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Immunoaffinity Purification using LC-MS/MS: Comparing Two Methods

Protein quantitation by liquid chromatography-tandem mass spectrometry (LC-MS/MS) has become an increasingly popular field for pharmacokinetics study in the drug discovery phase. Traditional method development using a ligand binding assay usually requires 2-3 months. By using LC-MS/MS techniques, development times can be reduced to one week. In this white paper, we discuss testing the robustness of membrane affinity versus magnetic beads by using both methods to analyze antibody samples.

The magnetic bead approach and membrane approach produced comparable validation parameters including linearity, intra-, inter-day accuracy and precision, carryover, dilution integrity, matrix effect, specificity, and selectivity.

The two validated strategies for protein immunoaffinity purification were compared and both methods offered the same LLOQ of 0.05 μg/mL. The main differences was with the sample processing time. Specifically, the sample immunoaffinity purification (IP) processing time for a 96-well plate with the magnetic bead method was 3-4 hours in comparison to the 10-20 minutes by the membrane approach. The magnetic bead method was also more prone to operational pipetting errors.


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WuXi AppTec is working to revolutionize bioresearch by advancing discovery efforts and delivering the high-quality data clients need. By working closely with chemistry, biology and pharmacology for effective lead selection during discovery, our team is able to provide you with the expertise across a diverse range of compound classes with a breadth of experience in a variety of species, strains, matrices and microsampling techniques.

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Established in December 2000, WuXi AppTec provides a broad portfolio of R&D and manufacturing services that enable companies in the pharmaceutical, biotech and medical device industries worldwide to advance discoveries and deliver groundbreaking treatments to patients. With industry-leading capabilities such as R&D and manufacturing for small molecule drugs, cell and gene therapies, and testing for medical devices, WuXi AppTec’s open-access platform is enabling more than 4,000 collaborators from over 30 countries to improve the health of those in need – and to realize our vision that “every drug can be made and every disease can be treated”.

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