Comparison of antibody, antigen, and metabolite assays in rat models of systemic and gastrointestinal candidiasis.

We compared serial measurements of antibodies to mannan and to a cytoplasmic antigen (enzyme-linked immunosorbent assays), detection of mannan and an unidentified candidal antigen (latex agglutination), and assays of mannose and arabinitol (gas chromatographic assay of per-O-acetylated aldonitrile derivatives). In a high-inoculum intravascular-infection model, antimannan assays were consistently positive beginning on day 2 postinoculation, anti-cytoplasmic antigen assays followed the same time course but were less sensitive, mannan was detected in all samples beginning on day 2 postinoculation, and serum mannose concentrations peaked on day 3 postinoculation and were less sensitive than mannan detection. Other assays were not useful. In a lower-inoculum intravascular-infection model, the antibody assays became positive after a similar interval and remained positive for 28 days, with antimannan again being the more sensitive. Mannan and mannose tests were positive in week 1 postinoculation only, with mannan detection being the more sensitive. In a gastrointestinal-colonization model, antimannan assays become positive after 2 weeks of colonization, whereas anti-cytoplasmic antigen and mannan tests remained negative. In a model of gastrointestinal colonization followed by invasive infection produced by induction of neutropenia, only mannan detection was diagnostically useful. These data, comparing this panel of modern serodiagnostic techniques in controlled models of clinically relevant syndromes of candidiasis, enhance understanding of previous efforts in serodiagnosis of candidiasis and provide a foundation for further prospective studies in patients.     

Value and limitations of anti-mannan antibodies research by co-immunoelectrodiffusion in the diagnosis of deep candidiasis

Several groups have evaluated detection of antibodies against Candida, with somewhat conflicting results. In this study, co-counterimmunoelectrodiffusion was used to detect antimannan antibodies specific of components of the Candida membrane. Study patients were divided into two groups according to whether their history for Candida infection was negative (population A, n = 102) or positive (population B). Different antigen levels were used in order to differentiate low and high antimannan antibody levels. Among the 102 sera in population A, 42 were positive for antimannan antibodies; the antimannan antibody titer was low in 40 cases and high in 2 cases. In population B (53 patients), antimannan antibodies were found in 97 of the 98 sera studied; titers were high in 95 cases. Use of an antigen level that detects only high titers of antimannan antibodies thus provides a sensitive and specific tool for the diagnosis of deep candidiasis. The simplicity and rapidity of this test are particularly valuable in situations where emergency treatment is needed.     

Comparison of enzyme immunoassay and gas-liquid chromatography for the rapid diagnosis of invasive candidiasis in cancer patients.

Three proposed quantitative markers for candidiasis, arabinitol, mannose, and mannan in serum, are compared in 50 normal blood donors and 38 high-risk patients, 23 with and 15 without invasive candidiasis. Arabinitol concentrations in serum, the arabinitol/creatinine ratio, and mannose concentrations in serum were significantly greater in the 15 patients without candidiasis than in the normal blood donors (P less than 0.05). The sensitivities and specificities were 26 and 87% for arabinitol, 13 and 93% for the arabinitol/creatinine ratio, and 39 and 87% for mannose. On the other hand, mannan concentrations in serum were less than 1 ng/ml in normal blood donors and patients without candidiasis (P = 0.344), and the sensitivity and specificity were 65 and 100%, respectively. Of 23 patients with proven or probable candidiasis, 16 had mannan levels in serum greater than the mean + 2 standard deviations (0.46 ng/ml) for the 15 controls. In 16 patients with invasive candidiasis and positive blood cultures for the Candida spp., only 13 had elevated levels of at least one of the three markers. The arabinitol/creatinine ratio, the mannose level, and the mannan level became elevated an average of 4 days before, 1 day before, and on the same day that the blood cultures were drawn, respectively. Conversely, mannan was detected in the sera of six of seven patients with invasive candidiasis and negative blood cultures. We conclude that the best approach to diagnosing invasive candidiasis involves obtaining blood cultures and carrying out serial assays for mannan in serum.     

