Amgen Inc.
SEQUENTIAL DIGESTION OF POLYPEPTIDES FOR MASS SPECTROMETRIC ANALYSIS

Abstract:

The disclosed methods are directed to preparing polypeptides for multi-attribute analysis. The polypeptides are optionally denatured, reduced, and/or alkylated before being subjected to a first digestion. Following the first digestion the large and small fragments resulting from the digestion are separated from each other. A second digestion is then performed on the larger of the fragments. All of the fragments from the two digestions are then analyzed chromatographically, electrophoretically, or spectrometrically, or a combination of these methods. The methods are especially useful for the preparation of therapeutic polypeptides for analysis, especially those that are not easily cleaved.