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Published on: February 27, 2020
Immunoaffinity purification and quantification of antibody-toxin conjugates
1Drug Targeting Laboratory, Institute of Cancer Research, Sutton, Surrey, UK.
Methods in Molecular Biology (Clifton, N.J.)
|February 22, 2012
Summary
Antibody-toxin conjugates, vital for targeted therapies, are purified using gel permeation chromatography. This method effectively removes impurities but struggles to separate unconjugated antibodies from the final conjugate product.
Area of Science:
- Bioconjugation Chemistry
- Protein Chemistry
- Immunotherapy
Background:
- Antibody-toxin conjugates are engineered using antibodies and cytotoxic proteins like ribosome-inactivating proteins (RIPs).
- Chemical crosslinking is a common method for preparing these antibody-drug conjugates.
Purpose of the Study:
- To describe the purification process of antibody-toxin conjugates.
- To highlight the limitations of current purification techniques.
Main Methods:
- Antibody-toxin conjugates are synthesized via chemical crosslinking.
- Gel permeation chromatography (GPC) is employed as an initial purification step.
Main Results:
- GPC effectively removes protein aggregates, excess RIPs, and low molecular weight byproducts.
- A significant portion of unconjugated antibody remains in the conjugate preparation.
- Size exclusion chromatography alone is insufficient for complete separation of conjugate from unconjugated antibody.
Conclusions:
- While GPC is a valuable initial step, further methods are needed to achieve high purity of antibody-toxin conjugates.
- The presence of unconjugated antibodies impacts the efficacy and specificity of these targeted therapies.
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