Enzyme-linked immunosorbent assay for the detection of influenza type-specific antibodies

An enzyme-linked immunosorbent assay (ELISA) using a purified ribonucleoprotein antigen is described for detection of type-specific anti-influenza virus antibodies. ELISA was found to be more sensitive than indirect immunofluorescent and complement fixation tests. A significant increase in antibody titer could be demonstrated by ELISA between serum specimens collected prior to and during the influenza season. ELISA appears to be useful for rapid and sensitive diagnosis of influenza infections.     

An enzyme linked immunosorbent assay for leucocyte and platelet antibodies

An enzyme linked immunosorbent assay (ELISA) method for the detection of antibodies to platelets and leucocytes is presented. The method can be used for large numbers of samples. The method is objective when photometers are used.Approximately 50% of all cases of possible autoimmune thrombocytopenic purpura (A.T.P.) and unexplained neutropenia showed positive results. The results obtained using ELISA and standard tests for leucocyte and platelet antibodies are compared. The ELISA tests may also detect immune complexes.     

Detection of M2 antimitochondrial antibodies by dot blot assay is more specific than by enzyme linked immunosorbent assay

AIMS: The objective of our study was, in one hand, to determine the sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of ELISA and dot blot assay to investigate IgG M2 antimitochondrial antibodies (M2 AMA) and, on the other hand, to compare these results with those of indirect immunofluorescence technique (IIF). METHODS: Sera from patients suffering from primary biliary cirrhosis (PBC) (n=55), systemic lupus erythematosus (n=21), celiac disease (n=30) and blood donors (n=75) were analyzed. M2 AMA were detected by ELISA and dot blot using pyruvate dehydrogenase purified from porcine heart and by IIF on cryostat sections of rat liver-kidney-stomach. RESULTS: IIF was more sensitive (98%) than ELISA (93%) and dot blot (91%). The specificity of AMA for PBC using IIF, ELISA and dot blot reached 100%, 92% and 100%, respectively. The PPV of IIF, ELISA and dot blot was 100%, 93% and 100%, respectively. The NPV was 98% for IIF, 92% for ELISA and 91% for dot blot. CONCLUSION: Dot blot, using purified pyruvate dehydrogenase, had a higher specificity than ELISA and may be useful in confirming the specificity of AMA in cases of doubt with IIF.     

An enzyme-linked immunosorbent assay for detection of flavivirus antibodies in chicken sera

An enzyme-linked immunosorbent assay (ELISA) was developed to determine the presence of flavivirus antibodies to Murray Valley encephalitis (MVE) and Kunjin viruses in sentinel chicken sera. The development of a quick, reliable assay to detect antibodies to MVE was an essential part of a large-scale surveillance programme to monitor arbovirus activity in Western Australia. This assay was developed for use with alkaline phosphatase conjugated goat anti-mouse IgG using mouse anti-chicken globulin in an intermediate step. There was a significant difference in absorbance values between neutralization test positive and pre-bled sera. However, some sera obtained from sentinel chickens and deemed negative by neutralization demonstrated adsorbance levels above the cutoff level in the ELISA, which reflects the increased sensitivity of this technique. The ELISA test detected antibodies to MVE in chicken sera 7-10 days after infection, whereas these antibodies were only consistently detected by the neutralization test 24 days after infection. Antibodies to both MVE and Kunjin reacted positively with the MVE antigen, but there was little cross-reactivity between this antigen and antibodies to other togaviruses. The main advantages of the ELISA over the neutralization test for detecting antibodies to MVE virus in the sera of sentinel chickens are its greater sensitivity and the speed with which tests can be performed. Results are available within 48 h of receiving specimens and emergency mosquito control measures may then be implemented.     

Detection of influenza virus neuraminidase-specific antibodies by an enzyme-linked immunosorbent assay.

We analyzed sera from 28 patients with various types of malignancies for the occurrence of antibodies against exotoxin A of Pseudomonas aeruginosa and two Pseudomonas proteases. A total of 27 of these individuals were colonized or infected with P. aeruginosa at one time or another during the study, whereas the remaining patient was colonized with four non-P. aeruginosa species of Pseudomonas. Sera were obtained from several of these patients before P. aeruginosa colonization or infection of these individuals was detected, which provided an opportunity to evaluate their responsiveness to pseudomonal exoproducts as they acquired the organism. Exotoxin A was purified from culture supernatant fluids of strain PA-103, and the two proteases were purified from an isolate of strain JR3, a highly proteolytic strain originally recovered from the sputum of a cystic fibrosis patient. Antibodies to the exotoxin A and the two proteases were detected in these sera, and sera which contained relatively high antibody levels to exotoxin A afforded mice complete protection against lethal challenges with this substance. Statistical analyses showed that patients infected with P. aeruginosa had consistently higher antibody levels (P less than 0.005) to the exoproducts than patients who were colonized with this organism. Also, patients colonized with P. aeruginosa possessed significantly higher antibody levels (P less than 0.003) to these three exoproducts than uninfected, hospitalized patients. Parke-Davis type 1 was the strain most commonly isolated from these patients (46%), but colonization or infection due to this organism usually resulted in the production of low levels of antibody to Pseudomonas exoproducts. However, infections with Parke-David type 7 organisms were always associated with intermediate- and high-responder sera to exotoxin A. These results indicated that potentially toxic products were elaborated during the course of cancer-related colonization and infection with P. aeruginosa.     

