scholarly journals Peptide-Based OspC Enzyme-Linked Immunosorbent Assay for Serodiagnosis of Lyme Borreliosis

1998 ◽  
Vol 36 (12) ◽  
pp. 3474-3479 ◽  
Author(s):  
Marianne J. Mathiesen ◽  
Michael Christiansen ◽  
Klaus Hansen ◽  
Arne Holm ◽  
Eva Åsbrink ◽  
...  
Keyword(s):  
Lyme Borreliosis ◽  
Immunosorbent Assay ◽  
Erythema Migrans ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Amino Acid Residues ◽  
Serum Samples ◽  
Igg Antibody ◽  
Acrodermatitis Chronica Atrophicans ◽  
Increased Sensitivity

Sera from 210 patients with Lyme borreliosis (LB) were studied by an enzyme-linked immunosorbent assay (ELISA) based on a synthetic peptide (pepC10) comprising the C-terminal 10-amino-acid residues of OspC of Borrelia burgdorferi. We found that 36.3 and 45.0% of the serum samples from patients with erythema migrans (EM) and neuroborreliosis (NB), respectively, displayed immunoglobulin M (IgM) anti-pepC10 reactivities, while these samples rarely (≤8%) displayed IgG antibody reactivities. Sera from patients with acrodermatitis chronica atrophicans did not contain anti-pepC10 antibodies. The diagnostic performance of this newly developed peptide ELISA was compared with those of an ELISA based on the full-length recombinant OspC protein (rOspC) and a commercially available ELISA based on theB. burgdorferi flagellum (Fla). The sensitivity of the IgM pepC10 ELISA was slightly lower (P < 0.04) than that of the rOspC ELISA for EM patients (36.3 versus 43.8%), while there was no difference for NB patients (45.0 versus 48.0%). However, the optical density values obtained by the pepC10 ELISA were generally higher than those obtained by the rOspC ELISA, leading to a significantly better quantitative discrimination between seropositive patients with NB and controls (P < 0.008). The specificity of the pepC10 ELISA was similar to those of the rOspC ELISA and the Fla ELISA for relevant controls including patients with syphilis and mononucleosis. Although the overall diagnostic sensitivity of the Fla ELISA was superior, 8.8 and 12.0% of the EM and NB patients, respectively, were antibody positive only by the pepC10 ELISA. Thus, use of a diagnostic test for LB based on the detection of IgM antibodies to pepC10 and Fla has increased sensitivity for the diagnosis of early LB.

scholarly journals Enzyme-Linked Immunosorbent Assay Using Recombinant OspC and the Internal 14-kDa Flagellin Fragment for Serodiagnosis of Early Lyme Disease

1998 ◽  
Vol 36 (4) ◽  
pp. 857-861 ◽  
Author(s):  
Sebastian Rauer ◽  
Nicole Spohn ◽  
Christiane Rasiah ◽  
Uwe Neubert ◽  
Arnold Vogt
Keyword(s):  
Lyme Disease ◽  
Immunosorbent Assay ◽  
Erythema Migrans ◽  
Cell Extract ◽  
Sensu Stricto ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Epstein Barr Virus Infection ◽  
Serum Samples ◽  
Antigen Elisa

The outer surface protein C (OspC) and the internal 14-kDa flagellin fragment of strain GeHo of Borrelia burgdorferisensu stricto were expressed as recombinant proteins inEscherichia coli and were purified for use in an immunoglobulin M (IgM) enzyme-linked immunosorbent assay (OspC–14-kDa antigen ELISA). No hint at disturbing protein-protein interferences, which might influence the availability of immunoreactive epitopes, was found when the recombinant antigens were combined in the ELISA. The recombinant OspC–14-kDa antigen ELISA was compared to a commercial IgM ELISA that used a detergent cell extract from Borrelia afzelii PKo as the antigen. According to the manufacturer’s information, the cell extract contains, in addition to other antigens, the following diagnostically relevant antigens: the 100-kDa (synonyms, 93- and 83-kDa antigens), 41-kDa, OspA, OspC, and 17-kDa antigens. The specificity was adjusted to 95% on the basis of data for 154 healthy controls. On testing of 104 serum samples from patients with erythema migrans (EM), the sensitivity of the recombinant ELISA (46%) for IgM antibodies was similar to that of the commercial ELISA (45%). However, when 42 serum samples from patients with polyclonal B-cell stimulation due to an Epstein-Barr virus infection were tested, false-positive reactions were significantly less frequent in the recombinant ELISA (10%) than in the whole-cell-extract ELISA (23%). OspC displays sequence heterogeneity of up to 40% according to the genomospecies. However, when the reactions of serum specimens from controls and EM patients with OspC from representative strains of B. burgdorferi sensu stricto (strain GeHo) and B. afzelii (strain PKo) were compared in an ELISA, almost no differences in specificity and sensitivity were seen. This demonstrates that the sera predominantly recognize the common epitopes of OspC tested in this study. In conclusion, we suggest that the OspC–14-kDa antigens ELISA is a suitable test for the detection of an IgM response in early Lyme disease.

