Acta Academiae Medicinae Sinica

Acta Academiae Medicinae Sinica

Acta Academiae Medicinae Sinica ›› 2008, Vol. 30 ›› Issue (3): 354-359.

• Technological Method • Previous Articles     Next Articles

Preparation and Activity Detection of Monoclonal Antibody against Anti-CD3 ScFv

SHAO Xiao-feng, GAO Ying-dai, LIU Juan-ni, WANG Jin-hong, XU Yuan-fu, FAN Dong-mei, YANG Chun-zheng, XIONG Dong-sheng   

  1. State Key Laboratory of Experimental Hematology, Institute of Hematology, CAMS and PUMC, Tianjin 300020 , China
  • Received:2007-09-30 Revised:1900-01-01 Online:2008-06-30 Published:2008-06-30
  • Contact: XIONG Dong-sheng

Abstract: ABSTRACT:Objective To prepare monoclonal antibody (McAb) against anti-CD3 ScFv for purifying and detecting serum anti-CD3 antibody concentration. Methods McAb against anti-CD3 ScFv was prepared by hybridoma technique and used to prepare affinity chromatography column, which was used to purify anti-CD3 ScFv and Diabody [CD3×Pgp] without E-tag. The binding activities of anti-CD3 ScFv, Diabody [CD3×Pgp] without E-tag, and Diabody [CD3×Pgp] purified by anti-CD3 affinity chromatography column or anti-E-tag affinity chromatography column against K562/A02 cell and Jurket cells were detected by fluorescence activated cell sorting (FACS) method. ELISA was used to identify the specificity of the McAb. Results McAb against anti-CD3 ScFv specifically detected serum anti-CD3 ScFv without reacting with sera. The anti-CD3 ScFv purified by anti-CD3 affinity chromatography column and purified by anti-E-tag affinity chromatography column had the same specific binding activity with Jurkat cells. The positive binding rates of Diabody [CD3×Pgp] without E-tag to K562/A02 and Jurkat cells were 89.87% and 83.95%,respectively. In the competitive binding experiments with K562/A02 and Jurkat cells, the binding rates of Diabody [CD3×Pgp] without E-tag decreased to 56.30% and 43.78%, respectively. Conclusion The McAb against anti-CD3 ScFv prepared in our lab can be used to purify and detect serum anti-CD3 antibody concentration.

Key words: anti-cd3 scfv, monoclonal antibody, affinity cdromatocrapdy, diabody [cd3×pcp]