Characterization of human tissue factor (TF)-specific monoclonal antibodies prepared using a rapid immunization protocol.
Tissue factor (TF) plays an important, physiological role in hemostasis. Recent studies have demonstrated the over-expression of TF in a number of solid tumor types and its pathological roles in angiogenesis and tumor metastasis. In this study, we report the development and characterization of a panel of murine MAbs that are specific for human TF, but do not inhibit TF-mediated blood coagulation. By using a modified repetitive immunizations at multiple sites (RIMMS) protocol in conjunction with an efficient hybridoma cloning procedure, anti-TF MAbs were generated within a relatively short time frame of 5-6 weeks. Following primary screening by ELISA, the binding of the MAbs to the native form of human TF was demonstrated in flow cytometry using a stable cell line expressing human TF. Several of these TF-specific MAbs did not inhibit blood coagulation in a blood coagulation assay and bound with high affinity (0.5-2 nM) to human TF in BIAcore analyses. Importantly, this study represents an independent evaluation of the RIMMS strategy for MAb generation and demonstrates that class-switched, high-affinity MAbs can be generated rapidly and reliably using RIMMS.
Chen
Chaoyuan
C
Department of Immunotherapeutics, Purdue Pharma L.P., Cranbury, New Jersey, USA.
Yang
Qifeng
Q
Patel
Sima
S
Lei
Ying
Y
McAleer
Lihong
L
Singleton
Judith
J
Soltis
Daniel
D
Wang
Baiyang
B
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D006825:Hybridomas; D007074:Immunoglobulin G; D051379:Mice; D008807:Mice, Inbred BALB C; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D013925:Thromboplastin / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D011994:Recombinant Proteins; D013925:Thromboplastin