Diagnosis of invasive candidiasis by detection of mannan antigen by using the avidin-biotin enzyme immunoassay.

The diagnosis of invasive candidiasis was attempted by detection of circulating mannan antigen by using an avidin-biotin-amplified enzyme-linked immunosorbent assay (AB-ELISA), and this method was compared with the conventional culture method. Mannan antigen was detected by AB-ELISA in the sera of 16 (84.2%) of the 19 patients with invasive candidiasis. On the other hand, for 34 immunocompromised candidiasis-free patients, including 8 with aspergillosis or cryptococcosis, mannan antigen was positive during only 1 of the 67 febrile episodes and in the serum of none of the 50 outpatients without infections. The results were also negative for all patients with deep-seated mycoses other than candidiasis. However, the mannan level was low (less than 2.0 ng/ml) in the serum of 63.2% of the patients with invasive candidiasis. The positivity rate of blood cultures was 31.6%, and that of blood cultures and/or cultures of samples from sterile sites combined was 47.4%. The advantages of the diagnosis based on antigen detection by AB-ELISA are considered to be a higher sensitivity and elimination of nonspecific reactions by the introduction of the avidin-biotin system and pretreatment of sera by heating. In addition, it is considered essential for high sensitivity that transient mannan antigenemia be determined frequently so that it is not overlooked. In light of its sensitivity and specificity, this method is considered to be clinically useful in the diagnosis of invasive candidiasis.     

Quantitation of antibody against cell wall mannan and a major cytoplasmic antigen of Candida in rabbits,mice, and humans.

Cell wall mannan of type A Candida albicans was purified, conjugated with tyramine, and labeled with 125I. Labeled cell wall mannan was used in a radioimmunoassay to measure serum antimannan antibody levels. An ammonium sulfate-soluble fraction of a cytoplasmic extract of C. albicans contained a large amount of a major cytoplasmic antigen of this organism. When the sulfate-soluble fraction was labeled with 125I, much more 125I attached to this major antigen than to the other antigens present in the sulfate-soluble fraction. Thus, when serum antisulfate-soluble fraction antibody levels were measured by a radioimmunoassay which used the iodine-labeled sulfate-soluble fraction, antibody against this major cytoplasmic antigen was quantitated. Both radioimmunoassays were used to measure antimannan and antisulfate-soluble fraction antibody levels in mice, rabbits, and humans. Irrespective of the procedure used to elicit antibody against C. albicans antigens, mice failed to produce antimannan antibody. By contrast, all strains of mice tested produced antisulfate-soluble fraction antibody after immunization, and the magnitude of this antibody response depended on the strain of mice immunized. Rabbits readily produced antibody against both mannan and sulfate-soluble fraction when immunized by a variety of methods. Antimannan antibody was detected in 100% of sera from a randomly selected sample of 50 hospitalized patients. Only 1 of 50 patients had antisulfate-soluble fraction antibody detectable by radioimmunoassay. In pooled normal human serum, most antimannan antibody was of the immunoglobulin G class.     

Diagnosis of systemic candidiasis by latex agglutination for serum antigen.

Three latex agglutination test procedures for detecting Candida antigen in human serum were compared in a retrospective study of 69 patients and 20 normal volunteers. Untreated human serum was reacted with two different latex reagents; one reagent also was reacted with serum treated with protease and heat. The test procedure with treated serum was best, detecting serum antigen in 17 of 21 patients (81%) with disseminated candidiasis. Judging by autopsy-proven cases, there was an increase in positive test results in the last 2 weeks of life. When untreated sera were tested with this reagent, only 3 (14%) of the 21 patients with disseminated candidiasis had detectable antigen in serum. A subset of these same sera was tested by a commercial latex reagent (Candida Detection System lot C001; Ramco Laboratories, Inc., Houston, Tex.) and untreated serum. Of 18 patients with disseminated candidiasis, 5 (28%) had at least one positive serum. Sera from patients with less severe clinical forms of candidiasis were usually negative regardless of the test procedure used. With one exception, sera from control patients were negative or were positive only in sera containing rheumatoid factor. Latex agglutination tests for Candida spp. in treated serum may prove to be a useful procedure for the rapid diagnosis of severe disseminated candidiasis.     

