Objective:To establish and verify a tech. method for determination of the purity of recombinant human serum albumin interferon α2 b fusion protein(rIFNα2 b-HSA) by ion exchange chromatog.(IECHPLC). Method:The IEC-HPLC chromatog. systems were applied for the quantification of α2 b protein by TSK gel Q-STAT column(3.0 mm x 10 cm) with mobile phase A[Tris(HCl)] buffer and B[Tris(HCl) NaCl buffer] under the gradient elution conditions: 0-3 min, 100%A; 3-18 min, 100%A → 0%A; 18-21 min, 0%A; 21-22 min, 0%A→ 100%A, and 20 μL of theα2 b sample was applied and detective on 25 °C, 0.5 mL·min∼(-1) flow rate and 214 nm. The system applicability,specificity,linearity,precision and limit of quantification of this method were investigated,and compared with the results obtained by reversed-phase chromatog. and electrophoresis. Results:System applicability:the separating degree of IEC-HPLC method for separation of impurities and main peak was higher than 1.5,the number of theor. plates was higher than 20 000;Specificity:the sample was treated with NaOH solution,the main peak and impurity peak are well separated;the linear range was 0.05-1 mg·m L∼(-1)(R2=0.997 4);method precision(RSD) was 0.52%(n=6);the limit of quantification was 0.001 8 mg·mL∼(-1). Three batches of samples were purified by 3 methods,and the purity was all higher than 99.0%. Conclusion:A methodol. study of this method is performed,and the results meet the relevant requirements of the"Chinese Pharmacopoeia(2015 edition)" and"Guidelines for the Verification of Drug Quality Standard Anal. Methods",and this method is suitable for determination of rIFNα2 b-HSA purity.