Fibroblast growth factor 21(FGF21) is a protein that inmammals is encoded by the FGF21 gene. The proteinencoded by this gene is a member of the fibroblastgrowth factor (FGF) family and specifically a memberof the endocrine subfamily which includes FGF23 andFGF15/19. FGF21 is the primary endogenous agonist ofthe FGF21 receptor, which is composed of the co-receptors FGF receptor 1 and β-Klotho. Besides, Klotho Beta (KLb) has been identified as an interactor of FGF21, thus a binding ELISA assay was conducted to detect the interaction of recombinant human FGF21 and recombinant human KLb. Briefly, FGF21 were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100μl then transferred to KLb-coated microtiter wells and incubated for 2h at 37℃. Wells were washed with PBST and incubated for 1h with anti-KLb pAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody, wells were aspirated and washed 3 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50μl stop solution to the wells and read at 450nm immediately. The binding activity of KLb and FGF21 was shown in Figure 1, and this effect was in a dose dependent manner.