Contribution of the Platelia Candida-specific antibody and antigen tests to early diagnosis of systemic Candida tropicalis infection in neutropenic adults

The Platelia Candida-specific antigen and antibody assays (Bio-Rad Laboratories) were used to test serial serum samples from seven neutropenic adult patients with hematological malignancies who had developed systemic Candida tropicalis infections. The diagnosis of candidiasis was based on a positive blood culture (all seven patients) and the isolation of C. tropicalis from a normally sterile site (six patients). All patients received early antifungal therapy with amphotericin B and/or an azole derivative and had successful outcomes. When the combined assays were applied to sera collected at different time points before and after the first positive blood culture, all patients tested positive. In six patients, at least one positive test was obtained with sera collected, on average, 5 days (range, 2 to 10 days) prior to the first positive blood culture, while blood cultures were constantly negative. High and persistent mannanemias were detected in all patients during the neutropenic period. In five patients, an increased antibody response was detected when the patients recovered from aplasia. Controls consisted of 48 serum samples from 12 febrile neutropenic patients with aspergillosis (n = 4), bacteremia (n = 4), or no evidence of infection (n = 4). A low level of mannanemia was detected in only one serum sample, and none showed significant Candida antibody titers. Our data thus confirm the value of the combined detection of mannanemia and antimannan antibodies in individuals at risk of candidemia and suggest that in neutropenic patients, an approach based on the regular monitoring of both markers could contribute to the earlier diagnosis of C. tropicalis systemic infection.     

Comparison of antibody, antigen, and metabolite assays for hospitalized patients with disseminated or peripheral candidiasis.

Repeat serum samples from 22 patients with proven disseminated candidiasis and 42 with simple peripheral colonization were assayed for Candida antibodies by coelectrosyneresis, immunoprecipitation, and A and B immunofluorescence, for metabolites by D-arabinitol measurement, and for antigens by the mannan immunoassay and Cand-tec latex agglutination (mean number of samples tested, 2.5 per patient). For the antibody and metabolite assays, the results showed no statistical difference between the two groups. By contrast, the results of both antigen assays were positive for a significantly larger number of patients with disseminated candidiasis than of those with simple peripheral colonization. Results were regardless of whether the patients were neutropenic. They were not predictive of death. We calculated that the mannan antigen assay had 29% sensitivity and 97% specificity for the diagnosis of disseminated candidiasis. Likelihood ratios of a positive and a negative result of this test were 9.2 and 0.7, respectively, for this diagnosis. In the latex agglutination test, likelihood ratios were 2.5, 1.5, 1.6, and 0.3 when the test was positive for dilutions of 1/8, 1/4, and 1/2 and was negative, respectively.     

Combined detection of mannanaemia and antimannan antibodies as a strategy for the diagnosis of systemic infection caused by pathogenic Candida species

A novel strategy for the diagnosis of systemic candidosis was evaluated, based on the combination of two enzyme immunoassays that detect a candida oligomannoside repetitive epitope expressed in large amounts by Candida albicans (Platelia Candida Ag), and antibodies against C. albicans mannan, the major cell-wall immunogen in which this epitope is present (Platelia Candida Ab). Sera were selected retrospectively from intensive care and haematology patients with clinically suspected systemic candidosis, and from whom Candida spp. had been isolated from normally sterile sites. Of the 21 patients infected with C. albicans, 13 had positive antigenaemia and 14 had a positive antibody response, including eight patients who were antigenaemia negative. The sensitivity of the combined tests was 100%. In patients infected with C. glabrata (n = 12) or C. tropicalis (n = 10), the sensitivity was 83% and 80%, respectively. For the remaining patients, infected with C. parapsilosis (n = 10), C. krusei (n = 8) or C. kefyr (n = 2), the sensitivity of the combined tests was 40%, 50% and 50%, respectively. At least one of the serological tests was positive before yeast growth occurred in 60% of patients for whom a serum sample was available before blood culture sampling. An increase in serological test positivity to >80% was observed for sera obtained around the date of positive culture, irrespective of the Candida species isolated. These results suggest that regular serological monitoring for both mannanaemia and anti-mannan antibodies in at-risk patients may contribute to the early diagnosis of candidosis.     