scholarly journals Development of an Enzyme-Linked Immunosorbent Assay for Immunoglobulin M Antibodies against Measles Virus

2003 ◽  
Vol 10 (3) ◽  
pp. 439-442 ◽  
Author(s):  
F. Roodbari ◽  
M. H. Roustai ◽  
A. Mostafaie ◽  
H. Soleimanjdahi ◽  
R. Sarrami Foroshani ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Neutralizing Antibodies ◽  
Measles Virus ◽  
Clinical Symptoms ◽  
Maculopapular Rash ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Vaccination Programs ◽  
Elisa System

ABSTRACT Measles is a highly contagious respiratory virus infection, with typical clinical symptoms including maculopapular rash, fever, cough, coryza, and conjunctivitis. Despite implementation of widespread vaccination programs throughout the world, the rates of global morbidity and mortality are still considerable. This study was performed to design a reliable indirect enzyme-linked immunosorbent assay (ELISA) to measure measles-specific immunoglobulin M (IgM). First, human IgM was purified, and then an anti-IgM antibody was produced in rabbits and purified in a multistep process. The rabbit IgG against human IgM was conjugated with peroxidase. Measles virus-infected Vero cells produced viral antigen. One hundred serum samples from infants of 9 to 18 months of age, mostly vaccinated, were evaluated for determining the presence of specific IgM antibodies against measles virus. The samples were also evaluated for neutralizing antibodies against measles virus by a microneutralization test (MNT). By comparing the results of the ELISA with those of MNT, it was demonstrated that ELISA had a sensitivity and specificity of 100 and 92%, respectively. On the other hand, when the results obtained by our ELISA system were compared with those of an imported measles virus IgM ELISA kit (EIAgen; Adaltis Italia SPa, Bologna, Italy), a high level of agreement was shown (k = 0.926).

scholarly journals Evaluation of an Immunoglobulin M Capture Enzyme-Linked Immunosorbent Assay for Diagnosis of Orientia tsutsugamushi Infection

2003 ◽  
Vol 10 (3) ◽  
pp. 394-398 ◽  
Author(s):  
Won-Jong Jang ◽  
Myung-Suk Huh ◽  
Kyung-Hee Park ◽  
Myung-Sik Choi ◽  
Ik-Sang Kim
Keyword(s):  
Immunosorbent Assay ◽  
Scrub Typhus ◽  
Microtiter Plate ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Orientia Tsutsugamushi ◽  
Serum Samples ◽  
Capture Elisa ◽  
Igm Antibodies ◽  
Immunodominant Antigen

ABSTRACT To differentiate scrub typhus from other acute febrile diseases, a rapid and reliable serological diagnosis is important. We developed an immunoglobulin M (IgM) capture enzyme-linked immunosorbent assay (ELISA) for diagnosis of recent Orientia tsutsugamushi infections in humans. The 56-kDa major outer membrane protein of O. tsutsugamushi is well known as the most immunodominant antigen in scrub typhus. The test is based on the use of the biotinylated recombinant 56-kDa protein of O. tsutsugamushi Boryong, Bor56, which was expressed as a fusion protein with a maltose-binding protein in Escherichia coli. In the test, the serum IgM antibodies were captured by anti-human IgM antibodies coated onto a microtiter plate. The captured IgM antibodies were revealed through sequential addition of biotinylated Bor56 antigen and peroxidase-conjugated streptavidin to the plate. The IgM capture ELISA was compared with the immunofluorescence antibody assay (IFA) by testing 176 serum samples from patients with diagnosed cases of rickettsial disease and patients with other acute febrile diseases. Of the 81 IgG IFA-positive samples, 78 tested positive (sensitivity, 96.3%) and all 31 IgM IFA-positive samples tested positive (sensitivity, 100%) by the IgM capture ELISA. The specificity of the IgM capture ELISA was 99%, and 1 of the 95 IFA-negative samples was positive in the assay. These results strongly suggest that IgM capture ELISA using the recombinant Bor56 antigen is a reliable and detailed method for the detection of early O. tsutsugamushi infection.

