Detection of IgM antibodies to measles virus by enzyme-immunoassay

A method of solid-phase enzyme-immunoassay (EIA) with horseradish-peroxidase-conjugated anti--globulin was used to determine IgM antibodies to measles virus in human sera. Antigens prepared from measles-infected and noninfected Vero cells passively adsorbed to polystyrene cuvettes were both important for the tests, because sera from many patients convalescing from viral infections and sera from rheumatoid arthritis patients were able to bind to control antigen.Sera from measles patients, patients with other viral infections, rheumatoid arthritis (RA) patients, and from blood donors were tested in a dilution of 1200. Almost all of the acute-phase measles sera (37/38) were positive. The first measles-IgM-negative specimen was found 9 weeks after the onset of rash. Seven percent (15/213) of patients with other viral infections gave positive results in these IgM tests. False-positive reactions were also found in 33 of 51 (65%) sera from RA patients. This nonspecific binding could be abolished by absorptions with latex-IgG particles. This treatment did not have any effect on specific IgM.The IgM-EIA test developed for measles virus antibodies, which requires only a single serum specimen, appears to be a useful diagnostic method for routine virus laboratories.     

The detection of measles specific immunoglobulin M antibodies using biotinylated antigens

The methods of reverse type enzymeimmunoassay (EIAs) with biotinylated antigens were used to determine IgM antibodies to measles virus in human sera. These antigens, either purified measles virus antigen or lysate type measles-vero antigen with lysate vero control antigen, were used in the two separate IgM-tests. Paired sera from 15 measles patients as well as 456 sera from patients with viral infections other than measles, with mycoplasma pneumoniae infections, from rheumatoid arthritis patients and blood donors, were assayed in a dilution of 1:200. Both the test systems detected all the 30 serum specimens from the measles patients as measles IgM positive, but the sera of all the other groups proved to be measles IgM negative. These tests developed for measles specific IgM antibodies, avoiding the interference of IgM-class rheumatoid factor, offer valuable tools for routine virus serology.     

IgM antibody capture haemadherence test (MACHAT) for the detection of measles specific IgM

An antibody capture haemadherence test (MACHAT) for detecting measles-specific IgM is described. The assay is based on the antibody capture principle with rhesus monkey erythrocytes as detector system in place of labelled antisera. MACHAT was compared with a commercial indirect enzyme immunoassay (EIA) for measles-specific IgM using 382 sera from patients notified as measles. There was good agreement between the two tests; 106 sera were found to contain measles IgM by both tests, 7 further sera were positive only in the commercial EIA and 9 only in MACHAT. One sera gave an equivocal result in MACHAT and another in the commercial EIA. Twelve of the 18 sera with discrepant results were also tested by MACRIA; in 7 MACRIA gave the same results as MACHAT, in 3 the MACRIA results agreed with the commercial test and in 2 the MACRIA results were equivocal. Specificity was established by a lack of MACHAT reactivity in sera collected from blood donors (n = 83) and from cases of recent rubella, dengue and parvovirus B19 infection (n = 51). The MACHAT is a simple, cheap test that can be read by eye and is suitable for measles surveillance programmes in the developing world.     

Development of a highly specific and sensitive rubella immunoglobulin M antibody capture enzyme immunoassay that uses enzyme-labeled antigen.

An enzyme immunoassay (EIA) for serum immunoglobulin M (IgM) antibodies to rubella virus based on enzyme labeling of viral antigen was developed. The sensitivity of the EIA for the detection of recent rubella virus infection was evaluated by using 115 rubella-IgM-antibody-positive serum specimens, which were confirmed as positive by Rubazyme M (Abbott Diagnostics). In addition, 12 individuals, 2 of whom were exposed to rubella through vaccination and 10 of whom were exposed through natural infection, were studied, and the results were compared with those obtained by indirect EIA (Rubelisa M; Electro-Nucleonics, Inc.) and immunoblotting. The sensitivity of the newly developed EIA with sera from these individuals was 100%. Serum specimens from two patients indicated that the IgM antibodies were detected by the newly developed EIA at the same time as IgM antibodies were detected by immunoblotting and before positive reactions were detected by an indirect EIA. The reference population consisted of 564 healthy blood donors and hospitalized patients (150 serum specimens). In addition, 145 serum specimens commonly giving false-positive reactions in conventional rubella IgM EIAs were studied. With these specimens, no false-positive reactions were observed. Positive IgM responses, which could not be confirmed by immunoblotting, were observed in two samples from the reference population. However, these two samples were rubella IgG positive. The overall specificity of the EIA was 99.8%.     

