IgG antibodies are immunoglobulin homodimers of heterodimers; they contain 2 heavy chains (50 kDa) each paired with a light chain (25 kDa). Heavy and light chains, as well as their dimerization, are stabilized through covalent disulfide bridges. Enzymatic cleavage of specific disulfide bonds results in fragments of differing composition.
The use of papain, an enzyme derived from papaya, results in reduction of exposed disulfide bonds in the hinge region of the antibody. This separation, or fragmentation, breaks the the antibody into three segments; 2 Fabs consisting of equal parts heavy chain and light chain, and 1 Fc region. Pepsin, another protease, preferentially digests the Fc region of the antibody, but leaves intact the hinge region, stabilizing the pairs of heavy and light chain regions that recognize antigen, resulting in bivalent Fabs or F(ab')2. Purification post-fragmentation is performed to remove unwanted Fc region, accomplished through protein A/G affinity or size exclusion chromatography.
| Antibody Fragment Format | Digestion Enzyme | Service Items |
|---|---|---|
| Fab | papain | Fab Fragmentation |
| F(ab')2 | pepsin | F(ab')2 Fragmentation |
Antibody fragmentation through enzymatic cleavage requires sufficient amounts of whole IgG. Although IgG is mostly uniform, fragmentation conditions for each antibody require optimization with initial pilot study analysis. Additionally, fragmentation reactions and purification have a typical yield of 40-60%. Should you require production of sufficient antibody quantities prior to fragmentation, our Antibody Manufacturing Solutions pair effortlessly with antibody fragmentation.
No antibody to fragment yet? Let GenScript make one for you. All our service offerings seamlessly integrate to create your custom solution and facilitate your research. Start with one of our popular antibody generation services below:
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