To further refine our current nanoparticle-based HIV-1 p24
antigen assay, we investigated immune responses to p24 to identify diagnostically significant immune dominant
epitopes (IDEs) in HIV-infected human sera, to address cross-reactivity of anti-p24
antibodies to different subtypes, and to identify new
biomarkers that distinguish acute from chronic
HIV infection for more accurate incidence estimation. We identified two major linear
epitope regions, located in the CypA binding loop and adjacent helices and at the end of the C-terminal domain. Most sera (86%) from acutely HIV-1-infected individuals reacted with multiple
peptides, while 60% and 30% of
AIDS patient samples reacted with multiple and single
peptides, respectively. In contrast, 46% and 43% of chronically HIV-1-infected individuals reacted with one and none of the
peptides, respectively, and only 11% reacted with multiple p24
peptides, indicating a progression of immune responses from polyclone-like during acute
infection to monoclone-like or a nonresponse to linear
epitopes during
chronic infection. Anti-p24
antibodies (subtype B) show broad cross-reactivity to different HIV-1 subtypes, and the synergistic action of different combinations of anti-
HIV antibodies improves capture and detection of divergent HIV-1 subtypes. Our results indicate that the modified
peptide immunoassay is sensitive and specific for the rapid identification of HIV-1 p24 IDEs and for investigation of immune responses to p24 during natural HIV-1
infection. The data provide the foundation for development and refinement of new assays for improved p24
antigen testing as future tools for rapid and accurate diagnosis as part of early intervention strategies and estimations of incidence.