[Preparation and identification of monoclonal antibody against human Flt-1].

AIM: To prepare functional monoclonal antibodies(mAb)against recombinant human Flt-1(rhFlt-1).METHODS: A cell line stable secreting mAb was established by using FLT-1 extracellular domain III as antigen and hybridoma technique.Then it was purified in large-scale from mouse ascites by protein G affinity chromatography.The characteristics of mAb were then determined by ELISA,Western blotting and FACS.RESULTS: The immunoglobin subtype of mAb XA12 was IgG1 with kappa(κ) light chains,and it could recognize rhFlt-1 specifically.Furthermore,mAb XA12 could bind to rhFlt-1with high affinity(K=1.28±0.05 nmol/L).It could also be used to detect Flt-1-positive cells,such as human umbilical vascular endothelial cells(HUVECs) and K562/A02 in a dose-dependent fashion.CONCLUSION: A hybridoma cell line secreting functional anti-rhFlt-1 mAb was successfully prepared.The antibody can be used to study the function of Flt-1 and further potentially optimized for clinical purpose.