Comparative serological and cutaneous reactivity of candidal cytoplasmic proteins and mannan separated by affinity for concanavalin A.

Yeast-form Candida albicans cells were disrupted for 1.5 min in a Braun homogenizer and centrifuged at 100,000 X g. The supernatant was concentrated by ammonium sulfate precipitation and then dialyzed. The resulting material (650 mg), containing 81.2% protein and 11.5% carbohydrate, was subjected to affinity chromatography on concanavalin A (Con A) linked to agarose. A protein fraction was eluted from the column with buffer, and a fraction containing mannan was eluted with 0.2 M alpha-methyl mannoside. The candidal soluble proteins had 19 components which were resolvable by polyacrylamide gel electrophoresis.The material with affinity for Con A contained mannan and 17% complexed protein. Antigenic differences between the soluble proteins and the mannan-protein complex were shown by lines of intersection in immunodiffusion. The soluble proteins devoid of mannan reacted in immunoelectrophoresis with sera from infected rabbits and patients with chronic candidiasis. These same sera also reacted with a mannan-protein complex eluted from the Con A column with alpha-methyl mannoside. The comparative ability of candidal proteins and cell wall-derived mannan to elicit skin test reactions in guinea pigs sensitized by infection or with formaldehyde-killed yeast was studied. Candidal proteins at a 10-mug dose elicited positive reactions at 6 and 21 days after sensitization. The reactions persisted for 48 h and showed minimal tendency to an arthus response, which was marked when mannan-containing antigens were used. The antigenicity of cell wall-derived mannans and candidal soluble proteins devoid of mannan was compared in immunodiffusion tests of sera from 39 patients with neoplastic disease. Of these patients with documented candidiasis, 13 of 20 reacted to one or more mannan antigens, and 3 of 20 reacted to candidal soluble proteins. In contrast, of those patients who were uninfected or had superficial Candida spp. infections, 5 of 19 reacted to candidal soluble proteins, and 16 of 19 reacted to one or more mannan antigens.     

Biological activities of naturally occurring antibodies reactive with Candida albicans mannan

Sera from normal adult humans may contain high levels of antibody reactive with Candida albicans mannan. This study examined selected biological activities of such antibodies, focusing on sera that were collected from 34 donors and analyzed individually. The results showed that antimannan titers were normally distributed. Reactivity as determined by enzyme-linked immunosorbent assay with serotype A mannan generally paralleled reactivity with serotype B. Analysis of the kinetics for activation of the complement system and deposition of complement component 3 (C3) onto serotype A and serotype B cells showed a decrease in the lag time that occurred before the onset of rapid accumulation of C3 that correlated with increasing antimannan titers. In contrast, there was a decrease in the overall rate of accumulation of C3 on serotype A cells that was strongly correlated with increasing antibody titers; serotype B cells showed no such decrease. An evaluation of the contribution of mannan antibody to opsonophagocytic killing showed that mannan antibody in individual sera and antimannan immunoglobulin G (IgG) affinity purified from human plasma contributed to killing by neutrophils in a dose-dependent fashion in the absence of a functional complement system. However, affinity-purified antibody in very high concentrations was inhibitory to both complement-dependent and complement-independent opsonophagocytosis, and this finding suggests a prozone-like effect. In contrast, if the complement system was functional, antimannan IgG was not needed for opsonophagocytic killing. These results suggest that naturally occurring mannan antibodies and the complement system are functionally redundant for opsonophagocytic killing by neutrophils.     

Collaborative evaluation of antigen detection by a commercial latex agglutination test and enzyme immunoassay in the diagnosis of invasive candidiasis.

