Preparation and identification of monoclonal antibodies against recombinant human thioredoxins.
Thioredoxin (Trx) is a member of the thioredoxin protein family and has a conserved catalytic domain (-Trp-Cys-Gly-Pro-Cys-Lys-) with reduction/oxidation (redox) activity. There are two main members in this family, Trx-1, a cytosolic and nuclear form, and Trx -2, a mitochondrial form. Trx-1 is a 104 amino acid multifunctional protein that has been extensively studied. Here we report the preparation of monoclonal antibodies (MAb) against recombinant human Trx-1(hTrx). To enhance its immunogenicity, Trx-1 was coupled to carrier protein bovine serum albumin by a two-step glutaraldehyde method. Using conventional procedures, we prepared three stable hybridoma cell lines that can produce and secret anti-Trx MAbs. We further analyzed their isotypes, titer, and affinity and found that those MAbs belong to the G1 subclass with kappa light chains, respectively. The MAbs were capable of recognizing hTrx-1, as determined by Western blotting.
Tang
Wen-Xin
WX
College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, China.
Chen
Xiao-Ping
XP
Zhao
Yan-Ying
YY
Wang
Jian-He
JH
Wang
Wei
W
Chen
Zheng-Wang
ZW
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D002417:Cattle; D002459:Cell Fusion; D002460:Cell Line; D045744:Cell Line, Tumor; D006801:Humans; D006825:Hybridomas; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000276:immunology*; D013879:Thioredoxins / Q000276:immunology*
D000911:Antibodies, Monoclonal; D011994:Recombinant Proteins; C491203:TXN protein, human; D013879:Thioredoxins