Fluoroimmunoassay studies with solubilized antigens from Borrelia burgdorferi

Hechemy, Karim E.; Harris, H L; Wethers, J.; Stevens, Roy W.; Stock, B R; Reilly, Andrew; Benach, Jorge L.

doi:10.1128/jcm.27.8.1854-1858.1989

Cited by 8 publications

(6 citation statements)

References 17 publications

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“…three species were grown for 10 to 13 days. The Borrelia cultures were then processed as described previously (12). T. phagedenis was grown in multiple cultures of 100 ml of pepticase-yeast-glucose semisolid-Smibert's salt solutionserum-cocarboxylase medium (as described by R. M. Smibert [26a]).…”

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confidence: 99%

“…SDS gel electrophoresis and electroblotting. Sodium dodecyl sulfate (SDS) gel electrophoresis and electroblotting of whole spirochetal antigens were performed as described previously (12). SDS gel electrophoresis of recombinant-P39 antigen was performed with 4% stacking and 12.5% separating gels by use of the Mini-Protean II system (Bio-Rad…”

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confidence: 99%

“…Immunoblotting. Immunoblotting was performed as described previously (12). The color was developed with 4-chloro-1-naphthol when the HRP-labeled probe was used or with fast red when the alkaline phosphatase-labeled probe was used.…”

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Monoclonal antibody to native P39 protein from Borrelia burgdorferi

et al. 1994

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We have produced, by using a sonicate of Borrelia burgdorferi, a monoclonal antibody (MAb), NYSP39H, that is specific for the P39 protein band. This MAb reacted with 13 isolates of B. burgdorfieri but not with eight different spirochetes (four borrelias, two leptospiras, and two treponemas). Surface labeling of B. burgdorfieri with biotin and subsequent treatment with Nonidet P-40 showed that P39 was not biotinylated but was extracted with Nonidet P-40, indicating that it is present within the outer membrane, but not on the surface of the spirochete. Immunoelectron microscopy revealed the immunogold probe primarily at the cytoplasmic membrane region of the spirochete. The MAb detected B. burgdorferi in the indirect fluorescent-antibody test only when the spirochetes from a culture or in a tick homogenate were fixed with polylysine and not with acetone. NYSP39H appears to be an appropriate probe for use in the specific detection of B. burgdoferi.

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Monoclonal antibody to native P39 protein from Borrelia burgdorferi

et al. 1994

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“…burgdorferi antibody screening and immunoblots. Lyme disease screening tests for B. burgdorferi antibodies were performed by fluorescent immunoassay with a whole-cell sonicate of the B31 strain B. burgdorferi (ATCC 2521) and the VIDAS Lyme screen (17). The cutoff for a positive VIDAS Lyme screen was a test value of 1.00, while samples with test values of between 0.75 and 1.00 were considered equivocal.…”

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confidence: 99%

Serological responses to Ehrlichia equi, Ehrlichia chaffeensis, and Borrelia burgdorferi in patients from New York State

1997

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Serological testing at the New York State Department of Health for human granulocytic ehrlichiosis in the residents of Westchester County, N.Y., was performed with specimens from 176 patients by the indirect fluorescent-antibody (IFA) technique with Ehrlichia equi MRK-infected neutrophils. To understand whether human monocytotropic ehrlichiosis also occurs in this northeastern geographic region, specimens were also tested for antibodies to Ehrlichia chaffeensis Arkansas. Screening tests and immunoblots for Lyme disease (Borrelia burgdorferi infection) were also performed. Thirty-two patients had antibodies only to E. equi and 21 patients had antibodies to both E. equi and E. chaffeensis, whereas 12 patients had only E. chaffeensis antibodies by the IFA technique. The remaining patients did not have antibodies to either ehrlichia. Eighteen serum samples from 13 of these patients were coded and sent to the Ehrlichia Research Laboratory (Baltimore, Md.) for repeat analysis by the IFA test and for E. equi and E. chaffeensis immunoblots. Immunoblot analysis for E. equi in samples with positive IFA test results confirmed the results for eight of the nine specimens. Immunoblot analyses for E. chaffeensis were negative for all 18 serum samples. Borrelia-reactive antibodies were found in sera both from patients with granulocytic ehrlichiosis and from patients with monocytotropic ehrlichiosis from New York State. Our results suggest that E. equi antigen is an appropriate substrate for identifying human granulocytic ehrlichiosis. E. chaffeensis antigen lacks appropriate sensitivity to serve as a surrogate substrate for the detection of human granulocytic ehrlichiosis and should be used solely for the diagnosis of human monocytotropic ehrlichiosis. Heat shock proteins may, in some cases, cause cross-reactivity between B. burgdorferi and ehrlichiae.

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“…Determination of antibody levels to B. burgdorferi is the most frequently used laboratory method for the diagnosis of Lyme disease. Established procedures include the indirect fluorescent-antibody test (18,25), formats of the enzymelinked immunosorbent assay (7,18) including the antibody capture assay (2), and the quantitative fluorescence immunoassay (11,17). These procedures, however, have demonstrated a lack of sensitivity, especially in the early stages of disease, and cross-reactivity with other spirochetal or autoimmune diseases (7,11,17,18,25).…”

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Detection of borreliacidal antibodies in hamsters by using flow cytometry

et al. 1992

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Flow cytometry can be used to detect antibody that kills Borrelia burgdorferi. Borreliacidal activity was detected within 3 h of incubating B. burgdorferi with immune serum and complement. Right-angle light scatter and propidium iodide fluorescence were the cytometric parameters which correlated best with in vitro killing of B. burgdorferi. Flow cytometry is a rapid method for determining the presence of borreliacidal activity and may lead to a better serodiagnostic test for the detection of Lyme disease.

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Fluoroimmunoassay studies with solubilized antigens from Borrelia burgdorferi

Cited by 8 publications

References 17 publications

Monoclonal antibody to native P39 protein from Borrelia burgdorferi

Monoclonal antibody to native P39 protein from Borrelia burgdorferi

Serological responses to Ehrlichia equi, Ehrlichia chaffeensis, and Borrelia burgdorferi in patients from New York State

Detection of borreliacidal antibodies in hamsters by using flow cytometry

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