scholarly journals Development of an Immunoglobulin M (IgM) Capture Enzyme-Linked Immunosorbent Assay for Detection of Equine and Swine IgM Antibodies to Vesicular Stomatitis Virus

2001 ◽  
Vol 8 (3) ◽  
pp. 475-481 ◽  
Author(s):  
En-min Zhou ◽  
Jose Riva ◽  
Alfonso Clavijo
Keyword(s):  
Vesicular Stomatitis Virus ◽  
Immunosorbent Assay ◽  
Polyclonal Antibodies ◽  
Competitive Elisa ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Igm Antibodies ◽  
Vesicular Stomatitis ◽  
Elisa Plate

ABSTRACT An immunoglobulin M (IgM) capture enzyme-linked immunosorbent assay (MC-ELISA) was developed for the detection of primary infection of vesicular stomatitis virus (VSV) in equine and swine sera. The test was based on the use of biotinylated sheep antibodies against equine or swine IgM molecules bound to a streptavidin-coated ELISA plate. The captured IgM antibodies were detected by application of antigens prepared from the New Jersey and the Indiana VSV serotypes (VSV-NJ and VSV-IN, respectively) and mouse polyclonal antibodies against VSV-NJ and VSV-IN. The MC-ELISA was compared to a competitive ELISA (C-ELISA) and the standard microtiter serum neutralization (MTSN) assay by testing serum samples from horses and pigs experimentally infected with VSV-NJ or VSV-IN. The MC-ELISA detected specific homologous IgM antibodies from equine and swine sera as early as 5 and 4 days postinfection (DPI), respectively, and as late as 35 DPI. The MTSN test also detected antibodies as early as 5 DPI and as late as 160 DPI. In a similar fashion, the C-ELISA detected antibodies from 6 to 7 DPI and as late as 160 DPI. These results demonstrated that the MC-ELISA is a useful test for serodiagnosis of primary VSV infection in horses and pigs.

scholarly journals Comparison of Capture Immunoglobulin M (IgM) and IgG Enzyme-Linked Immunosorbent Assay (ELISA) and Nonstructural Protein NS1 Serotype-Specific IgG ELISA for Differentiation of Primary and Secondary Dengue Virus Infections

2003 ◽  
Vol 10 (4) ◽  
pp. 622-630 ◽  
Author(s):  
Pei-Yun Shu ◽  
Li-Kuang Chen ◽  
Shu-Fen Chang ◽  
Yi-Yun Yueh ◽  
Ling Chow ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Immunosorbent Assay ◽  
Primary Infection ◽  
Nonstructural Protein ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Virus Infections ◽  
Igg Antibody ◽  
Previous Infection ◽  
Specific Igg

ABSTRACT We have found that NS1 serotype-specific immunoglobulin G (IgG) enzyme-linked immunosorbent assay (ELISA) can be used to differentiate primary and secondary dengue virus infections. This is due to the fact that the NS1-specific IgG antibody cannot be detected before day 9 of illness for primary infection, so the NS1-specific IgG antibodies measured in acute-phase sera must come from previous infection. Comparison of NS1 serotype-specific IgG ELISA with envelope- and membrane-specific capture IgM and IgG ELISA in the differentiation of primary and secondary dengue virus infections showed good correlation (95.90% agreement). Most important, we have found that the serotype of the dengue virus from the majority of patients with primary infection could be correctly identified when convalescent-phase or postinfection sera were analyzed by NS1 serotype-specific IgG ELISA. These findings suggested that NS1 serotype-specific IgG ELISA could be reliably applied for serodiagnosis and seroepidemiological study of dengue virus infection.

