Expression and characterization of human rheumatoid factor single-chain Fv

The variable region of heavy chain (V-H) of human rheumatoid factor (hRF) I gM was connected with the variable region of light chain (V-L) with the pep tide-linker (GGGSGGGSGGGS) by genetic engineering method and the single-cha in Fv (scFv) was expressed in E, coil, On design, scFv and scFv (tag) were planned; the Latter had a detection marker at the carboxyl-terminal, These scFvs were expressed as inclusion bodies in E. coil, purified in the presen ce of 8 M urea by gel filtration and renatured to the active form in vitro, As a control, the Fv, non-covalently associated V-H and V-L fragments, was also constructed. The 3 derivatives showed almost the same binding activit y to rabbit-IgG to which hRF is cross-reactive. ScFv (tag) was the most sta ble against urea among the 3 derivatives.