FAQ: How much Rapid PNGase F should I use to deglycosylate my antibody sample?

The quantity of enzyme recommended is sufficient for the deglycosylation of up to 100 μg of a mouse monoclonal antibody isotype IgG2a. The optimal amount of starting material will be determined by the nature of a sample (glycan diversity) and the particular downstream N-glycan analysis that will be performed.

Typically, 30-50 µg of antibody are sufficient for complete N-glycan quantitation (after labeling by reductive amination) by HILIC followed by ESI-MS. For proteins with additional glycan sites, and/or a high glycosylation variability, more starting material might be required to quantitate all minor glycan species.

Reactions may be scaled-up linearly to accommodate larger amounts of glycoprotein and/or larger reaction volumes.