The COP9 signalosome (CSN) is a multimeric protein complex that occurs in all 
eukaryotic cells. Originally described in plants as a regulator of 
photomorphogenesis, its purification and characterization from mammalian cells 
revealed significant sequence homologies to subunits of the 26S proteasome lid 
complex, as well as of the eukaryotic translation initiation factor 3. Recent 
studies disclosed its participation in processes such as DNA repair, cell cycle 
regulation, development, and angiogenesis. At the moment, the pleiotropic 
effects of the CSN point to a regulatory role in the ubiquitin/26S proteasome 
system, but its exact function still remains to be clarified. This chapter 
describes the method to purify human CSN from red blood cells. Two outdated 
erythrocyte concentrates are sufficient to prepare approximately 0.5 mg of CSN. 
Washed cells are first lysed and then proteins are separated by a DEAE 
anion-exchange column. The CSN-containing fractions are pooled and subjected to 
an ammonium sulfate precipitation followed by dialysis. The concentrated 
proteins are then loaded onto a glycerol density gradient and 
ultracentrifugation is performed. The purification procedure is continued using 
two succeeding anion-exchange columns, resulting in a sufficiently pure CSN 
complex. Optionally, an additional density gradient centrifugation can be 
attached. The purified CSN complex possesses kinase, deneddylase, and 
deubiquitinase activities and can be stored for at least 2 months on ice at 4 
degrees .