http://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 DING^1,2, Mei-Yun LIU², Wei HAN^1,2, Sheng-Li YANG^1,2, 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]