T7 RNA polymerase uses double-stranded DNA containing the T7 promoter sequence (5’-TAATACGACTCACTATAG*-3’) as a template and NTPs as substrates to synthesize RNA complementary to the reverse strand of DNA downstream of the promoter. Both linear double-stranded DNA with blunt ends or 5’ overhangs can serve as template substrates for T7 RNA polymerase, meaning linearized plasmids or PCR products can be used as templates for in vitro RNA synthesis. This kit employs a sandwich enzyme-linked immunosorbent assay (ELISA) principle to detect residual T7 RNA polymerase. T7 RNA polymerase standards and test samples are added to microplate wells pre-coated with anti-T7 RNA polymerase antibodies (36705-A). Diluted biotin-labeled T7 RNA polymerase detection antibodies (36705-C) are then added, followed by Streptavidin-HRP (SA-HRP) (36705-D), forming an antibody + antigen + antibody-biotin + SA-HRP complex. After washing, TMB substrate (36705-H) is added for color development. TMB is catalyzed by HRP to transition from colorless to blue and finally to yellow upon addition of stop solution (36705-I). The intensity of the yellow color is positively correlated with the amount of T7 RNA polymerase detected in the sample. Validation Report Features High Sensitivity: Quantitative lower limit (LOQ) of 1 ng/mL; detection limit (LOD) as low as 0.318 ng/mL. High Precision: Intra-assay CV
