Measurement of antibodies to herpesvirus types 1 and 2 in human sera by microradioimmunoassay.

The binding of 125I-labelled anti-human antibodies against the fc IgG fragment to unlabelled antiviral immunoglobulins in the surface of infected cells was used to quantitate antibodies against herpes simplex virus type (HSV-1) and type 2 (HSV-2) in sera from patients with cervix carcinoma. The microradioimmunoassay technique (micro-RIA) proved to be 5-10 times more sensitive than the microneutralization test. Antibody titres determined by micro-RIA correlated with neutralizing antibody titres to both HSV-1 and HSV-2. The relative antibody titres to HSV-1 and HSV-2, as determined by micro-RIA, could be used to distinguish persons previously infected with HSV-2 by means of II/I indices.     

Immunofluorescent IgG and IgM antibodies in human sera by enterovirus infections

IgG and IgM antibodies to selected enterovirus serotypes (P2, CB 1-5, E2, 4, 20), to prototype collection strains and to fresh isolates were examined by immunofluorescence in paired sera of 11 children and 4 adults in whom enterovirus had been isolated, or a rise of specific antibodies had been proved by neutralization test. One six-month- and one seven-month-old child had antibodies of both classes to the isolated virus strain and to poliovirus only. Children from eight month onwards and adults had antibodies to the majority of enterovirus serotypes tested. Among all enteroviruses tested, heterologous reactions were observed not only with IgG but also with IgM class antibodies.     

An enzyme-linked immunosorbent assay (ELISA) for measurement of antibodies against equine herpesvirus 2 in equine sera

An indirect enzyme-linked immunosorbent assay (ELISA) was developed for the detection of antibodies against equine herpesvirus type 2 (EHV-2) in equine sera. The optimal conditions of antigen concentration, and serum and conjugate dilutions were established by chequerboard titrations. When the standard ELISA test was used for titration of test sera, it was found to give titres approximately 1500 times higher than those obtained in the virus neutralization (VN) test, and a correlation coefficient of 0.815 was obtained between these two tests on 42 equine sera. All the positive serum samples by the VN were also positive by the ELISA, and one negative serum in the former test was found to be positive in the latter. Under field conditions, the test also detected increases in antibody titres against EHV-2 in 13 out of 14 foals soon after these animals excreted the virus.     

Enzyme-linked immunosorbent assay for determination of antibodies against herpes simplex virus types 1 and 2 in human sera.

A rapid and reproducible enzyme-linked immunosorbent assay (ELISA) is described for determining antibodies in human sera against herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2). The sera were absorbed for 30 min with heterologous virus-infected-cell extracts to remove cross-reacting antibodies and then were applied to ELISA plates containing the target antigens, immunoaffinity-purified HSV-1 glycoproteins gC and gD and HSV-2 glycoproteins gD and gF. The absorbance index, defined as the ratio of A414 generated by a serum sample absorbed with a heterologous virus-infected-cell extract versus the A414 of a serum sample absorbed with an uninfected-cell extract, was used to determine the presence or absence of antibodies to HSV-1 and HSV-2. Results of the ELISA for detecting antibodies against HSV-2, when compared with results obtained for the same sera by the microneutralization test, showed an index of overall agreement of 91%. Results of the ELISA for detecting antibodies against HSV-1, when compared with microneutralization test results for sera negative for HSV-2 antibodies but positive for HSV antibodies by ELISA, showed an index of agreement of 99%.     

Anti-albumin antibodies in normal rabbit sera

Antibodies reacting with homologous glutaraldehyde (GA)-modified albumin were demonstrated in normal rabbit sera (NRS) by passive hemagglutination and direct binding assay. The ability of rabbit anti-albumin antibodies (AAA) to react with homologous GA-modified albumin was found to increase with the degree of albumin polymerization. AAA did not react with GA-treated monomeric albumin. It was proved that rabbit AAA are not species-specific since they react also with heterologous GA-polymerized albumin. The possible role of AAA in the catabolism of in vivo aged albumin molecules, antigenically similar with GA-induced albumin polymers, is discussed.     

