Factor VIII/Von Willebrand Factor Protein Sensitivity Of Periodic Acid Schiff Stain To Carbohydrate Deficiency

Carbohydrate (C) deficiency of the factor VUI/von Willebrand factor (f. VIII/vWf) protein has been implicated as one of the molecular defects of von Willebrand’s disease. The periodic acid Schiff (PAS) reaction in polyacrylamide gel electrophoresis (PAGE) has been used as a screening test for C deficiency. To ascertain the sensitivity of this reaction to the C content of the f. VIII/vWf protein we have purified the normal protein and compared the Coomassie to PAS staining reaction of the subunit of the intact, asialo (neuraminidase-treated), asialo-agalacto (neuraminidase and β-galactosidase-treated) derivatives and a fourth derivative which was treated with a mixed glycosidase which included neuraminidase, β-galactosidase and N-acetyl-glucosaminidase. Gels were run simultaneously, one stained with Coomassie blue and the other with PAS and at various time intervals after staining the gels were scanned.

Studies of the intact f. VIII/vWf protein revealed that the PAS and Coomassie stains could be compared from 24 hours up to 7 days with no significant changes. The desialated f. VIII/vWf protein had insignificant differences in the PAS stain compared to normal (range 81-108% of intact). However, the asialo-agalacto derivative (36% of intact) and the f. VIII/vWf treated with the mixed glycosidase (24% of intact) had significantly reduced PAS staining in relation to normal. C analysis of the different f. VIII/vWf proteins revealed that removal of the sialic acid reduced the C 30% while in the asialo-agalacto the C was reduced 47% and in the mixed glycosidase-treated material the C was reduced 78%. The PAS stain is not very sensitive to the sialic acid content of the f. VIII/vWf protein and reduction of 30% of the C does not affect the PAS stain. Since the PAS stain is relatively insensitive to significant carbohydrate deficiencies of the f. VIII/vWf protein, the use of the PAS stain in PAGE may not be an adequate test to ascertain C content of the f. VIII/vWf protein.