SMT3 (yeast SUMO) sandwich ELISA with immobilized SMT3-A1(Cys)
This application involves the identification of antigens in solution by sandwich ELISA using nanoCLAMPs covalently conjugated to maleimide coated microtiter wells via their lone C-terminal Cys. Antigen bound by the immobilized nanoCLAMP is identified by an enzyme-conjugated secondary antibody followed by development with the appropriate secondary substrate.
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Analysis of the interaction between SMT3-A1(Cys) and SMT3-GFP-V5 by sandwich ELISA. SMT3-A1(Cys) was conjugated to maleimide coated microtiter wells of 96 well strips via the lone C-terminal Cys, at 5 µg/ml. Dilutions of SMT3-GFP-V5 were incubated on the strips, washed, and then incubated with anti-V5-HRP and developed with TMB Ultra. Reactions were stopped with H2S04 and absorbance measured at A450. The blank wells were processed the same, except they included no SMT3-A1(Cys).