Immune response to bovine herpes herpesvirus type 1 infections: virus-specific antibodies in sera from infected animals.

The virus specificity of antibodies against bovine herpes virus type 1 was determined with a radioimmunoprecipitation assay and serum collected from natural and experimentally induced infections. By using sequentially collected sera, the development of antibodies to 4 to 5 viral glycoproteins and 11 to 12 nonglycosylated proteins was followed for the first 50 days after infection. The major and most consistent responses in experimentally and naturally infected animals were to four glycoproteins with molecular weights of 102,000, 96,000, 69,000, and 55,000, as well as to a major virion 115,000-molecular-weight nonglycosylated protein. The four glycoproteins were all coprecipitated by a neutralizing monoclonal antibody and were probably involved as target antigens in virus neutralization. Another antigenically unrelated glycoprotein with a molecular weight of 82,000 and a nonglycosylated protein with a molecular weight of 91,000 were also precipitated, but the immune response to these two proteins was transient. Reactivity to gp82 was only weakly detected in serum from naturally infected animals. Contact control animals which did not contract a bovine herpes virus type 1 infection but were exposed to infected animals with signs of severe illness had antibodies which recognized gp102, gp96, gp69 and gp55 as well as p115. These antibodies were present in low amounts and, in contrast to infected animals, did not increase between acute and convalescent sampling.     

Enzyme-linked immunosorbent assay for quantitation of toxoplasma antibodies in human sera.

An enzyme-linked immunosorbent assay(ELISA) was developed for the detection of toxoplasma antibodies using a single serum dilution (1:800) in conjunction with a standard curve Antigen was prepared from Toxoplasma gondii cultivated in human cell cultures. A nearly linear relationship was found between the logarithms of the absorbance values of 120 human sera at a dilution of 1/800 and the titres as determined by an end point dilution ELISA. The reproducibility of the single dilution ELISA was excellent; the coefficients of variation for within-day and day-to-day tests were less than 15%. A close correlation was found between the results obtained with ELISA, indirect immunofluorescence (IF), and complement fixation (CF). The titres in ELISA were 20 to 40 times higher than in IF and 200 to 1000 times higher than in CF.     

Synthetic-peptide-based enzyme-linked immunosorbent assay for screening human serum or plasma for antibodies to human immunodeficiency virus type 1 and type 2.

A synthetic-peptide-based enzyme-linked immunosorbent assay (EIA) capable of screening for antibodies to both human immunodeficiency virus type 1 (HIV-1) and HIV-2 has been developed for use in blood banks and diagnostic laboratories. Microtiter wells are coated with two synthetic peptides, one corresponding to the highly conserved envelope region of HIV-1 and another corresponding to the conserved envelope region of HIV-2. Overall, sensitivity was 100% in 303 individuals diagnosed with AIDS and 96 individuals diagnosed with AIDS-related complex, 14.8% in a study of 500 high-risk group members, 99.9% in 600 EIA repeatedly reactive (RR)-HIV-1 Western blot (WB)-positive repository specimens, and 100% for 222 geographically diverse HIV-1 specimens and 216 confirmed HIV-2-positive specimens evaluated. The specificity was determined to be 99.72% for a total of 13,004 serum and plasma samples from random volunteer donors evaluated across five blood banks. Forty donors who were found to be EIA RR-WB indeterminate but nonreactive on the United Biomedical, Inc., test (UBI HIV 1/2 EIA) were prospectively followed as an additional measure of specificity. None of the 40 low-risk cases evolved into a positive WB pattern at follow-up. The sensitivity and specificity of this new assay are comparable to those of other Food and Drug Administration-licensed HIV-1 and HIV-1-HIV-2 assays that are currently available in the United States. The UBI HIV 1/2 EIA affords laboratories another choice in the detection of antibodies for HIV-1 and HIV-2 with a test based on an alternative antigen format.     