Development of an indirect enzyme-linked immunosorbent assay test for detecting antibodies to chicken astrovirus in chicken sera

The development of an indirect enzyme-linked immunosorbent assay (ELISA) for the serological diagnosis of Group B chicken astrovirus (CAstV) infections is described. The test was based on the use of an affinity-purified capsid antigen, specific to CAstV isolate 11672, produced as a glutathione-S-transferase N-terminal fusion protein by a recombinant baculovirus. Strongly positive ELISA signals were elicited against experimentally produced antisera raised to CAstVs from Group B (subgroups i and ii) but were negative for antisera raised to a Group A CAstV. Using a panel of 240 selected serum samples, 99% agreement was observed when the results obtained by ELISA were compared to those from an indirect immunofluorescence test for CAstV 11672. The ELISA test was applied to 68 serum sets comprising 1864 samples, which were obtained from parent and grandparent flocks originating mainly in the UK. Of the 52 sets containing ELISA-positive samples, 24 sets had >75% samples positive and nine sets had <25% samples positive and were regarded as having high and low seropositivities, respectively. Of the 1864 serum samples tested 1090 (58.5%) were ELISA positive and of these, 234 sera (21.5%) produced strongly positive signals, whereas moderately positive and weakly positive signals were produced by 562 (51.5%) and 294 (27%) sera. When used for flock screening purposes, this ELISA test can be used to (i) investigate the occurrence of first-time CAstV infections of parent flocks during lay and the possible adverse effects caused by vertically transmitted CAstV infections on broiler hatchability and performance and (ii) diagnose Group B CAstV infections within specific pathogen free flocks.     

Detection of anti-gag antibodies of feline immunodeficiency virus in cat sera by enzyme-linked immunosorbent assay

Summary Usinggag protein of feline immunodeficiency virus (FIV) expressed inEscherichia coli, an enzyme-linked immunosorbent assay (ELISA) system was developed for detection of antibodies to FIVgag protein in cat sera. With serum samples from cats experimentally infected with several strains and an infectious molecular clone of FIV, increases of the antibody titers to FIVgag protein were observed in all cases by the ELISA at early stage of infection. When we examined a total of 415 field cat sera which were previously tested by an indirect immunofluorescence assay (IFA), 9 (12.9%) out of 70 IFA positive sera were judged as negative by the ELISA. However, all 3 serum samples tested among the 9 IFA positive sera had antibodies to gp130 but not to p26 by a radioimmunoprecipitation assay. The results indicated that some IFA positive sera did not have antibodies to the p26 though they have antibodies to other proteins specific for FIV.     

Detection of herpes simplex virus type-specific antibodies by an enzyme-linked immunosorbent assay based on glycoprotein G

In order to develop a simple and quantitative method to detect herpes simplex virus (HSV) type-specific antibodies, the usefulness of an enzyme-linked immunosorbent assay (ELISA) using HSV glycoprotein G (gG) captured on a plate by monoclonal antibodies as antigen was studied. The gG1- and gG2-specific IgG antibody activities were measured by the ELISA for 54 sera which had been collected from culture-proven genital herpes patients and pre-characterized by an immunodot assay using purified gG antigens. Thirty control sera without antibodies against the HSV whole antigens were also included. In comparison with the immunodot assay as standard, the sensitivities of the ELISA were 88.9% (32/36) for HSV-1 antibody and 89.2% (33/37) for HSV-2 antibody and the specificities were both 100%. Sera taken within a few months after primary infection tended to give false negative results. The HSV type-specific ELISA based on easy-to-prepare gG antigens might be useful to help improve the serological assessment of HSV infections. J. Med. Virol. 53:319–323, 1997. © 1997 Wiley-Liss, Inc.     

Comparison of enzyme linked immunosorbent assay and enzyme linked fluorescence immunoassay for detection of antibodies against Chlamydia trachomatis.

