Separation of mitochondrial creatine kinase (MiMi-CK) from cytosolic creatine kinase isoenzymes by Cibachrome-Blue affinity chromatography.
The mitochondrial isoenzyme of creatine kinase (MiMi-CK) was separated by affinity chromatography on Cibachrome-Blue-Sepharose (Sepharose-Blue, Pharmacia). While the soluble CK isoforms (BB-CK and MM-CK) were specifically eluted by raising the pH of the column buffer from pH 6.0 to pH 8.0, MiMi-CK remained bound under these conditions but was specifically eluted by subsequent addition of ADP to the pH 8.0 buffer. This one-step method allows a fast and efficient separation of MiMi-CK from MM-and BB-CK isoenzymes and at the same time an enrichment of MiMi-CK by about 50-fold. Since MiMi-CK can be assayed separately after isolation by affinity chromatography on Sepharose-Blue, this method may be of clinical importance.
D000818:Animals; D002846:Chromatography, Affinity; D003402:Creatine Kinase / Q000302:isolation & purification*; D003600:Cytosol / Q000201:enzymology*; D007527:Isoenzymes; D008928:Mitochondria / Q000201:enzymology*; D012685:Sepharose / Q000031:analogs & derivatives
D007527:Isoenzymes; C017842:cibacron blue-sepharose; D012685:Sepharose; D003402:Creatine Kinase