Anti-cholesterol antibodies in human sera

In the last 30 years many research showed that high serum cholesterol level is a great risk for the atherosclerosis. In recent years, it has become clear that the immune system has a major role in atherosclerosis development and progression, and has binding capacity to cholesterol as well. It has been demonstrated in animal experiments, that anti-cholesterol antibodies (ACHA) can prevent cholesterol diet induced atherosclerosis. Our group is looking for the answer, whether ACHA have the same function in animals and in humans, or not. In this review we summarize our studies in human sera. We measured serum ACHA levels in different groups of patients with atherosclerotic diseases in patients with viral infections and in healthy population. In the summary we write about the possible functions of ACHA in the human immune system.     

Relationship between type specific antibodies to herpes simplex virus and type specificity in human sera

A quantitative analysis was carried out on the relationship between type specific neutralizing antibodies to herpes simplex virus and the type specificity, using a mutual absorbing procedure. Forty-two samples were selected among human sera, on the basis of type specificity expressed by the value of II/I index. All sera with values of less than 90 of II/I index contained only type 1 specific antibody, while those with values over 111 contained only type 2 specific antibody. When the values were between 90 and 110, type 1 specific antibody was present in all, and type 2 specific antibody was present in some serum samples.     

Enzyme-linked immunosorbent assay for detection of antibodies to influenza A and B and parainfluenza type 1 in sera of patients.

An enzyme-linked immunosorbent assay (ELISA) was developed for the detection of antibodies to influenza A/Hong Kong/1/68, influenza A/Victoria/3/75, influenza B/Hong Kong/5/72, and parainfluenza type 1 viruses. Development and standardization of the method indicated that an acetone-ethyl alcohol mixture was a suitable fixative for the preparation of the solid-phase coupled antigen. The addition of sodium azide to the enzyme-conjugated solution and the concentrations of the enzyme-conjugate antiglobulin and test sera employed were all critical factors in the success of the ELISA procedure. The ELISA test was specific; there was no cross-reaction between influenza A and B or parainfluenza type 1 viruses. The concordance between ELISA and hemagglutination inhibition results suggested that both tests probably detected the same type of antibodies. The ELISA procedure was 8 to 64 times more sensitive than complement fixation and/or hemagglutination inhibition tests. Low levels of antibody in patients' sera were detected only by the ELISA test. During the course of the testing period false positive reactions were not encountered. The results of ELISA could be obtained within 3 h. The ELISA test required a very small amount of serum and, therefore, offered an opportunity to detect the presence of maternal antibodies to influenza viruses in blood collected from infants by heel prick.     

Immunofluorescence assay for detection of antibodies to human immunodeficiency virus type 2.

A total of 215 serum samples were tested for antibodies against human immunodeficiency virus type 2 (HIV-2) with an immunofluorescence assay (IFA). Some samples originated from Denmark and some originated from Guinea-Bissau. The IFA results were compared with enzyme-linked immunosorbent assay (ELISA) and Western (immuno-) blot (WB) results. Twenty-nine serum samples were found to be true positive for HIV-2 antibodies as judged from WB and radioimmunoprecipitation results; all of these were also found to be positive in the HIV-2 IFA. Of 80 serum samples originating from HIV-1-infected persons, 60% showed reactivity in the HIV-2 ELISA, and 51% cross-reacted with at least one band in the HIV-2 WB. None of the sera cross-reacted in the HIV-2 IFA. A total of five serum samples (three African and two Danish) gave unspecific results in the HIV-2 IFA. It is concluded that the HIV-2 IFA is more specific and at least as sensitive as a first-generation ELISA and that IFA is superior to WB in discriminating between HIV-1 and HIV-2 infections.     

HSV-1 gB and VZV gp-II crossreactive antibodies in human sera

Summary The specificity and prevalence of human IgG antibodies crossreactive between HSV-1 (ANG) and VZV (Ellen) was examined in immunoblots. Using antibody fractions purified on HSV- and VZV-coated affinity chromatography columns and by preadsorption of sera with HSV and/or VZV lysates a cross-reactivity between HSV-1 gB and VZV gp-II was demonstrated. Crossreaction of human IgG antibodies among other structural and nonstructural viral proteins, however, was not detected. The frequency of human IgG antibodies crossreactive between HSV-1 gB and VZV gp-II was highest in HSV-seropositive patients experiencing an acute primary VZV infection (4 out of 5 sera tested). In contrast, no crossreactive antibodies were found in sera of HSV-seronegative patients with acute primary VZV infection (0/6) or in sera from individuals with acute recurrent HSV or VZV infection (0/12). Analysis of sera from individuals with previous HSV and/or VZV infection showed the presence of antibodies crossreactive between HSV-1 gB and VZV gp-II in 3 out of 30 sera tested.     

