Skip to main content
. 2013 Nov 6;3:72. doi: 10.3389/fcimb.2013.00072

Figure 5.

Figure 5

Western blotting and affinity determination. (A) Western blotting analysis of the binding activity of scFv obtained by Skp co-expression to the TLH antigen. Left panel: SDS-PAGE results for the extracted TLH; Lane M: Protein molecular weight markers; Lanes 1, 2: extracted TLH. Right panel: Western blotting results; Lanes 3, 4: TLH band at 47 kDa bound by scFv obtained by Skp co-expression. (B) Specificity analysis of scFv obtained by Skp co-expression. BSA, KLH, and associated antigen proteins (VP1668 and YscF) of V. parahaemolyticus were coated on 96-well plates in triplicate (5 μ g/mL, 100 μ L/well). The purified scFv were added to the reaction wells and incubated for 2 h at 37°C. Specificity of Skp co-expressed scFv was determined using an anti-6×His tag antibody. (C) Affinity Determination of Skp co-expressed scFv. The affinity of anti-TLH Skp co-expressed scFv for the TLH antigen was studied by Microcalorimetry using the Nano ITC-LV. The measured data was used for the quantitative determination of the affinity constant of the anti-TLH Skp co-expressed scFv antibody.