The Cand-Tec Candida detection system and enzyme immunoassay for serum mannan were retrospectively compared in a controlled collaborative evaluation of antigen detection in 32 patients with candidiasis proven by biopsy or culture from a normally sterile site and with sera drawn within 7 days of inclusion. With a threshold titer of 1/8, which excluded false-positive results in 17 hospitalized patients without candidiasis, sensitivities for all 32 patients with candidiasis were 44% for the Cand-Tec assay and 17% for the enzyme immunoassay. Both assays provided greater sensitivity when sera were drawn within 24 h of inclusion in the study and in the category of patients with invasive candidiasis (57% by Cand-Tec and 33% by enzyme immunoassay). The Cand-Tec assay gave false-positive results (titer, greater than or equal to 1/8) in 4 of 6 patients with transient candidemia, in 1 of 20 otherwise healthy patients with rheumatoid factor, and in 1 patient with a positive cryptococcal latex agglutination test. Three serum specimens from 3 of 32 patients with candidiasis contained rheumatoid factor and gave titers of greater than or equal to 1/8 by the Cand-Tec assay. Detection of serum mannan by enzyme immunoassay was less sensitive but more specific than the Cand-Tec Candida detection system for the diagnosis of invasive candidiasis.     

Clearances ofCandida albicans-derived α- and β-linked mannose residues in sera from patients with candidiasis

Two monoclonal antibodies (MAbs) reacting with differentCandida albicans mannan oligomannosidic epitopes were used to detect antigens in human sera. The first MAb reacted with -linked mannose residues shared by mannoproteins, and the second displayed reactivity against -1,2-linked oligomannosides shared by phospholipomannan. Two latex agglutination tests performed after dissociation of serum immune complexes were 100% specific. In a retrospective analysis of 39 sera from 20 patients with candidiasis, each test detected seven patients; in combination, they detected ten. More than 60% of positive samples were positive with only one test. These results demonstrate that the clearance ofCandida albicans antigens from the blood differs according to the type of oligomannoside and glycoconjugate. Antigen detection kits with MAbs to differentCandida albicans mannan epitopes could provide a logical rationale to compensate for the transitory character of mannanemia detection during candidal infections.     

Analysis of complement fixation and commercial enzyme immunoassays for detection of antibodies to Mycoplasma pneumoniae in human serum

The Meridian ImmunoCard (IC), GenBio ImmunoWELL-IgM, and Remel EIA commercial antibody tests are qualitative enzyme immunoassays that detect antibodies to Mycoplasma pneumoniae in serum. These tests were compared to an M. pneumoniae complement fixation (CF) assay, which uses a commercially available antigen component. The Meridian IC and the ImmunoWELL-IgM detect immunoglobulin M (IgM) only; the Remel EIA and the CF test detect both IgM and IgG antibodies. Detection of specific IgM antibody, which appears early in infection, can be, but is not always, indicative of a recent or current infection. Paired serum samples from 64 adult patients with probable M. pneumoniae infection were examined with each of the four tests. Thirty (47%) of the 64 acute-phase sera were IgM positive by Meridian IC, 26 (41%) were positive by Remel EIA, 24 (38%) were positive by CF, and 15 (23%) were positive by ImmunoWELL-IgM. When both the acute- and convalescent-phase serum samples from each patient were examined, 61 (95%) of the 64 patients were positive by CF, 60 patients (94%) were positive by Remel EIA, 52 patients (81%) were IgM positive by the Meridian IC, and 29 patients (45%) were IgM positive by the ImmunoWELL-IgM assay. The Meridian IC was more sensitive than the other tests for early detection of IgM antibodies. However, after examining paired serum samples, we concluded that the detection of IgM alone may not be useful for all cases of mycoplasma infection, especially in an adult population.     

Enzyme-linked immunosorbent assay measurement of fluctuations in antibody titer and antigenemia in cancer patients with and without candidiasis.

