Say Hello to IgASAP™ Sub1+2 Lyophilized!

IgASAP is a family of IgA-digesting enzymes. IgASAP Sub1 is an IgA1-specific enzyme that digests human IgA1 at one specific site above the hinge, and the enzyme we’re launching today, IgASAP Sub1+2, specifically digests IgA1 and IgA2m1 above the hinge. Both enzymes generate intact and homogenous Fab and Fc fragments.
 
The IgASAP enzymes enable generation of intact monovalent Fab fragments as well as middle-level analysis of IgA1, which facilitates IgA1 characterization during, for example, the development of vaccines, therapeutics, and diagnostics.
 

• Unique enzymatic tools that enable middle-level characterization and quality control of IgA
• Generates homogeneous Fab and Fc fragments with preserved immunoreactivity
• Highly specific – one digestion site above the hinge of human IgA

Site-specific Digestion of Human IgA2m1

To demonstrate the site-specificity of IgASAP Sub1+2, a commercially available recombinant human IgA2m1 was digested with IgASAP Sub1+2 Lyophilized overnight at 37°C, and compared to the undigested antibody. To facilitate data interpretation, the samples were treated with PNGaseF Lyophilized under denaturing conditions for removal of all Fab and Fc N-glycans. The denatured samples were analyzed with LC-MS (Fig. 1), and the acquired masses of the homogenous scFc and Fd fragments generated from IgASAP Sub1+2 hydrolysis confirm the single digestion site on human IgA2m1 at VTVPCP / VPPPPP (aa 102/103 of constant heavy chain acc. to Uniprot A0A0G2JMB2). The high-resolution mass data demonstrates the value of using middle-level analysis for characterization of the IgA2m1 antibody.
 

 
Figure 1. Site-specific digestion of IgA2m1. a) Schematic illustration of the workflow. Deconvoluted mass spectra of N-deglycosylated b) heavy chain (HC) of a commercially available recombinant IgA2m1 and the corresponding c) scFc fragment and d) Fd fragment after digestion using IgASAP Sub1+2 in an overnight reaction at 37°C. The samples were separated by reversed-phase chromatography (ACQUITY Premier Protein BEH C4, 300 Å, 1.7 µm 2.1 x 100 mm, Waters™) and analyzed with ESI-QTOF MS (Bruker Impact II).
 

IgASAP Sub1+2 Lyophilized – Above hinge digestion of IgA1 and IgA2m1.