https://www.abbs.info
E-mail: [email protected]
ISSN
1672-9145
Acta Biochim Biophys Sin
2005, 37(3): 205–209
CN 31-1940/Q
Application of Phage-displayed Single Chain Antibodies in Western
Blot
Yan-Li DING1,2, Mei-Yun LIU2, Wei HAN1,2,
Sheng-Li YANG1,2, Hui LIU2*,
and Yi GONG2*
1School of Pharmaceutical Engineering, Shenyang Pharmaceutical
University, Shenyang 110015, China;
2Shanghai Research Center of Biotechnology, Chinese Academy of
Sciences, Shanghai 200233, China
Abstract A phage display single chain fragment variable library constructed
on pIII protein of M13 filamentous phage was screened using B-lymphocyte
stimulator and FP248 as selective molecules. After four rounds of panning,
there was a remarkable enrichment in the titer of bound phages. Twenty phage
clones were selected from the last round and screened by means of phage-ELISA.
With the antibody phages as primary antibodies in Western blot, we developed a
method for detecting the specific antigen. The dilutions of antibody phages
depend on the affinity between antibody-displayed phage particles and antigens.
Key words phage-display; antibody; Western blot
—————–
Received: November 23, 2004 Accepted: January 7, 2005
Abbreviations: BLyS, B-lymphocyte stimulator; ELISA, enzyme-linked
immunosorbent assay; HRP, horseradish peroxidase; IPTG, isopropyl b–D-thiogalactopyranoside;
PBS, phosphate-buffered saline; PBST, phosphate-buffered saline containing 0.5%
Tween-20; PEG, polyethylene glycol; pfu, plaque forming unit; PVDF,
polyvinylidene difluoride; scFv, single-chain fragment of variation; TMB,
tetramethylbenzidine
*Corresponding authors:
Yi GONG: Tel, 86-21-64369607; Fax, 86-21-64700244; E-mail, [email protected]
Hui LIU: Tel, 86-21-54971071; Fax, 86-21-64700244; E-mail, [email protected]