Antibody titers against purified sulfate-soluble fraction (PSSF) obtained from cytoplasmic extracts of Candida albicans were determined retrospectively over a 2-year period for 123 cancer patients by enzyme-linked immunosorbent assay. Antibody against cell wall mannan (CWM) was also measured by the hemagglutination test and the production of precipitins by a serum interacting with a yeast cell homogenate by immunodiffusion. Invasive candidiasis determined by histological evidence at autopsy was present in 10 patients. Fourfold or greater rises in anti-CWM and anti-PSSF antibodies were detected for eight of the patients with invasive candidiasis at 14 to 22 days after the onset of fever. The immunodiffusion test was positive for four patients with invasive candidiasis. For patients with no evidence of candidiasis, significant rises in anti-CWM and anti-PSSF antibodies were observed at a frequency of 20 and 10%, respectively. The concentrations of serum mannan were sequentially measured by the enzyme-linked immunosorbent assay. Antigenemia (greater than or equal to 3 ng/ml) was found in 9 of the 10 patients with invasive candidiasis and in 2 of the 4 patients with thrush, whereas the serum of 1 of the 36 patients with no evidence of candidiasis was positive for antigen. The first antigenemia antedated significant rises in antibody levels against Candida species by 6 to 23 days.     

Human recombinant antimannan immunoglobulin G1 antibody confers resistance to hematogenously disseminated candidiasis in mice

Mannan is a major cell wall component found in Candida species. Natural antimannan antibody is present in sera from most normal adults, but its role in host resistance to hematogenously disseminated candidiasis is unknown. The purpose of this study was to develop recombinant human antimannan antibody and to study its protective function. A phage Fab display combinatorial library containing Fab genes from bone marrow lymphocytes was screened with Candida albicans yeast cells and chemically purified mannan. One antimannan Fab, termed M1, was converted to a full-length immunoglobulin G1 antibody, M1g1, and M1g1 was produced in CHO cells. The M1g1 epitope was found in C. albicans serotypes A and B, Candida tropicalis, Candida guilliermondii, Candida glabrata, and Candida parapsilosis. Its expression was active at both 23 degrees C and 37 degrees C and uniform over the cell surface. BALB/c mice passively immunized with M1g1 were more resistant than control mice to a lethal hematogenous infection by C. albicans, as evidenced by extension of survival in an M1g1 dose-dependent manner (P, 0.08 to <0.001) and by reduction in number of infection foci and their size in the kidney. In vitro studies found that M1g1 promoted phagocytosis and phagocytic killing of C. albicans yeast cells by mouse peritoneal macrophages and was required for activation of the mouse complement cascade. Thus, human antimannan antibody may have a protective role in host resistance to systemic candidiasis.     

Diagnosis of Helicobacter pylori infection by using pyloriset EIA-G and EIA-A for detection of serum immunoglobulin G (IgG) and IgA antibodies.

We evaluated the performance of new enzyme immunoassay (EIA) kits (Pyloriset; Orion Corporation, Orion Diagnostica, Espoo, Finland) for the detection of immunoglobulin G (IgG) and IgA antibodies to Helicobacter pylori in serum. Serum samples from 195 patients with upper abdominal complaints were collected. Biopsy specimens of the gastric mucosae were taken for histological analysis and bacterial culture. The sensitivity, specificity, and positive and negative predictive values, and efficacy of the Pyloriset EIA-G in detecting IgG antibodies to H. pylori were 92, 84, 88, 90, and 89%, respectively, when compared with those of the reference methods used. The corresponding data for detection of IgA antibodies were 80, 89, 89, 79, and 84%, respectively. The overall prevalence of defined H. pylori positivity was 54%. Moreover, the antibody tests showed a very good correlation with the biopsy findings. IgG antibodies were found in 93% of sera from patients with documented gastritis and H. pylori positivity, whereas only 4% of the sera from patients with documented gastritis and H. pylori-negative patients was positive. The results obtained for IgA antibodies were 81 and 6%, respectively. We conclude that the Pyloriset EIA-G, the test for IgG antibodies, is a good and reliable test for the detection of antibodies to H. pylori and as an indication of H. pylori infection. The determination of IgA antibodies may be used as a test that complements the IgG antibody assay.     

