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Biochemistry (Moscow)

, Volume 73, Issue 6, pp 702–710 | Cite as

Single-chain antibody against human lipocalin-type prostaglandin D synthase: Construction, expression, purification, and activity assay

  • De-Yu Chen
  • Li-Min Liu
  • Sheng-Jie Liu
  • Mao-Ying Zhu
  • Lan Xu
  • Tian-Hua Huang
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Abstract

An active form of single-chain antibody (ScFv) from murine monoclonal antibody 4A7, which is specific for lipocalin-type prostaglandin D synthase (L-PGDS), was produced in Escherichia coli. The complementary DNA fragments encoding the variable regions of heavy chain (VH) and light chain (VL), which amplified from hybridoma 4A7 producing a monoclonal antibody (IgG1) against L-PGDS, were connected by a (Gly4Ser)3 linker using an assembly polymerase chain reaction. The resultant ScFv were cloned into the vector pGEM and expressed in E. coli as inclusion bodies. The expressed ScFv fusion proteins were purified by Ni2+-nitrilotriacetic acid chromatography. The purity and activity of purified ScFv were confirmed by SDS-PAGE and ELISA. The result revealed that 4A7 ScFv conserved the same characteristics of specific recognition and binding to sperm as the parental 4A7 monoclonal antibody.

Key words

prostaglandin D synthase (L-PGDS) single-chain antibody (ScFv) expression 

Abbreviations

L-PGDS

lipocalin-type prostaglandin D synthase

MAb

monoclonal antibodies

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Copyright information

© MAIK Nauka 2008

Authors and Affiliations

  • De-Yu Chen
    • 1
    • 2
  • Li-Min Liu
    • 1
  • Sheng-Jie Liu
    • 2
  • Mao-Ying Zhu
    • 2
  • Lan Xu
    • 1
  • Tian-Hua Huang
    • 1
  1. 1.Research Center for Reproductive Medicine, Department of Cell Biology and GeneticsShantou University Medical CollegeShantouChina
  2. 2.Department of BiologyFuyang Normal CollegeFuyangChina

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