[Screening and identification of a single chain antibody fragment].
OBJECTIVE
To screen and identify anti- Aflatoxin B1 single chain antibodies (scFv) from Tomlinson (I) library.


METHODS
The phages absorbed on the ELISA plate were eluted by four methods including glycine buffer elution, trypsin elution, AFB1 competitive elution and competitive elution following trypsin treatment. The phage positive clones were transformed into Escherichia coli HB2151 and soluble scFv protein was expressed with the induction of Isopropyl beta-D-1-Thiogalavtopy ranoside (IPTG). The scFv was identified by ELISA and DNA sequence.


RESULTS
Comparing the four methods, we found that the most efficient way to get positive clones was competitive elution following trypsin treatment. Obtainning two positive clones that could bind specifically with free AFB1, which their relative affinity were 0.4 microg/mL and 0.7 microg/mL, respectively. DNA sequencing results showed that the scFv belonged to human immunoglobulin variable region.


CONCLUSIONS
The specific human scFv could be obtained with phage antibody library, and our method provided an alternative for screening recombinant antibody against anti-hapten.
Wang
Tiebin
T
State Key Laboratory of Food Science and Technology, School of Food Science, Jiangnan University, Wuxi 214122, China. wangtiebin000@sina.com
Ding
Husheng
H
Yang
Lian
L
Ji
Qing
Q
Chen
Wei
W
Zhang
Hao
H
D016604:Aflatoxin B1 / Q000276:immunology*; D000906:Antibodies / Q000235:genetics / Q000276:immunology*; D000917:Antibody Formation; D000918:Antibody Specificity; D019411:Clinical Laboratory Techniques; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D007128:Immunoglobulin Fragments / Q000276:immunology / Q000302:isolation & purification*; D007135:Immunoglobulin Variable Region; D007158:Immunologic Techniques
D000906:Antibodies; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D016604:Aflatoxin B1