High-throughput antibody generation using multiplexed immunization and immunogen array analysis.
In this work, the authors developed a new screening approach using multiplexed immunization and immunogen array analysis to improve the efficiency of antibody screening for high-throughput antibody generation. The immunogen array is based on a 96-well format in which different immunogens and negative as well as positive controls are immobilized in each well, thus making it possible to screen hundreds of antibody candidates simultaneously. To demonstrate this approach, a model of 4 mixed immunogens immunization was employed. In total, 675 antibody candidates were screened before and after established antibody hybridomas in parallel with immunogen arrays and enzyme-linked immunosorbent assay. The signal intensity, specificity, and cross-reactivity of produced antibody candidates were analyzed using a hierarchical cluster algorithm to track the characteristics of antibody candidates during antibody generation, which might reduce the number of false-positive and false-negative binding of antibodies. Moreover, 4 monoclonal antibodies that were produced successfully recognized their corresponding target antigens.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing, China.
Zhe
Z
Yu
Li
L
Liu
Xiaobo
X
Yu
Jun
J
Chi
Huanhuan
H
Han
Ying
Y
Liu
Wei
W
He
Qihong
Q
Sun
Jianen
J
Gao
Danke
D
Xu
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D016000:Cluster Analysis; D019084:Fluorescent Antibody Technique, Indirect; D056945:Hep G2 Cells; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D007114:Immunization / Q000379:methods*; D007118:Immunoassay; D051379:Mice; D008807:Mice, Inbred BALB C; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000276:immunology; D054464:Peroxiredoxins / Q000276:immunology; D040081:Protein Array Analysis; D012680:Sensitivity and Specificity
D000911:Antibodies, Monoclonal; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D054464:Peroxiredoxins