scholarly journals Humoral Immune Response Associated with Lyme Borreliosis in Nonhuman Primates: Analysis by Immunoblotting and Enzyme-Linked Immunosorbent Assay with Sonicates or Recombinant Proteins

2002 ◽  
Vol 9 (6) ◽  
pp. 1348-1355 ◽  
Author(s):  
A. R. Pachner ◽  
D. Dail ◽  
L. Li ◽  
L. Gurey ◽  
S. Feng ◽  
...  
Keyword(s):  
Immune Response ◽  
Antibody Response ◽  
Lyme Borreliosis ◽  
Recombinant Proteins ◽  
Diagnostic Testing ◽  
Sensu Stricto ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Igg Antibody ◽  
Humoral Immune

ABSTRACT The immune response to Borrelia burgdorferi, the causative agent of Lyme disease, is complex. We studied the immunoglobulin M (IgM) and IgG antibody response to N40Br, a sensu stricto strain, in the rhesus macaque(nonhuman primate [NHP]) model of infection to identify the spirochetal protein targets of specific antibody. Antigens used in enzyme-linked immunosorbent assays were whole-cell sonicates of the spirochete and recombinant proteins of B. burgdorferi. Immunoblotting with a commercially available strip and subsequent quantitative densitometry of the bands were also used. Sera from four different groups of NHPs were used: immunocompetent, transiently immunosuppressed, extended immunosuppressed, and uninfected. In immunocompetent and transiently immunosuppressed NHPs, there was a strong IgM and IgG response. Major proteins for the early IgM response were P39 and P41 and recombinant BmpA and OspC. Major proteins for the later IgG response were P39, P41, P18, P60, P66, and recombinant BmpA and DbpA. There was no significant response in the NHPs to recombinant OspA or to Arp, a 37-kDa protein that elicits an antibody response during infection in mice. Most antibody responses, except for that to DbpA, were markedly diminished by prolonged dexamethasone treatment. This study supports the hypothesis that recombinant proteins may provide a useful adjunct to current diagnostic testing for Lyme borreliosis.

scholarly journals Sensitive and Specific Enzyme-Linked Immunosorbent Assay for Detecting Serum Antibodies against Mycobacterium avium subsp. paratuberculosis in Fallow Deer

2014 ◽  
Vol 21 (8) ◽  
pp. 1077-1085 ◽  
Author(s):  
José M. Prieto ◽  
Ana Balseiro ◽  
Rosa Casais ◽  
Naiara Abendaño ◽  
Liam E. Fitzgerald ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Mycobacterium Avium ◽  
Cost Effective ◽  
Fallow Deer ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Igg Antibody ◽  
Tissue Samples ◽  
Content Type ◽  
Mycobacterium Avium Subsp Paratuberculosis

ABSTRACTThe enzyme-linked immunosorbent assay (ELISA) is the diagnostic test most commonly used in efforts to control paratuberculosis in domestic ruminants. However, commercial ELISAs have not been validated for detecting antibodies againstMycobacterium aviumsubsp.paratuberculosisin wild animals. In this study, we compared the sensitivities and specificities of five ELISAs using individual serum samples collected from 41 fallow deer with or without histopathological lesions consistent with paratuberculosis. Two target antigenic preparations were selected, an ethanol-treated protoplasmic preparation obtained from a fallow deerM. aviumsubsp.paratuberculosisisolate (ELISAs A and B) and a paratuberculosis protoplasmic antigen (PPA3) (ELISAs C and D). Fallow deer antibodies bound to the immobilized antigens were detected by using a horseradish peroxidase (HRP)-conjugated anti-fallow deer IgG antibody (ELISAs A and C) or HRP-conjugated protein G (ELISAs B and D). A commercially available assay, ELISA-E, which was designed to detectM. aviumsubsp.paratuberculosisantibodies in cattle, sheep, and goats, was also tested. Although ELISAs A, C, and E had the same sensitivity (72%), ELISAs A and C were more specific (100%) for detecting fallow deer with lesions consistent with paratuberculosis at necropsy than was the ELISA-E (87.5%). In addition, the ELISA-A was particularly sensitive for detecting fallow deer in the latent stages of infection (62.5%). The antibody responses detected with the ELISA-A correlated with both the severity of enteric lesions and the presence of acid-fast bacteria in gut tissue samples. In summary, our study shows that the ELISA-A can be a cost-effective diagnostic tool for preventing the spread of paratuberculosis among fallow deer populations.