Analysis of false negative results in the immunoassay for anti-acetylcholine receptor antibodies in myasthenia gravis

Possible causes for the failure of immunoassays to detect anti-acetylcholine receptor activity in serum from confirmed myasthenia gravis (MG) patients were investigated. A more sensitive assay, using Protein A to trap immune complexes (ARIA), was applied to 65 MG sera which were negative in the usual assay and to 42 normal human sera. Normal and negative MG sera had antibody (Ab) activity in the same range (50-70 pM). Titers present in 70% of normal sera appeared to be specific antireceptor antibodies as defined by tests for antigen specificity. Thus, higher sensitivity assays did not improve discrimination of MG from normals. In a second group of 108 MG sera studied, 48 were negative by the usual assay criteria in a rat acetylcholine receptor immunoassay. Further detailed analysis of this negative group showed that 3/48 had IgG3 antibody not detectable in the test, 14/48 had Ab's recognizing human receptor determinants exclusively, 29/48 had toxin blocking Ab's not determined by immunoassays, and 6/48 were negative in all tests. The results indicate that the exclusive occurrence of toxin-blocking antibodies in MG subjects is a major factor contributing to false negatives in the ARIA test. Estimates of the amount of Ab's with this functionality indicated that they are present in very much smaller amounts than other classes of anti-receptor Ab's. Degree of blocking activity in patient serum showed a fair correlation with severity of disease. Thus, blocking antibodies appear capable of causing all degrees of disease severity in the absence of other types of antireceptor Ab's. The development of a sensitive and quantitative in vitro assay for blocking antibodies combined with the usual immunoassay would be a major improvement for a MG diagnostic test, with greater than 94% positivity predicted.     

Competitive radioimmunoassay to detect antibodies to herpes B virus and SA8 virus.

A monoclonal competitive radioimmunoassay (CompRIAm) which detects antibody to herpesvirus simiae (B virus) in monkey and human sera and antibody to SA8 virus in monkey sera but not antibody to herpes simplex virus in human sera is described. Of 232 serum samples from wild-caught cynomolgus monkeys, 117 serum samples were positive when tested by CompRIAm. The results were in close agreement (97.5%) with B virus neutralizing antibody results on the same sera. Sera from 97 wild-caught rhesus monkeys and 92 wild-caught baboons were also tested. The CompRIAm was able to differentiate between sera that had neutralizing antibody to B virus and SA8 virus and those that did not, although the discrimination was not as clear as that in the tests on cynomolgus monkey sera. Sequential sera from two humans with confirmed cases of B virus infection were tested by CompRIAm. B virus antibody was detected in sera from both humans. None of 237 other serum samples from blood donors and patients attending sexually transmitted disease clinics reacted in the CompRIAm.     

Analysis of eight commercial enzyme immunoassay tests for detection of antibodies to Mycoplasma pneumoniae in human serum

Mycoplasma pneumoniae is an important etiologic agent of primary atypical pneumonia in children and adults. The diagnosis of M. pneumoniae infection is commonly confirmed through serologic testing. In this study, we used paired sera from 51 patients (all with confirmed M. pneumoniae infection and positive complement fixation [CF] titers) to compare the results of eight enzyme immunoassays (EIAs) available commercially in the United States. We compared two single-use EIAs and six plate-type EIAs. Results from acute-phase sera ranged from only 7 (14%) positive by ImmunoWELL (GenBio) immunoglobulin M (IgM) EIA to 23 (45%) positive by Zeus IgG EIA. When both the acute-phase and convalescent-phase serum samples were examined, positive results ranged from 20 (39%) by the ImmunoWELL (GenBio) IgM assay to 45 (88%) positive by the Remel IgG-IgM EIA. In this study, the single-use EIAs by Remel and Meridian were more reliable than were the plate-type EIAs. Among the plate-type EIAs, the Zeus and DiaSorin assays (which detect antibodies to protein antigens) were more sensitive than the ImmunoWELL assay (which detects antibodies to glycolipid antigens). In general, IgG EIAs on convalescent-phase sera were more concordant with one another than were IgM EIAs with one another. Scatter plot analysis of convalescent-phase sera showed that, as the CF titer dropped, the IgM assays identified fewer positive convalescent-phase sera. In contrast, the IgG assays provided fairly consistent positive results for convalescent-phase sera with CF titers of 64 and above. Results of individual tests and overall limitations of serodiagnostics for M. pneumoniae infections are discussed.