Persistent infection with bovine herpesvirus type 1: rabbit model.

Persistent infection with bovine herpesvirus type 1 (BHV-1) was established in all rabbits after conjunctival inoculation of virus. Spontaneous reactivations of BHV-1 with and without the appearance of recurrent ocular lesions were observed in persistently infected rabbits. BHV-1 was reactivated predictably and shed from all persistently infected rabbits after the administration of dexamethasone. During all reactivations, BHV-1 isolation was restricted to the inoculated eye.     

Anti-horse globulin antibodies in human sera

Organ transplant patients who had received ALG, patients with rheumatoid arthritis and normal persons were studied for serum antibodies to horse globulins. Although normal individuals rarely show the presence of anti-horse antibody, rheumatoid patients with high titres of rheumatoid factor usually show anti-horse globulin agglutinins in their IgM globulins and these agglutinins are considered to be a manifestation of their `rheumatoid factors'. These agglutinins are readily absorbable with aggregated human γ-globulin. On the other hand, although most transplant patients administered ALG produce anti-horse antibody, it is usually produced as an IgG globulin and it is not absorbable with aggregated human γ-globulin.     

Maternal antibodies in human neonatal sera

Three hundred and sixty-three pairs of neonatal and maternal sera collected at delivery just after an influenza A2/Hongkong virus epidemic, were tested by complement fixation for influenza A2 antibodies. The results confirm an earlier suggestion. In this study the neonatal serum level of IgG class antibodies exceeded that of the mother in 109 of 329 cases, when the maternal value was low or normal, but in the case of a high maternal titre, the newborn had a higher titre than that of the mother in only four of thirty-four cases.     

Rapid, sensitive, competitive serologic enzyme-linked immunosorbent assay for detecting serum antibodies to bovine herpesvirus type 1.

An enzyme-linked immunosorbent assay (ELISA) for the detection of cattle antibodies to bovine herpesvirus type 1 was developed on the basis of competition between serum antibody and a virus-neutralizing mouse monoclonal antibody. The assay showed improved sensitivity over the virus neutralization (VN) test and over an enhanced VN test in which incubation of antibody-virus mixtures was carried out for 24 h. With the ELISA, antibodies in sera from experimentally infected cattle were detected earlier after infection and showed more rapid increases in levels. A comparison of the ELISA with the VN tests by using a set of 85 field sera with low levels of antibodies demonstrated that the ELISA was the most sensitive test, detecting 10 positive serum samples that were negative by the VN tests. The ELISA was inexpensive, rapid, and highly reproducible and showed a significant improvement in sensitivity over VN tests.     

ELISA for detection of IgG and IgM antibodies to HSV-1 and HSV-2 in human sera

A rapid, enzyme-linked immunoassay (ELISA) was applied to identify and measure specific IgG and IgM antibodies to herpes simplex viruses types 1 and 2 (HSV-1 and HSV-2). Detergent solubilized infected cells and mock-infected cells were used as antigens in the assay. Identification of type-specific antibodies was achieved by a competition assay in which clinical sera mixed with HSV-1 or HSV-2 antigens were assayed for reactivity to identical antigens coating wells of polystyrene microtiter plates. Reactivity and the specificity of the reactive immunoglobulin class was quantitated using biotinylated goat anti-IgG and biotinylated goat anti-IgM. Five paired sera from patients with diagnosed herpes simplex genital infections and one human anti-HSV-1 reference serum were tested with this assay and results were compared to results previously obtained using a complement fixation test and micro-SPRIA. The results indicate that the ELISA is a specific, sensitive and simple test which confirms the herpes simplex virus infection history of patients.