Multiplexed serologic assay for nine anogenital human papillomavirus types.
A multiplexed human papillomavirus (HPV) immunoassay has been developed for the detection of human IgG antibodies to HPV type 6, 11, 16, 18, 31, 33, 45, 52, and 58 virus-like particle (VLP) types in serum following natural infection or immunization with VLP-based vaccines. The VLP antigens were covalently conjugated to carboxyl Luminex microspheres (MS) using a carbodiimide chemistry. Antibody (Ab) titers were determined in a direct binding format, in which an IgG1- to -4-specific, phycoerythrin (PE)-labeled monoclonal antibody (MAb) (HP6043) binds to human serum IgG antibodies. Pooled serum samples from rhesus macaques immunized with a 9-valent VLP-based vaccine served as the reference standard. The overall specificity of the assay was >99%, and the linearity (parallelism) of the assay was <7% per 10-fold dilution. Total assay precision was <19% across 3 different VLP-microsphere lots, 2 secondary antibody lots, and 2 different operators over a period of 3 weeks. Three different methods were used to evaluate serostatus cutoffs (SCO): (i) a clinical sensitivity/specificity analysis based on "likely negative" and "likely positive" samples from nonvaccinees, (ii) stringent upper tolerance limits on samples from "likely negatives," and (iii) stringent upper tolerance limits from the same "likely negative" sample set after VLP adsorption. Depending on the method to set the serostatus cutoff, the percentage of seropositive samples at the month 48 time point following vaccination with the HPV 6/11/16/18 quadrivalent vaccine ranged from 70% to 100%. This assay has proven useful for measuring the levels of serum antibody to the nine HPV VLPs following natural infection or administration of VLP-based vaccines.
Opalka
David
D
Wayne Clinical Support, Merck Research Laboratories, Wayne, PA 19087-8630, USA.
Matys
Katie
K
Bojczuk
Paul
P
Green
Tina
T
Gesser
Richard
R
Saah
Alfred
A
Haupt
Richard
R
Dutko
Frank
F
Esser
Mark T
MT
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral; D005260:Female; D006801:Humans; D007118:Immunoassay / Q000379:methods; D007074:Immunoglobulin G / Q000097:blood; D008253:Macaca mulatta; D008297:Male; D027383:Papillomaviridae / Q000145:classification / Q000276:immunology*; D030361:Papillomavirus Infections / Q000175:diagnosis* / Q000276:immunology*; D053918:Papillomavirus Vaccines / Q000276:immunology*; D012680:Sensitivity and Specificity; D014773:Virology / Q000379:methods*; D022701:Virosomes
D000914:Antibodies, Viral; D000956:Antigens, Viral; D007074:Immunoglobulin G; D053918:Papillomavirus Vaccines; D022701:Virosomes