EXAMINATION OF CAPILLARY ZONE ELECTROPHORESIS, CAPILLARY ISOELECTRIC-FOCUSING AND SODIUM DODECYL-SULFATE CAPILLARY ELECTROPHORESIS FOR THE ANALYSIS OF RECOMBINANT TISSUE-PLASMINOGEN ACTIVATOR

The microscale techniques of CZE, cIEF and SDS capillary electrophores is have been evaluated for the analysis of a complex glycoprotein, rec ombinant tissue plasminogen activator (rtPA). A series of omega-amino acid buffers (pH approximate to 5) was found suitable for the CZE sepa ration of rtPA on coated capillaries. rtPA could be resolved into a se ries of major and minor peaks in an epsilon-aminocaproic acid buffer c ontaining 0.01% (v/v) Tween 80. For cIEF, a two step method with press ure mobilization was utilized. Using a commercial instrument, either a polymer solution with a 50 mu m I.D. capillary or narrow bore capilla ries without a polymer solution (25 mu m I.D.) were employed. rtPA was resolved into at least eight species within a pi range of 6.4-9.2 usi ng Ampholine 3.5-10. Migration time precision for the major peaks rang ed from 0.2% for CZE to less than or equal to 2-3% R.S.D. for cIEF. To tal recovery of rtPA from the capillary was also demonstrated for both methods. Analysis of rtPA, rtPA Type I, rtPA Type II and the desialyl ated forms resulted in the expected elution profiles. Finally, the pot ential of SDS capillary electrophoresis using a coated capillary for a n rtPA Type I/Type II purity assay was shown.