A suspension of Sepharose (40-190 j

A suspension of Sepharose (40-190 jm) in deionized water was
centrifuged at 600 x g for 10 min to pack the gel. Ten
milliliters (settled gel volume) of the gel was suspended in 10
ml of deionized water, taken to pH 10.5-11 with 4 M NaOH,
and stirred mechanically in a well ventilated hood with 1.0 g
of cyanogen bromide dissolved in 15 ml of 1-methyl-2-pyrrolidinone. The pH was maintained between 10 and 11 for
12 min. The cyanogen bromide-activated gel was transferred
to a sintered glass funnel and washed with 10 vol of ice-cold
100 mM sodium carbonate/bicarbonate buffer (pH 9.5) (buffer A)