High level expression of recombinant mumps nucleoprotein in Saccharomyces cerevisiae and its evaluation in mumps IgM serology.

To develop improved reagents for mumps serology a high-level yeast expression system was employed to produce recombinant mumps nucleoprotein (rNP). The rNP was purified by CsCl gradient centrifugation and yielded approximately 15 mg/l of yeast culture. Electron microscopy of the rNP revealed characteristic herring-bone structures. The electrophoretic mobility of rNP in yeast cells was similar to native NP in SDS-PAGE. Monoclonal antibodies to rNP reacted with native mumps virus nucleoprotein by immunofluorescence assay. A monoclonal antibody to native mumps virus NP reacted with rNP by Western blot assay. The rNP was investigated as antigen in an IgM capture enzyme immunoassay (EIA) using a horseradish peroxidase conjugate of monoclonal antibody to the rNP. Eighteen sera previously found to be positive by IgM capture radioimmunoassay (MACRIA) and 30 sera that were mumps IgM negative by MACRIA were tested by mumps IgM capture EIA. The results for the two test were concordant. In addition, 26 rheumatoid factor positive sera and 35 sera that were IgM positive for measles, rubella or parvovirus B19 were tested. Fifty-nine sera were negative by mumps IgM capture EIA but two sera collected from two infants 3 and 6 weeks after mumps, measles and rubella vaccination were positive. Mumps MACRIA confirmed these results. Compared to MACRIA the overall sensitivity was 100% (20/20) and specificity was 96.8% (30/31). The yeast expressed rNP was highly immunogenic and suitable for use in IgM capture EIA for the diagnosis of mumps.     

Evaluation of five commercial tests for detection of immunoglobulin M antibodies to human parvovirus B19.

The following commercial tests for detection of immunoglobulin M antibodies to human parvovirus B19 were evaluated: Ideia Parvovirus B19-IgM, MRL Diagnostics Human Parvovirus B19 IgM ELISA, Parvoscan-B19, and Biotrin Parvo B19 IgM EIA and IF. A total of 203 serum specimens from patients who probably have current B19 infections or have other viral infections and sera with rheumatoid factor were investigated. Between 75 and 79 of 102 serum samples from patients thought to have current B19 infections yielded positive results with the different tests. Ideia had the highest specificity (94.8%), while Parvoscan showed a specificity of only 70.1%. Our evaluation results show that Ideia, MRL, and Biotrin EIA and IF can be recommended for diagnostic purposes.     

Comparative detection of measles and rubella IgM and IgG derived from filter paper blood and serum samples.

We compared the use of serum and filter paper blood spots as specimen sources for the detection of measles- and rubella-specific IgM and IgG. We collected capillary blood into microtainer tubes and onto filter paper spots from 60 children and 60 healthy adults. The blood was collected from 12-15-month-old children approximately 3 weeks after primary vaccination with measles, mumps, rubella vaccine, and the sample-pairs were tested for measles-specific IgM and IgG antibodies by using a capture antibody EIA and an indirect EIA, respectively. We tested sample-pairs from a subset of participants for rubella- specific IgM and IgG antibodies by using commercially available capture IgM (Captia) and indirect IgG (Wampole) assays. The concordance of results from serum and filter paper blood spots was high for all assays: 98% for measles IgM, 93% for measles IgG, 94% for rubella IgM, and 93% for rubella IgG, and increased to between 96-100% for all four assays when indeterminate samples were excluded. The correlation coefficients for EIA signals were 0.99 and 0.77 for measles IgM and IgG, respectively, and 0.92 and 0.94 for rubella IgM and IgG, respectively. The cut-off values used for filter paper samples were the same as those used for serum samples for all tests except for the rubella IgM assay. The use of filter paper blood spots is a promising future option for the detection of measles- and rubella-specific antibodies.     

