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lon-exchange chromatography . Separation basis: net charge of proteins at a given pH cation exchange chromatography . Resin: negatively charged resin: e.g., CM-cellulose, S-Sepharose positively charged resin: e.g., DEAE-cellulose, Q-Sepharose Macmillan Learning Positively charged . How to elute: protein binds to negatively charged Increase salt concentration (e.g., NaCl) to compete with protein binding. bead Gradually change pH to alter protein charge and reduce binding affinity. CH3 H,C H2 H2 \\+H Macmillan Learning Negatively charged CH3 Cellulose Cellulose protein flows through or or H2 H 2 agarose agarose Carboxymethyl Diethylaminoethyl (CM) group (DEAE) group (ionized form) (protonated form) 8 Figure 4.4 and Unnumbered 4.2 page 104 and Figure 3.27, Biochemistry, 10th ed