Top-Down MS and MS Methods for Bruker Instruments

This documents shows standard methods for protein mixture preparation, separation by LC as well as parameters for successful MS and MS measurements.  These parameters are not set for ultimate performance but rather offer a dependable starting point to help you become successful in your research.

Top-Down MS and MS Methods for Thermo Instruments

This documents shows standard methods for protein mixture preparation, separation by LC as well as parameters for successful MS and MS measurements.  These parameters are not set for ultimate performance but rather offer a dependable starting point to help you become successful in your research.

Fragmentation of the hinge region of IgG, producing a F(ab’)2 fragment and two Fc fragments

FabRICATOR® is a digestive enzyme that cleaves IgG only at one specific site below the hinge region resulting in F(ab’)2 and Fc-fragments. Since FabRICATOR® only cleaves at one specific site below the hinge region, there is no risk of over digestion if the incubation time is increased for several hours.

Fragmentation of human IgG1 between K223 and T224 producing a homogenous pool of Fab and Fc fragments

GingisKHAN™ digests human IgG1 at one single site in the upper hinge (…KSCDK / THTCPPCP…). A second cleavage site on the Fc may appear if the N-glycans are removed. GingisKHAN™ is a cysteine protease and requires reducing environment to be active. Intact Fab and Fc fragments are obtained with GingisKHAN™ digestion of human IgG1 since mild reducing conditions (i.e. 2 mM cysteine) is sufficient for enzyme activity.