Hemadsorption immunosorbent technique for determination of mumps immunoglobulin M antibody

We used hemadsorption immunosorbent technique (HIT) to detect mumps immunoglobulin M (IgM) antibody. IgM from human sera was adsorbed into anti-human IgM-coated wells in plates, and mumps-specific IgM was detected by adding mumps virus hemagglutinin and guinea pig erythrocytes consecutively. Specific IgM-positive sera showed hemadsorption, whereas negative sera showed hemagglutination. All 81 patients with current mumps infections tested showed mumps-specific IgM antibody, with titers ranging from 160 to 327,680. In most cases IgM antibody was already present on day 1 or 2 after the onset of illness. IgM antibody persisted for 6 to 10 weeks. Of 57 patients with acute respiratory illnesses caused by parainfluenza virus, 5 showed cross-reactions in the hemadsorption immunosorbent technique assay for mumps IgM. The hemadsorption immunosorbent technique assay is specific for the IgM class of antibody and avoids false-positive results due to rheumatoid factor. This test is an efficient and sensitive method for rapid and early diagnosis of mumps infections.     

Development and evaluation of a capture enzyme-linked immunosorbent assay for determination of rubella immunoglobulin M using monoclonal antibodies.

A capture enzyme-linked immunosorbent assay (ELISA) for detection of virus-specific immunoglobulin M (IgM) antibody was developed which used a panel of labeled monoclonal antibodies to rubella virus hemagglutinin. The rapidity of the test system was increased by using, after 1-h incubation of the test serum, a second 1-h incubation of the serum with a mixture of viral antigen and labeled monoclonal antibody. The new assay was tested for specificity on 371 human sera from people without any recent contact with rubella virus; of these, 66 were sera selected from people with rheumatoid factor or IgM antibody to human cytomegalovirus, Epstein-Barr virus, or other viruses. In parallel, the new assay was performed on 191 sera from patients having recent contact with rubella virus. Results were compared with those obtained by an indirect ELISA method on IgM serum fractions, using purified rubella virus as a solid phase. Of the 371 sera tested for specificity, 5 (1.3%) gave false-positive results with indirect ELISA (1 rheumatoid factor, 2 heterophil antibody, and 2 human cytomegalovirus sera positive for IgM), and none were false-positive with the capture assay. Two sera from a patient with primary cytomegalovirus infection, which were positive for rubella IgM antibody with both methods and were initially interpreted as false-positive, were finally considered to be true-positive, since they were reactive only in the presence of IgM antibody and viral antigen. Of the 191 sera from 92 patients (84 patients with acute rubella, four newborns from mothers with rubella during pregnancy, and four vaccinees), 136 (71.2%) were found to be positive for IgM by direct ELISA, and 128 (67.0%) were positive by capture ELISA; 12 sera drawn during the first 2 days of disease, or at least 40 days after onset (or after vaccination), were detected only by indirect ELISA, and 4 sera were detected only by capture ELISA. Thus, specificity and sensitivity, respectively, were 100 and 91.4% for capture ELISA and 98.6 and 97.1% for indirect ELISA. However, when the number of patients was considered, 86 were detected as IgM positive by indirect ELISA, and 87 were detected positive by capture ELISA. The overall agreement between the two assays was 96.2%. Capture ELISA using monoclonal antibody appears preferable over indirect ELISA on IgM serum fractions because of its higher specificity and shorter time for test performance; furthermore, there is no need for serum fractionation or virus purification for the capture ELISA.     

Chronic biological false-positive reactions to serological tests for syphilis in blood donors

Problem sera from 375 blood donors were investigated for biological false-positive reactions to serological tests for syphilis. Problem sera are those sera in which submitting laboratories have found a reactive result to a screening test for syphilis. On each serum a cardiolipin Wassermann reaction, a Venereal Disease Research Laboratory test, a Reiter protein-complement fixation test, a Treponema pallidum immobilization test, and a fluorescent treponemal antibody absorption test were performed.Of the sera 49.9% were found to be non-reactive in all five tests, 28.8% gave results indicating a diagnosis of syphilis, and 21.3% showed biological false-positive reactions.There were 80 sera from blood donors which gave biological false-positive reactions. A further specimen of serum from 67 of these donors was tested after an interval of a minimum of six months. Sixty-six of the sera showed chronic biological false-positive reactions. Some donors were only found reactive for the first time after they had given very many donations of blood.It is concluded that sudden blood loss, as in blood donation, appears to stimulate the production of excess reagin in certain individuals, causing a chronic biological false positive reaction to serological tests for syphilis. This may only appear after several blood donations have been made.     

