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. 2020 Jul 14;79(10):1370–1380. doi: 10.1136/annrheumdis-2020-217072

Figure 1.

Figure 1

ELISA assay for N-terminal (NT)/C-terminal (CT) SIRT1 ratio. Diluted serum or conditioned media are incubated with either NT-reactive or CT-reactive antibodies and concentrations are calculated based on the respective standard curve. The plate configuration is added next to the standard curve illustration (A). Each serum sample is either incubated with an NT-reactive or CT-reactive antibody (B), wherein ratio between the NT and CT concentrations denoted NT/CT SIRT1 ratio. (C) Possible scenarios related to this ELISA method: (1) an equal NT and CT ratio of 1 indicates that there is no increase in either protein domains of SIRT1 in examined samples; (2) cleavage of both NT and CT domains will render undetected; (3) increased NT concentration as compared with CT concentration rendering a value over 1; (4) increased CT concentration as compared with NT concentration rendering a value less than 1. Notably, using this method, one cannot distinguish between full length and cleaved variants of SIRT1, rather NT-reactive or CT-reactive polypeptides of SIRT1.