An enzyme linked fluorescence immunoassay (ELFA) has been evaluated for the detection of antibodies against Chlamydia trachomatis. Reticulate bodies and elementary bodies from C trachomatis L2/434 strain were used as antigens. An enzyme linked immunosorbent assay (ELISA) has also been evaluated using the same antigens. Results obtained by ELISA and ELFA for human sera with these two antigens were compared with each other and with the results obtained by a micro-immunofluorescence (micro-IF) test. Serum IgG antibodies against C trachomatis L2 reticulate bodies and elementary bodies were found in 32 (20.0%) and 11 (6.9%), respectively, of 160 serum samples from pregnant women by the micro-IF test (titre greater than or equal to 1/32). All of these 32 pregnant women had IgG antibodies to C trachomatis reticulate bodies (titre greater than or equal to 1/100), whereas 20 (12.5%) had IgG antibodies to elementary bodies in the ELISA. On the other hand, 25 (15.6%) and 19 (11.9%) of them had IgG antibodies to C trachomatis L2 reticulate bodies and elementary bodies, respectively, by the ELFA (titre greater than or equal to 1/500).     

Tissue transglutaminase antibodies in celiac disease, comparison of an enzyme linked immunosorbent assay and a dot blot assay

AIMS: The purpose of our study is to determine the sensitivity, specificity and predictive values of an enzyme linked immunosorbent assay (ELISA) and a dot blot assay for the detection of IgA class anti-tissue transglutaminase antibodies (IgA-AtTGA) and to compare these results with those of IgA class anti-endomysium antibodies (IgA-AEA), IgA class anti-reticulin antibodies (IgA-ARA) and IgA class anti-gliadin antibodies (IgA-AGA). PATIENTS: Serum samples from 143 patients (97 children, 46 adults) with untreated celiac disease (CD) confirmed by intestinal biopsy and 74 disease controls (64 children, 10 adults) were studied. Methods. - The anti-tissue transglutaminase antibodies were detected by dot blot assay and an ELISA using guinea pig tissue transglutaminase (gp-tTG) as antigen. The anti-endomysium antibodies were detected by an indirect immunofluorescence technique on cryostat sections of human umbilical cord. The anti-reticulin antibodies were also investigated by indirect immunofluorescence on cryostat sections of kidney, liver and stomach of rat. The anti-gliadin antibodies were determined by an ELISA. RESULTS: The sensitivity of an ELISA for the detection of anti-tissue transglutaminase antibodies was 86% in children and 87% in adults and the sensitivity of dot blot assay was 57% in children and 54% in adults. The specificity of an ELISA and dot blot for the detection for anti-tissue transglutaminase antibodies was, respectively, 96% and 88% lower than that of anti-endomysium antibodies (100%). The sensitivity of anti-gliadin antibodies was 97% in children and 91% in adults and their specificity was 85%. The sensitivity of anti-reticulin antibodies was 94% in children and 87% in adults. Their specificity was 100%. CONCLUSIONS: The sensitivity and specificity of an ELISA for the detection of anti-tissue transglutaminase antibodies were better than that of dot blot assay. However, this dot blot assay could screen four celiac patients who have not had anti-tissue transglutaminase antibodies by an ELISA. The sensitivity of anti-endomysium antibodies was better than that of anti-tissue transglutaminase antibodies, anti-reticulin antibodies and anti-gliadin antibodies but in children aged less than 2 years, the sensitivity of anti-gliadin antibodies was better than that of anti-tissue transglutaminase antibodies.     

Determination of herpes simplex virus type-specific antibodies by enzyme-linked immunosorbent assay.

We determined type-specific antibodies to herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) by an indirect enzyme-linked immunosorbent assay, using as antigens HSV-1 glycoprotein gC-1 and a HSV-2-specific polypeptide purified on affinity columns of monoclonal antibodies. All sera were initially screened for HSV antibodies by the enzyme-linked immunosorbent assay with a pool of Triton X-100-extracted antigens of HSV-1- and HSV-2-infected HEp-2 cells. The titer of HSV antibodies was predicted from a linear regression curve based on the absorbance of the initial 1:50 serum dilution. The sensitivity and specificity of the screening assay and of the assay for type-specific antibodies were established.     

Detection of antibodies to influenza virus M protein by an enzyme-linked immunosorbent assay.

An enzyme-linked immunosorbent assay test system was developed in which purified influenza virus M protein was used for the detection of M antibody in human sera. Antibody levels to influenza A virus M protein were monitored in sera from a vaccine study population by using an enzyme-linked immunosorbent assay technique with purified M protein as the adsorbent antigen. A 10-fold variation in titers of preexisting M antibody was observed in this population of young adults. Increases of anti-M titer of 7- to 24-fold were observed upon immunization with Formalin-inactivated vaccine or after natural infection. The antibody response to M protein was dissociated from the response to the hemagglutinin or neuraminidase antigens. The M antibody response preceded or was coincident with the antibody response to H1 hemagglutinin upon natural exposure to circulating virus.     

