Analyzing antibody specificity with whole proteome microarrays.
Although approximately 10,000 antibodies are available from commercial sources, antibody reagents are still unavailable for most proteins. Furthermore, new applications such as antibody arrays and monoclonal antibody therapeutics have increased the demand for more specific antibodies to reduce cross-reactivity and side effects. An array containing every protein for the relevant organism represents the ideal format for an assay to test antibody specificity, because it allows the simultaneous screening of thousands of proteins for possible cross-reactivity. As an initial test of this approach, we screened 11 polyclonal and monoclonal antibodies to approximately 5,000 different yeast proteins deposited on a glass slide and found that, in addition to recognizing their cognate proteins, the antibodies cross-reacted with other yeast proteins to varying degrees. Some of the interactions of the antibodies with noncognate proteins could be deduced by alignment of the primary amino acid sequences of the antigens and cross-reactive proteins; however, these interactions could not be predicted a priori. Our findings show that proteome array technology has potential to improve antibody design and selection for applications in both medicine and research.
Michaud
Gregory A
GA
Protometrix, 688 East Main Street, Branford, Connecticut 06405, USA.
Salcius
Michael
M
Zhou
Fang
F
Bangham
Rhonda
R
Bonin
Jaclyn
J
Guo
Hong
H
Snyder
Michael
M
Predki
Paul F
PF
Schweitzer
Barry I
BI
D000906:Antibodies / Q000032:analysis* / Q000737:chemistry / Q000276:immunology*; D000918:Antibody Specificity / Q000276:immunology*; D000936:Antigen-Antibody Complex / Q000032:analysis* / Q000737:chemistry / Q000276:immunology*; D005656:Fungal Proteins / Q000276:immunology; D007118:Immunoassay / Q000295:instrumentation / Q000379:methods; D015335:Molecular Probes / Q000737:chemistry / Q000276:immunology; D040081:Protein Array Analysis / Q000295:instrumentation / Q000379:methods*; D025941:Protein Interaction Mapping / Q000379:methods*; D020543:Proteome / Q000737:chemistry / Q000276:immunology*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D000906:Antibodies; D000936:Antigen-Antibody Complex; D005656:Fungal Proteins; D015335:Molecular Probes; D020543:Proteome