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http://dx.doi.org/10.7845/kjm.2015.5015

The development of murine recombinant single-chain variable domain fragment (ScFv) specific to acute non-lymphocytic leukemia (ANLL) cell line HL60  

Kim, Cheol Hong (Department of Microbiology, College of Natural Sciences, Changwon National University)
Han, Seung Hee (Department of Microbiology, College of Natural Sciences, Changwon National University)
Kim, Hyeong Min (Department of Microbiology, College of Natural Sciences, Changwon National University)
Han, Jae Yong (Department of Agricultural Biotechnology, Seoul National University)
Lim, Myeong Woon (Joongkyeom: Antibody Therapy Inc.)
Kim, Jin-Kyoo (Department of Microbiology, College of Natural Sciences, Changwon National University)
Publication Information
Korean Journal of Microbiology / v.51, no.2, 2015 , pp. 115-125 More about this Journal
Abstract
A monoclonal antibody AP64 IgM binds to human acute nonlymphocytic leukemia (ANLL) cell line HL60 and also cross-reacts with the homologous antigen in a rat ANLL cell. This antibody mediated by complement, has leukemia a suppression effect. In this study, we generated a recombinant single-chain variable domain fragment (ScFv) which were derived from $V_H$ and $V_L$ cDNA of AP64 IgM-secreting hybridoma by RT-PCR. The two variable regions were joined with a single 15 amino acid linker $(G_4S)_3$. This recombinant ScFv was expressed as a single polypeptide chain from Escherichia coli BMH 71-18. The recombinant ScFv was purified by applying the periplasmic extract to $Ni^+$-NTA-agarose affinity column and detected with westernblot. The purified recombinant ScFv recognized a surface antigen (about 30 kDa) of HL60 cell line which is the same antigen detected by parental AP64 IgM. But the affinity of ScFv for a surface antigen of HL60 was lower than that of the parental AP64 IgM, which needs to be further improved. Overall, the recombinant ScFv specific to HL60 might be a useful bioreagent for either diagnostic or therapeutic purposes.
Keywords
human acute nonlymphocytic leukemia; HL60; IgM; $Ni^+$-NTA-agarose; single-chain Fv; westernblot;
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