Complex 4 ab109876 enzyme activity dipstick assay kit

Activity of complex I and complex IV was assessed according to the protocol of the Complex I (ab109720)/Complex IV (ab109876) Enzyme Activity Dipstick Assay Kit (Abcam, Cambridge, United Kingdom). In this technique the proteins from cellular lysates migrate through a nitrocellulose membrane. Then, Complex I is immunocaptured (i.e., immuno-precipitated in active form) on the dipstick. Then, the dipstick is immersed in Complex I activity buffer solution containing NADH as a substrate and nitrotetrazolium blue (NBT) as the electron acceptor. Immunocaptured complex I oxidizes NADH and the resulting H+ reduces NBT to form a blue-purple precipitate at the Complex I antibody line on the dipstick. The signal intensity of this precipitate corresponds to the level of Complex I enzyme activity in the sample. Dipsticks images were taken with a Molecular Imager ChemiDoc XRS+ System (BIORAD, Hercules, CA, United States) and quantified by the Image Lab software.

Activity of complex I and complex IV was assessed according to the protocol of the Complex I (ab109720)/Complex IV (ab109876) Enzyme Activity Dipstick Assay Kit (Abcam, Cambridge, UK) starting from a cellular pellet. In this technique the proteins from cellular lysates migrate through a nitrocellulose membrane. Then, Complex I is immunocaptured (i.e. immuno-precipitated in active form) on the dipstick. Then, the dipstick is immersed in Complex I activity buffer solution containing NADH as a substrate and nitrotetrazolium blue (NBT) as the electron acceptor. Immunocaptured complex I oxidizes NADH and the resulting H+ reduces NBT to form a blue-purple precipitate at the Complex I antibody line on the dipstick. The signal intensity of this precipitate corresponds to the level of Complex I enzyme activity in the sample. 30 μg of protein were used for each assay, according to the protocol range. Three replicates were performed for each respiratory complex assay.
Images of the dipsticks were acquired with a Molecular Imager ChemiDoc XRS+ System (BIORAD, Hercules, CA, USA) and quantified with the Image Lab software.