3. Sample Preparation- The sample is prepared in the mobile phase and it should be free of suspended particle to avoid clogging of the column. The most recommended method to apply the sample is to inject the sample with a syringe.

4. Elution- There are many ways to elute a analyte from the ion-exchange column. (1) Isocratic elution (2) Step-wise gradient (3) Continuous gradient either by salt or pH (4) affinity elution (5) displacement chromatography 

5. Column Regeneration- After the elution of analyte, ion-exchange chromatography column require a regeneration step to use next time. column is washed with a salt solution with a ionic strength of 2M to remove all non-specifically bound analytes and also to make all functional group in a iononized form to bind fresh analyte.

Figure 30.4 : Operation of the Ion-exchange Chromatography. (A) Chromatography system to perform gradient elution of analytes to give an (B) elution profile.