Comparison of immunodiffusion and enzyme linked immunosorbent assay for antibodies to four Aspergillus species.

Antigenic extracts were prepared from Aspergillus fumigatus, A niger, A flavus and A terreus for use in enzyme linked immunosorbent assay (ELISA) and immunodiffusion (ID) tests for Aspergillus antibodies to determine whether the use of antigenic extracts from species other than A fumigatus increased the sensitivity of the ELISA. ELISA titres correlated well with positive ID tests. Patient titres by ELISA were significantly higher than control titres for all species. Patient titres to A niger were also significantly higher than titres to the other species. Total number of ID bands to A fumigatus correlated significantly with anti-A fumigatus ELISA titres. It is concluded that the use of antigenic extracts from species other than A fumigatus improves the sensitivity of the ELISA.     

Detection of aldrin/dieldrin in milk by competitive enzyme‐linked immunosorbent assay (ELISA)

Polyclonal antibodies raised against 6,7‐dihydro‐6‐carboxyaldrin can be used to detect aldrin and dieldrin. These analytes are preferentially fat soluble. An immunoassay is described which provides a method for detecting these pesticides in milk, a fat‐rich matrix. The enzyme‐linked immunosorbent assay (ELISA) can detect aldrin/dieldrin in milk in the range 1 ng ml^‐1‐5 μg ml^‐1 simply and reliably. The detection range differs in skimmed and semi‐skimmed milk and in cream, reflecting the differences in fat content between these samples.     

Detection of rotavirus-specific IgG antibodies by immunoperoxidase assay and enzyme-linked immunosorbent assay

An indirect immunoperoxidase assay (IPA) has been developed for determination of IgG antibodies to rotavirus. The technique employed as antigen, SA-11 infected MA 104 cells, which were air-dried on glass slides and acetone-fixed. In parallel, rota-specific IgG antibodies were determined by enzyme-linked immunosorbent assay (ELISA). Specific IgG antibodies to rotavirus were determined in sera of healthy children and in sera of patients suffering from gastroenteritis. A good correlation (r = 0.92) and (r = 0.98) for healthy children and patients, respectively, was found between IPA and ELISA techniques. The IPA technique is rapid and simple and positive results, because of the intensive staining, are easily read by low-power light microscope. The potential application of IPA and ELISA methods in serodiagnosis of rotavirus infections is discussed.     

Rapid detection of IgG and IgM antibodies for cytomegalovirus by the enzyme linked immunosorbent assay (ELISA)

Summary A simple solid phase enzyme immunoassay for the detection of immunoglobulin G and M to cytomegalovirus (CMV) is described.Using this test IgM antibodies to CMV were detected in 0.7 per cent of newborns and regularly after CMV infection in transplant patients, furthermore in these latter patients IgM production was prolonged for several months. For the determination of IgG the enzyme immunoassay was more sensitive than the complement fixation test (CF) and the antibody titres were 4 to 8 fold higher.Since the ELISA test is rapid, specific and unexpensive it can become an acceptable routine diagnostic procedure.With 1 Figure     

Rapid detection of antibodies against cytomegalovirus induced immediate early and early antigens by an enzyme linked immunosorbent assay

An enzyme linked immunosorbent assay (ELISA) for detecting antibodies against cytomegalovirus induced immediate early antigens and early antigens was developed using purified nuclear antigens and was compared with the indirect immunofluorescence test. The tests were comparable in their ability to detect positive and negative sera, and antibody titres determined by both assays were similar. The use of ELISA for the detection of antibodies against cytomegalovirus induced immediate early and early antigens is advocated in diagnostic and research laboratories.     

Quantitation of antibodies to Toxoplasma gondii in swine sera by enzyme-linked immunosorbent assay

An enzyme-linked immunosorbent assay (ELISA) was developed for quantitation of antibodies to Toxoplasma gondii in swine sera. Because a commercial anti-swine IgG conjugate was directed also against swine IgM, the conjugate was absorbed with the IgM fraction to eliminate the interference of naturally occurring IgM antibodies that appeared consistently in sera collected from slaughtered pigs at an abattoir. The ELISA values of 0.2 or more observed in most of the sera successfully decreased to less than 0.2 by the use of absorbed conjugate. An attempt to use a protein A conjugate has failed. Evaluation of this system by comparing it with the latex agglutination test provided a high significant correlation, indicating its usefulness for serodiagnosis of swine toxoplasmosis.