PROCESS (Steps) Preparation of Gel Column A column is filled with gel beads (e.g., Sephadex / Agarose / Polyacrylamide). These beads have tiny pores. Equilibration The gel column is washed with buffer to create a stable environment (pH & salt). Sample Loading The mixture of molecules (proteins etc.) is applied at the top of the column. Elution (Running Buffer Through the Column) Buffer is allowed to flow through the column. Molecules move down through the gel: Large molecules cannot enter the pores → move faster. Small molecules enter the pores → move slower. Collection of Fractions The solution coming out of the bottom of the column is collected in tubes. Each tube contains molecules separated by size. Analysis Fractions are checked (e.g., by spectrophotometer) to identify where the required molecule is present.