Evaluation of sodium dodecyl sulfate non-acrylamide, polymer gel-filled capillary electrophoresis for molecular size separation of recombinant bovine somatotropin.
Sodium dodecyl sulfate (SDS) non-acrylamide, polymer gel-filled capillary electrophoresis was examined as an alternative to the high-performance size-exclusion chromatographic (HPSEC) method for the analysis of recombinant bovine somatotropin (bovine growth hormone, rbSt). A calibration curve for the molecular mass of protein standards (M(r) 14,000 to 97,000) was linear (r2 = 0.998) when the molecular mass of the proteins and their peak migration time were plotted on a logarithmic (log-log) scale. Relative standard deviation (R.S.D.) of the molecular mass determination was approximately 2-3%. A pre-production as well as a production lot of the SDS gel-filled capillary columns were examined. Performance of both of these columns were equivalent. Peak migration time remained relatively constant beyond 140 sample injections. Gradual loss of theoretical plates was noted over the course of the assay; however, the peak resolution remained adequate for the analysis of rbSt. Peaks corresponding to monomer, dimer, trimer, and tetramer of rbSt were base-line resolved by the SDS gel-filled capillary electrophoresis. Theoretical plate of the monomer peak was approximately 28,000-30,000 per 40 cm column. Both the monomer and the dimer recovery studies indicated that the calibration curves are linear (r2 > 0.993) and the slopes are not different from one. Amounts of the components in rbSt determined by the SDS gel-filled capillary electrophoresis compared well with those of the HPSEC method. The results of this study indicated that the SDS non-acrylamide gel-filled capillary electrophoresis may be a viable alternative to the HPSEC method for the molecular size separation and analysis of recombinant proteins.
Tsuji
K
K
Pharmaceutical Product Control Division, Upjohn Company, Kalamazoo, MI 49001.
D000818:Animals; D002417:Cattle; D004586:Electrophoresis / Q000379:methods*; D005069:Evaluation Studies as Topic; D013006:Growth Hormone / Q000302:isolation & purification*; D008970:Molecular Weight; D011994:Recombinant Proteins / Q000302:isolation & purification; D012967:Sodium Dodecyl Sulfate
D011994:Recombinant Proteins; D012967:Sodium Dodecyl Sulfate; D013006:Growth Hormone