Evaluation of commercial methods of enzyme immunoassay (EIA) for the measurement of rubella-specific IgM

Four commercial EIA methods for measuring rubella-specific IgM (three indirect tests and one anti-mu capture test) were evaluated, using sucrose gradient centrifugation and hemagglutination inhibition as the reference method. Evaluation was conducted with the aid of four serum panels, including 53 primary rubella cases, 30 healthy pregnant women, 21 sera positive for rheumatoid factor(s) (RF) and 35 sera from 29 cases of heterophil-positive infectious mononucleosis with EBV-specific IgM detected by immunofluorescence. All EIA methods were more sensitive than the reference method when applied to very early samples (1-5 days post-exanthema) and no differences in sensitivity were found between them. On the other hand, we observed a significant incidence of false-positive results if an indirect EIA method is applied to RF-positive samples. False positivity is significantly reduced, but not totally eliminated, when samples are preabsorbed with anti-human IgG serum and, in all cases, the absorbance values obtained were low. In contrast, there were no false-positive results using an anti-mu capture method, even in sera from cases of infectious mononucleosis. The basis for choosing between an indirect method and an anti-mu capture method for the diagnosis of congenital and post-natal rubella virus infection is discussed.     

Comparative evaluation of commercial Premier EIA and microimmunodiffusion and complement fixation tests for Coccidioides immitis antibodies.

A total of 409 serum and cerebrospinal fluid specimens from human subjects with proven coccidioidomycosis, with other infections, or with no apparent illness were tested for antibodies to Coccidioides immitis by the Premier EIA (Meridian Diagnostics, Inc., Cincinnati, Ohio), which tests for immunoglobulin G (IgG) and IgM responses to coccidioidal antigens, and by the conventional complement fixation (CF) or immunodiffusion (ID) assays for antibodies corresponding to those detected by the tube precipitin (TP) or CF tests. Of the 409 specimens, 47 were from persons with confirmed coccidioidomycosis and all were positive for C. immitis antibodies in IDCF tests and enzyme immunoassays (EIAs) for both IgG and IgM. The EIA for detecting both IgG and IgM antibodies proved to be sensitive for detecting coccidioidomycosis case sera positive by the IDCF, IDTP, and CF tests. Maximal sensitivity for diagnosing coccidioidomycosis is dependent upon detection of both IgG and IgM antibodies in the EIA. The EIA, however, was not absolutely specific, since some sera from patients with confirmed blastomycosis and some from patients with noncoccidioidal disease produced false-positive reactions.     

Comparative evaluation of tests for detection of parvovirus B19 IgG and IgM

The aim of this study was to evaluate enzyme immunoassays (EIA) (Euroimmun, Lübeck, Germany) and chemiluminiscent immunoassays (CLIA) (Diasorin, Saluggia, Italy) in their application to detect B19V‐IgM and ‐IgG. For this purpose, one hundred and ninety samples were studied. Of them, 101 came from recent infection cases (B19V‐specific IgM (86) and/or PCR (87), 42 from past infections, 18 from non‐infected, and 29 from other viral recent infections (Epstein‐Barr virus, measles, and rubella). Samples were characterized by capture (for IgM), or indirect (for IgG) EIA (Biotrin, Dublin, Ireland); indeterminate samples were classified by indirect immunofluorescence (IIF) (Biotrin). All the samples were used for testing IgM assays, and all but the cases from other viral infections were used for IgG tests. For IgM, CLIA, and EIA identified 76 and 62 of 86 IgM positives, respectively (sensitivity 88.4% and 72.1%). Considering B19V IgM negative samples, negative result was obtained in 95 and 92 of 104, being the specificity values of CLIA and EIA 91.3% and 88.5%, respectively. For IgG, CLIA and EIA identified correctly 114 and 115 of the 122 positive samples (sensitivity 93.4% and 94.3%, respectively), and 39 and 36 of 39 negative samples (specificity 100% and 92.3%). As conclusion, CLIA methods can be used in clinical laboratories as adequate alternatives to the well‐established Biotrin EIAs.     

Mechanisms of smooth muscle antibody production: a clinical study in children with infections, haemolytic syndromes, and idiopathic thrombocytopenic purpura.

