Generation of Homogeneous Fab Fragments using FabALACTICA™

Performance Activity

Simple preparation of highly pure Fab fragments from IgG1 antibodies using FabALACTICA digestion combined with Fc affinity purification.

Fab fragments preserve the antigen-binding specificity of antibodies while eliminating Fc-mediated interactions that can introduce confounding biological effects. Generating them in a highly pure and homogeneous form is critical to ensure that measured binding behaviour accurately reflects Fab-antigen interactions, enabling robust, reproducible, and interpretable analytical and functional studies.

Here we demonstrate how we can use the FabALACTICA Fab Kit to generate and then purify homogeneous Fab fragments. The kit comprises two column types: FabALACTICA Immobilized for antibody digestion and a CaptureSelect™ Fc column for affinity purification. The CaptureSelect Fc column will bind the Fc fragments and any undigested antibody, allowing the Fab fragments to pass through the column.

Preparation of homogeneous Fab fragments

We have demonstrated efficient Fab preparation from three human IgG1 antibodies; adalimumab, trastuzumab and cetuximab. In all cases we have analysed the digested material eluted from the immobilized FabALACTICA column and the purified Fab fragments to visualise the purification process. Both the efficiency of the digestion and the purity of the Fab fragments can easily be observed here.

Robust digestion of human IgG1 using FabALACTICA

Preparation of human IgG1 Fab fragments using FabALACTICA Fab Kit. Schematic representation of how to prepare Fab fragments using FabALACTICA Fab Kit. Samples are digested using FabALACTICA Immobilized before affinity purification using CaptureSelect™ Fc (left). SDS-PAGE demonstrating the homogeneity of the purified Fab fragments from three human IgG1 antibodies; adalimumab, trastuzumab and cetuximab.

Related Products

Resources