title: Simple objective detection of human lyme disease infection using immuno-PCR and a single recombinant hybrid antigen.
abstract: A serology-based tiered approach has, to date, provided the most effective means of laboratory confirmation of clinically suspected cases of Lyme disease, but it lacks sensitivity in the early stages of disease and is often dependent on subjectively scored immunoblots. We recently demonstrated the use of immuno-PCR (iPCR) for detecting Borrelia burgdorferi antibodies in patient serum samples that were positive for Lyme disease. To better understand the performance of the Lyme disease iPCR assay, the repeatability and variability of the background of the assay across samples from a healthy population (n = 36) were analyzed. Both of these parameters were found to have coefficients of variation of <3%. Using eight antigen-specific iPCR assays and positive call thresholds established for each assay, iPCR IgM and/or IgG diagnosis from Lyme disease patient serum samples (n = 12) demonstrated a strong correlation with that of 2-tier testing. Furthermore, a simplified iPCR approach using a single hybrid antigen and detecting only IgG antibodies confirmed the 2-tier diagnosis in the Lyme disease patient serum samples (n = 12). Validation of the hybrid antigen IgG iPCR assay using a blinded panel of Lyme disease and non-Lyme disease patient serum samples (n = 92) resulted in a sensitivity of 69% (95% confidence interval [CI], 50% to 84%), compared to that of the 2-tier analysis at 59% (95% CI, 41% to 76%), and a specificity of 98% (95% CI, 91% to 100%) compared to that of the 2-tier analysis at 97% (95% CI, 88% to 100%). A single-tier hybrid antigen iPCR assay has the potential to be an improved method for detecting host-generated antibodies against B. burgdorferi.
authors.affiliation: Burnett School of Biomedical Sciences, University of Central Florida College of Medicine, Orlando, Florida, USA.
authors.forename: Micah D
authors.initials: MD
authors.lastname: Halpern
authors.affiliation: Diagnostic and Reference Laboratory, Bacterial Diseases Branch, Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, Colorado, USA.
authors.forename: Claudia R
authors.initials: CR
authors.lastname: Molins
authors.affiliation: Diagnostic and Reference Laboratory, Bacterial Diseases Branch, Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, Colorado, USA.
authors.forename: Martin
authors.initials: M
authors.lastname: Schriefer
authors.affiliation: Burnett School of Biomedical Sciences, University of Central Florida College of Medicine, Orlando, Florida, USA mollie.jewett@ucf.edu.
authors.forename: Mollie W
authors.initials: MW
authors.lastname: Jewett
mesh_terms: D000907:Antibodies, Bacterial / Q000097:blood*; D000942:Antigens, Bacterial / Q000235:genetics / Q000276:immunology; D001426:Bacterial Proteins / Q000235:genetics / Q000276:immunology; D025065:Borrelia burgdorferi / Q000276:immunology; D002460:Cell Line; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood / Q000276:immunology; D007075:Immunoglobulin M / Q000097:blood / Q000276:immunology; D008193:Lyme Disease / Q000097:blood / Q000175:diagnosis* / Q000276:immunology; D016133:Polymerase Chain Reaction / Q000379:methods*; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D012680:Sensitivity and Specificity
chemical_list: D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D011994:Recombinant Proteins