Antigen-binding characteristics of AbCD71 and its inhibitory effect on PHA-induced lymphoproliferation.
OBJECTIVE
To investigate the anti-lymphoproliferative effect of the prepared recombinant chimeric human/murine anti-cluster of differentiation(CD) 71 monoclonal antibody (AbCD71), which is composed of mouse-derived, antigen-binding variable regions and human-derived constant regions.


METHODS
After plasmids construction and transfection, the expression of AbCD71 in the transfectoma supernatant was determined by the sandwich ELISA. Indirect immunofluorescence assay was used to measure the antigen-binding characteristic and the percent CD71-expressed peripheral blood mononuclear cells (PBMC). The antibodies were purified from the ascites via diethylaminoethyl(DEAE)-Sephadex A-50 chromatography and then identified by SDS-PAGE. At last, inhibitory effect of AbCD71 on PHA-induced PBMC proliferation was calculated by methyl thiazolyl tetrazolium(MTT) assay.


RESULTS
Constant domain of heavy chain (C(H)) and light chain (C(L)) of AbCD71 were in the human Cgamma family and human Ckappa family, respectively. AbCD71 could compete with its original murine mAb to bind with CD71-positive human leukemia cell line CEM cells. AbCD71 could inhibit the peripheral blood mononuclear cell proliferation induced by phytohemagglutinin(PHA) in vitro in a dose-dependent manner, especially at time-points 0 and 12 h after induction. There was no statistical difference when compared with original murine mAb.


CONCLUSIONS
The AbCD71 is a promising immunosuppressant. Our approach to blocking CD71 with the chimeric human/murine mAb provides a novel strategy for prolonging allograft survival.
Lei
Ping
P
Department of Immunology and Pharmacology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
He
Yong
Y
Ye
Qing
Q
Zhu
Hui-Fen
HF
Yuan
Xiao-Mei
XM
Liu
Jing
J
Xing
Wei
W
Wu
Sha
S
Dai
Wei
W
Shen
Xin
X
Wang
Guo-Bin
GB
Shen
Guan-Xin
GX
D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology / Q000494:pharmacology*; D015703:Antigens, CD / Q000235:genetics / Q000276:immunology*; D049109:Cell Proliferation / Q000187:drug effects*; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry; D007147:Immunoglobulin Light Chains / Q000737:chemistry; D007166:Immunosuppressive Agents / Q000494:pharmacology; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000187:drug effects*; D051379:Mice; D008807:Mice, Inbred BALB C; D010835:Phytohemagglutinins / Q000494:pharmacology; D010957:Plasmids; D011485:Protein Binding / Q000276:immunology; D011990:Receptors, Transferrin / Q000235:genetics / Q000276:immunology*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D014162:Transfection
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; C068126:CD71 antigen; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D007166:Immunosuppressive Agents; D010835:Phytohemagglutinins; D011990:Receptors, Transferrin; D011993:Recombinant Fusion Proteins