Sera from 530 children suffering from various diseases and from 64 controls were tested for smooth muscle autoantibodies (SMA) by indirect immunofluorescence. A high incidence of SMA (51-86%) was found in patients with viral and bacterial infections (viral hepatitis, infectious mononucleosis, measles, mumps, chickenpox, typhoid fever, and brucellosis), independently of liver invovlvement, and in patients with acute haemolytic anaemia due to G-6-PD deficiency (48%). By contrast, the incidence of SMA from patients with beta-thalassaemia major and idiopathic thrombocytopenic purpura was no higher than in the controls. The discrepancy in incidence in haemolytic anaemias due to different causes may reflect the effect of endogenous and extrinsic agents. In the viral infections, SMA were mainly of the IgM class and gave an 'SMA-V' staining pattern. In bacterial infections (typhoid fever and brucellosis), SMA were either IgG only or IgM and IgG, and the staining pattern was also mainly 'SMA-V'. In infections which affect or may affect the liver (viral hepatitis, infectious mononucleosis, typhoid fever, and brucellosis), SMA was present at high titres (1:80-1:320), whereas in infections not affecting the liver (measles, mumps, and chickenpox) the titres were lower (less than or equal to 1:80). In most patients SMA occurred transiently and without apparent pathogenetic significance. The antigen against which infection-induced SMA is directed is not actin; its nature has yet to be identified.     

Simultaneous IgM reactivity by EIA against more than one virus in measles, parvovirus B19 and rubella infection.

BACKGROUND: A clinical diagnosis of rash-causing infections is not always possible and reliance has to be placed on serological evidence of infection, especially on the presence of specific immunoglobulin (Ig)M. However, despite the use of modern serological methods and validated commercial kits, reports appear in the literature of simultaneous IgM reactivity against more than one virus in cases of Epstein Barr virus, rubella, cytomegalovirus, human parvovirus B19 (HPV B19) and measles infections, all with implications for the pregnant woman. OBJECTIVES: We decided to evaluate the extent of the problem in rubella, measles and HPV B19 infections in a routine diagnostic laboratory. STUDY DESIGN: We tested sera from cases with initial clinical and serological evidence of infection with measles, HPV B19 or rubella for evidence of simultaneous IgM reactivity against more than one virus. We confirmed primary infections with specific-IgG antibody avidity tests, and subjected sera with IgM reactivity against more than one virus to avidity tests to identify which, if any, of the three viruses was the cause of the primary infection. Groups of monoreactive IgM sera were randomly selected from the presented sera to demonstrate that the avidity of the IgG specific for the other two viruses would be of high avidity compared with the low avidity of the IgG specific for the virus against which specific IgM had been detected. RESULTS: Our results confirm that simultaneous IgM reactivity against more than one virus does occur in these three infections, and that this is unlikely to be caused by the presence of rheumatoid factor. CONCLUSIONS: In the absence of seroconversion, reliance on specific IgM results alone for diagnosis of these infections should be avoided and tests such as specific IgG antibody avidity should also be employed. The simultaneous occurrence of IgM reactivity against more than one virus is also important for epidemiological and surveillance reasons as the widespread use of the mumps, measles and rubella vaccine makes its impact on the population. Falsely diagnosed cases of apparent measles or rubella could throw into question the efficacy of the vaccine.     

Enzyme-linked immunosorbent assay for detection of immunoglobulin M antibody against the Reiter treponeme flagellum in syphilis

An enzyme-linked immunosorbent assay (ELISA) for detection of immunoglobulin M (IgM) antibodies against the periplasmatic flagellum of the Reiter treponeme is described. IgM in the test samples was bound to anti-IgM-coated microtest plates, and flagellum-specific IgM antibody was subsequently detected by incubation with a purified flagellum preparation and monospecific anti-flagellum conjugate. Rheumatoid factor, antinuclear antibodies, or flagellum-specific IgG did not interfere. The specificity of the ELISA for IgM antibodies was 99.5% for sera from 200 blood donors and 98.6% for 147 patient sera that gave false-positive reactions in other syphilis serological tests. The sensitivity was 88.5% for sera from 87 patients with first-time primary syphilis, 93.5% for sera from 62 patients with first-time secondary syphilis, 21.4% for sera from 42 patients who were reinfected, and 0% for sera from 13 patients with late syphilis. Of the sera from 153 patients with treated syphilis, 7.2% had IgM antibodies, and sera from patients with primary or secondary syphilis generally had no IgM antibodies 6 months after treatment. The finding of IgM antibodies indicates that patients should receive antisyphilis treatment if they have not been treated recently, but a negative result does not exclude the possibility of active syphilis. The method may prove useful for the diagnosis of congenital syphilis in newborns.     

