The F(ab)₂ Fragmentation kits are designed for the generation and isolation of F(ab)₂ fragments from IgG molecules. The kit utilises Immobilised Pepsin resin. Pepsin is a proteolytic enzyme that is routinely used for the generation of F(ab)₂ fragments from immunoglobulin G (IgG). One or more of the disulfide bonds that join the heavy chains in the hinge region are preserved, so the two Fab regions of the antibody remain joined together, yielding a divalent molecule (containing two antibody binding sites), hence the designation F(ab)₂. The light chains remain intact and attached to the heavy chain, whereas the Fc fragment is digested into small peptides. The Immobilised Pepsin offers the distinct advantage of eliminating enzyme contamination of the F(ab)₂ fragments.
Following pepsin digestion the F(ab)₂ fragments are separated from undigested IgG and the large Fc fragments with the supplied Protein A Spin Column. The Protein A Resin binds the IgG and Fc molecules and the F(ab)₂ are rapidly collected due to the spin-format design. In addition, SpinOUT™ GT-600 desalting columns are supplied to ensure the initial antibody sample is in the optimal condition for F(ab)₂ Fragmentation. The F(ab)₂ Fragmentation Kit is optimised for mouse, rabbit and human IgG, using 0,25 to 4 mg/ 0,5 ml sample or using 25 to 250 μg/125 μl sample with the F(ab)₂ Fragmentation (Micro) Kit.