Journal of Pure and Applied MicrobiologyVol. 9 No. 2

Production of Single Chain Fragment Variable (scFv) Monoclonal Antibody against Cry2B through Phage Display Technology

Chidanand A. Rabinal1, Narayan Moger1, Satish Verma1, P. U. Krishnaraj2 and K. N. Chandrashekara3

1Department of Biotechnology, College of Agricultural Dharwad, University of Agricultural Sciences, Dharwad, Karnataka, India. 2 Department of Agricultural Microbiology, College of Agricultural Vijayapur, University of Agriculutral Sciences, Dharwad, Karnataka, India. 3 Department of Plant Physiology and Biotechnology UPASI TRI TRF, Valparai, Tamil Nadu, India.

Received on 13 April 2015 and accepted on 09 May 2015

 

ABSTRACT

Phage display technology was used to produce recombinant monoclonal antibody against Bacillus thuringiensis crystalline protein (Cry2B) from Tomlinson I antibody library. The polyclonal ELISA had revealed the fourth round of biopan shown highest specificity (1.480) and 92.5 folds greater than that of control. Total forty-five monoclonal antibodies were screened from fourth round of biopanning against Cry2B. Among forty-five clones, pscFvCry2B19 and pscFvCry2B43 had highest specificity against target. Further, the clones pscFvCry2B19 and pscFvCry2B43 were validated for their cross reactivity with other Cry proteins and the clones were sequenced with LMB3 forward and pHEN reverse primers. The scFv fragments of pscFvCry2B19 and pscFvCry2B43 were of 746 bp and 747 bp long respectively and almost similar to each other at DNA and amino acid level except at C? terminal end. The produced scFv monoclones have the potential to detect the Cry2B antigen.

Keywords : Phage display technology; scFv; Biopanning; ELISA; Cry2B antigen; Bacillus thuringiensis.