When working with native IgG, we recommend using 1 µL of Endo S per 100µg of native IgG. When working with other various glycoproteins (such as Fetuin) we recommend using a modified denaturing protocol, detailed below. Combine 100 µg of glycoprotein, 1 µL of 400mM DTT and H2O (if necessary to make a 10 µL total reaction volume. Denature glycoprotein by heating reaction at 55°C for 10 minutes. Make a total reaction volume of 20 µL by adding 2 µL 10X GlycoBuffer 1, H2O and 1-2 µL Endo S. Incubate reaction at 37°C for 1 hour. Note: To deglycosylate various glycoproteins, longer incubation times as well as more enzyme may be required. Reactions may be scaled-up linearly to accommodate larger reaction volumes.