antibody titers, as compared to neutralization and HLI antibody titers, to be higher in samples from patients with SSPE than from cases of MS. No significant differences were found between antibody titers recorded in neutralization, HLI, and HI tests carried out with two different measles virus strains, Edmonston and a strain (LEC) derived from a case of SSPE. Comparison of antibodies against measles virus products and, as a reference, against a group-specific vertex capsomer antigen of adenovirus in matched serum and CSF samples revealed a production of measles virus-specific antibodies within the central nervous system of all cases of SSPE and 8 out of 15 cases of MS.