Sensitive microplate enzyme-linked immunosorbent assay for detection of antibodies against the scrub typhus rickettsia, Rickettsia tsutsugamushi

1979 ◽  
Vol 9 (1) ◽  
pp. 38-48 ◽  
Author(s):  
G A Dasch ◽  
S Halle ◽  
A L Bourgeois
Keyword(s):  
Optical Density ◽  
Immunosorbent Assay ◽  
Scrub Typhus ◽  
Fluorescent Antibody ◽  
Antibody Test ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Igg Antibody ◽  
Serum Dilution ◽  
End Point

A microtiter enzyme-linked immunosorbent assay (ELISA) has been developed for the titration of antibodies against scrub typhus in human and animal sera. Scrub typhus rickettsiae were grown in monolayers of irradiated mouse LM3 cells and separated from host cell materials by differential centrifugation, filtration through a glass filter (AP-20, Millipore Corp.), and isopycnic banding in Renografin density gradients. The scrub typhus ELISA antigens were obtained from the purified viable rickettsiae by French pressure cell disruption and addition of 0.2% Formalin to the soluble extract. Antisera prepared in rabbits against the prototype Karp, the Kato, and the Gilliam strains of scrub typhus were used to standardize the ELISA and to compare its sensitivity and specificity to that of the indirect fluorescent antibody test (IFA). ELISA titers were measured as the greatest serum dilution showing an optical density 0.25 above controls or by the optical density achieved at a fixed serum dilution. The IFA and ELISA end point titers were quite similar, and all three measures of titer had comparable specificity for the strains of scrub typhus. No cross-reactions between the typhus and scrub typhus wera were observed by ELISA. Both the immunoglobulin M (IgM) and IgG antibody titers of 12 sequential sera from four patients with scrub typhus were obtained by IFA and ELISA. The IFA and ELISA end point titers for IgM and IgG had correlation coefficients of 0.91 and 0.97, respectively, whereas the ELISA optical density values at a serum dilution of 1:100 had slightly lower correlations with IFA titers (0.80 and 0.94). Early rising IgM titers followed by rising IgG titers were demonstrated by ELISA in three patients with primary scrub typhus infections, whereas the IgG response predominated in a patient with a reinfection. It is concluded that the ELISA for scrub typhus is a very satisfactory alternative to the IFA test.

scholarly journals Comparison of a Commercial Enzyme-Linked Immunosorbent Assay with Immunofluorescence and Complement Fixation Tests for Detection of Coxiella burnetii (Q Fever) Immunoglobulin M

2000 ◽  
Vol 38 (4) ◽  
pp. 1645-1647 ◽  
Author(s):  
Peter R. Field ◽  
Jody L. Mitchell ◽  
Avelina Santiago ◽  
David J. Dickeson ◽  
Sau-Wan Chan ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Coxiella Burnetii ◽  
Complement Fixation ◽  
Q Fever ◽  
Fluorescent Antibody ◽  
Reference Method ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Igm Elisa

A commercially available enzyme-linked immunosorbent assay (ELISA) for the diagnosis of Q fever (PanBio Coxiella burnetiiimmunoglobulin M [IgM] ELISA, QFM-200) was compared to the indirect fluorescent antibody test (IFAT) for C. burnetii IgM and the complement fixation test (CFT). The ELISA demonstrated 92% agreement with the reference method (IFAT), and gave a sensitivity of 99% (69 of 70 samples) and a specificity of 88% (106 of 121). Specificity can be increased with confirmation by IFAT. CFT was found to have a specificity of 90% (107 of 119), although it was lacking in sensitivity (73%; 51 of 70). No cross-reactivity was observed in the ELISA with serum samples from patients with mycoplasma (n = 6), chlamydia (n = 5), or legionella (n = 4) infections, although 2 of 5 patients with leptospirosis and 1 of 4 samples containing rheumatoid factor (RF) demonstrated positive results in the ELISA. Results indicate that the performance of the PanBio C. burnetii (Q fever) IgM ELISA (F = 187) is superior to that of CFT (F = 163), and consequently the ELISA should be a useful aid in the diagnosis of acute Q fever.

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