SMCC (N-Succinimidyl 4-(N-maleimidomethyl)cycl) is a heterobifunctional protein crosslinker.

N-Succinimidyl 4-(N-maleimidomethyl)cycl（SMCC) is an amine-to-sulfhydryl crosslinker that contains NHS-ester and maleimide reactive groups at opposite ends of a medium-length cyclohexane-stabilized spacer arm (8.3 angstroms). SMCC conjugation occurs via a maleimide group that is sulfhydryl (thiol; -SH) reactive and a NHS ester group that is amine reactive and forms stable, covalent protein crosslinks.

Branched PEI (25 kDa) was dissolved in PBS buffer to yield a concentration of 1 mg/mL. No-weigh Sulfo-SMCC was dissolved in Milli-Q water with a concentration of 10 mg/mL, then a certain of the prepared no-weigh SulfoSMCC solution was added into 2 mL PEI solution to incubate for 30 min at 37 C. After the PEI-SMCC polymer was formed, TAT solution was added into the polymer at the given molar ratio and incubated for 2 h at 4 C. The concentration of unreacted SMCC and unreacted TAT were removed using Zeba Spin Desalting Columns. The polymer PEI-SMCC-TAT was also characterized and analyzed through FTIR to check if each element was added to the PEI backbone.