Accuracy of immunoglobulin M immunoassay for diagnosis of chlamydial infections in infants and adults

An improved solid-phase enzyme immunoassay (EIA) with Chlamydia trachomatis L2 434/Bu elementary bodies was developed for the measurement of immunoglobulin M (IgM) antibody to C. trachomatis in serum. Comparison of EIA and microimmunofluorescence IgM antibody titers of 156 serum samples revealed an EIA sensitivity and specificity of 100% for infants, but reduced sensitivity (85%) and specificity (76%) for sera from adults. Sera containing IgM class rheumatoid factor produced false-positive IgM results which could easily be eliminated by pretreatment of the sera with anti-human IgG. Analysis of sera from infants with chlamydial infections revealed that 17 of 17 infants with C. trachomatis pneumonia had high IgM antibody titers (geometric mean titer, 1:64,812), whereas two infants with conjunctivitis only lacked detectable IgM antibody. EIA detected IgM antibody to several serovar groups in serum, including serovars B, BDE, FG, and J. IgM antibody to C. trachomatis in serum was detected as early as 5 days after the infection that was acquired at delivery and persisted for 3 months. The availability of an EIA possessing good sensitivity and specificity for the detection of IgM antibody to C. trachomatis may permit more laboratories to diagnose perinatal chlamydial infections.     

Enzyme-linked immunosorbent assay (ELISA) for detection of herpes simplex virus-specific IgM antibodies

Herpes simplex virus (HSV)-specific IgM in human serum could be detected by a microplate enzyme-linked immunosorbent assay, using extracts of HSV-infected cells as antigen. Peroxidase-conjugated anti-human IgM was used to detect human IgM bound to viral antigen. Pretreatment of sera with protein A-bearing staphylococcus or with aggregated human IgG was necessary to eliminate false-positive results caused by the presence of rheumatoid factor. Specificity controls included sera of patients with other herpes group virus infections.     

Detection of measles,mumps, and rubella antibodies in saliva using antibody capture radioimmunoassay

Antibody capture radioimmunoassays were developed for detecting virus specific IgM (MAC-RIA) and IgG (GACRIA) to measles, mumps, and rubella and used to investigate saliva as an alternative specimen to serum for diagnosis. Saliva was collected from 63 patients with measles, 19 with mumps, and 150 with rubella, which were all clinically diagnosed and serologically confirmed. Virus specific IgM was detected in 92% of measles, 75% of mumps, and 100% of rubella saliva samples collected during the first week of illness. Between 1 and 5 weeks after onset virus specific IgM was detected in 100% of saliva specimens. After the 5th week the proportion of reactive specimens declined. The specificity of the MACRIA tests was established by testing saliva samples collected from blood donors for measles (88), mumps (88), and rubella IgM (91). All of the saliva specimens tested for measles and rubella specific IgM were unreactive, 1/88 specimens tested for mumps specific IgM contained significant reactivity. Saliva specimens collected from acute cases of MMR were tested in all 3 MACRIAs. A small proportion of saliva samples contained detectable IgM of more than one virus infection. Rubella and measles specific IgG was detected in the saliva of all cases from the 4th or 5th day of illness, respectively. Detection of mumps specific IgG was less successful. We have demonstrated that virus specific IgM can be reliably detected in saliva samples collected from acute cases of measles, mumps, and rubella and identified 1–5 weeks after onset of illness as the optimum time for collection of samples. © 1993 Wiley-Liss, Inc.     

Enzyme-linked immunosorbent assay for mumps IgM antibody: comparison of IgM capture and indirect IgM assay

We have established two enzyme-linked immunosorbent assays (ELISAs) for detection of mumps IgM antibody, i.e., indirect IgM ELISA and IgM capture ELISA, for serodiagnosis of recent mumps infection. In the latter method, peroxidase-conjugated monoclonal antibody to mumps virus was employed. Both methods detected mumps antibody of IgM class only in serum fractions separated by centrifugation through a sucrose density gradient. Optical density values given by both ELISAs were correlated for most sera examined. Indirect IgM ELISA, however, gave a false positive reaction for sera containing both rheumatoid factor and mumps IgG antibody, while giving a false negative reaction for sera containing high titers of mumps IgG antibody. This technique was, therefore, less reliable than IgM capture ELISA. IgM antibody detectable by IgM capture ELISA was present in all patients with mumps by the fifth day of illness and persisted for up to 3 mth in most and up to 5 mth in same cases.