Assessment of Two Immunodepletion Methods: Off-Target Effects and Variations in Immunodepletion Efficiency May Confound Plasma Proteomics

作者：Patel Bhavinkumar B; Barrero Carlos A; Braverman Alan; Kim Phillip D; Jones Kelly A; Chen Dian Er; Bowler Russell P; Merali Salim; Kelsen Steven G; Yeung Anthony T^*

来源：Journal of Proteome Research, 2012, 11(12): 5947-5958.

DOI：10.1021/pr300686k

• immunodepletion
• Seppro
• IgY
• Qproteome
• iTRAQ
• EMMOL normalization
• off-target

Immunodepletion of abundant plasma proteins increases the depth of proteome penetration by mass spectrometry. However, the nature and extent of immunodepletion and the effect of off-target depletion on the quantitative comparison of the residual proteins have not been critically addressed. We performed mass spectrometry label-free quantitation to determine which proteins were immunodepleted and by how much. Two immunodepletion resins were compared: Qproteome (Qiagen) which removes albumin + immunoglobulins and Seppro IgY14 + SuperMix (Sigma-Aldrich) which removes 14 target proteins plus a number of unidentified proteins. Plasma collected by P100 proteomic plasma collection tubes (BD) from 20 human subjects was when using only albumin + immunoglobulins removal (Qproteome), while lower abundance proteins were evaluated better using exhaustive immunodepletion (Seppro IgY14 + SuperMix). The latter resin removed at least 155 proteins, 38% of the plasma proteome in protein number and 94% of plasma protein in mass. The depth of immunodepletion likely accounts for the effectiveness of this resin in revealing low abundance proteins. However, the more profound immunodepletion achieved with the IgY14 + SuperMix may lead to false-positive fold-changes between comparison groups if the reproducibility and efficiency of the depletion of a given protein are not considered.

• 出版日期2012-12