Rapid, On-Line Desalting of Intact Proteins prior to Characterization by Mass Spectrometry

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Paul D. Rainville, Thomas E. Wheat, Claude R. Mallet, and Jeff R. Mazzeo
WCBP, January 10th-13th, 2005
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Therapeutic proteins can undergo structural modification during production, storage, and stability testing. Alterations can affect safety and efficacy, therefore reliable determination and continuous monitoring of the protein properties is essential. Since structural changes are reflected in molecular weight, measuring this property is one convenient way to monitor modifications. Methods including electrophoresis, chromatography, and ultracentrifugation have been used; however, these methods measure average relative errors from 10-100%. This large error range is due to these methods being affected by properties other than molecular weight such as conformation, Stokes’ radius, and hydrophobicity. Mass spectrometry is a technique whereby more exact molecular weight information can be obtained. However, the proteins are often dissolved in buffers containing non-volatile salts. These salts cause ion suppression and adduct formation, which complicates molecular weight determination. Many methods have been developed for the purpose removing salts from protein. These methods include SEC, centrifugal filtration and buffer exchange, yet these techniques are limited to off-line use only. An alternative technique has been developed that can be used either off-line or in an on-line approach coupled directly to a mass spectrometer. This 2.1 x 10 mm device, packed with polymer sorbent, was evaluated using acidic, (bovine serum albumin), basic, (cytochrome c), and large globular (monoclonal IgG1) protein samples.

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