Microcolumn chromatofocusing using slurry-packed Mono P anion-exchanger beads as chromatographic matrix

Chromatofocusing of proteins in a microcolumn format (750 mum i.d.) was evaluated with slurry-packed Mono P anion-exchanger beads (10 pin bead size) as chromatographic matrix. Evaluation was performed with three standard proteins in terms of flow rate, Polybuffer 96-acetic acid, pH 6.0, concentration (eluent buffer strength), and detection limit. A column with the dimension 750 pin i.d. x 50 mm length, a flow rate of 20 muL min(-1),and a 1:10-1:20 Polybuffer 96-acetic acid aqueous dilution showed the best resolution between the proteins. Detection limits were 3 mug mL(-1) for carbonic anhydrase 1 and 6 mug mL(-1) for myoglobin A employing UV-defection at 280 nm.