A High Throughput Protein Microarray Approach to Classify HIV Monoclonal Antibodies and Variant Antigens.
In recent years, high throughput discovery of human recombinant monoclonal antibodies (mAbs) has been applied to greatly advance our understanding of the specificity, and functional activity of antibodies against HIV. Thousands of antibodies have been generated and screened in functional neutralization assays, and antibodies associated with cross-strain neutralization and passive protection in primates, have been identified. To facilitate this type of discovery, a high throughput-screening tool is needed to accurately classify mAbs, and their antigen targets. In this study, we analyzed and evaluated a prototype microarray chip comprised of the HIV-1 recombinant proteins gp140, gp120, gp41, and several membrane proximal external region peptides. The protein microarray analysis of 11 HIV-1 envelope-specific mAbs revealed diverse binding affinities and specificities across clades. Half maximal effective concentrations, generated by our chip analysis, correlated significantly (P<0.0001) with concentrations from ELISA binding measurements. Polyclonal immune responses in plasma samples from HIV-1 infected subjects exhibited different binding patterns, and reactivity against printed proteins. Examining the totality of the specificity of the humoral response in this way reveals the exquisite diversity, and specificity of the humoral response to HIV.
Division of Infectious Diseases, University of California Irvine, Irvine, California, United States of America.
Emmanuel Y
EY
Dotsey
Division of Infectious Diseases, University of California Irvine, Irvine, California, United States of America.
Andrea
A
Gorlani
Department of Cell and Molecular Biology, The Scripps Research Institute, La Jolla, California, United States of America.
Sampat
S
Ingale
Division of Infectious Diseases, Northwestern University, Chicago, Illinois, United States of America.
Chad J
CJ
Achenbach
Division of Infectious Diseases, University of California Irvine, Irvine, California, United States of America.
Donald N
DN
Forthal
Division of Infectious Diseases, University of California Irvine, Irvine, California, United States of America.
Philip L
PL
Felgner
Division of Infectious Diseases, University of California Irvine, Irvine, California, United States of America.
Johannes S
JS
Gach
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity / Q000276:immunology; D016000:Cluster Analysis; D004797:Enzyme-Linked Immunosorbent Assay; D015483:HIV Antibodies / Q000276:immunology*; D015488:HIV Antigens / Q000276:immunology*; D015658:HIV Infections / Q000097:blood / Q000276:immunology*; D015497:HIV-1 / Q000276:immunology; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D011134:Polysaccharides / Q000276:immunology; D040081:Protein Array Analysis / Q000379:methods*; D016415:Sequence Alignment
D000911:Antibodies, Monoclonal; D015483:HIV Antibodies; D015488:HIV Antigens; D010455:Peptides; D011134:Polysaccharides