Avoiding Nonspecific Interactions in Studies of the Plasma Proteome: Practical Solutions to Prevention of Nonspecific Interactions for Label-Free Detection of Low-Abundance Plasma Proteins

摘要

The molecular constitution of blood can be highly representative of the physiological state of anindividual and offers an ideal target for studies of biomarkers. High-abundance plasma proteins,particularly albumin, dominate the plasma proteome, but it is the low-abundance proteins (such ascytokines) that are commonly associated with many pathophysiological states. Several detectionstrategies, and particularly those that involve label-free detection, are available for low-abundanceprotein detection in plasma, but all can be severely compromised by the high-abundance of serumalbumin. In the present study, we examine the effect of albumin interference on accurate label-freedetection by protein microarrays. Albumin was found to disrupt specific antigen-antibody bindinginteractions of low-abundance proteins. In clinical analysis, where it is imperative to preserve theintegrity of samples, depletion of albumin may further undermine quantitative measurements. We haveoptimized procedures that permit accurate analysis to be undertaken without the need for prior treatmentof samples. The emphasis is placed on disrupting nonspecific interactions including both electrostatic(i.e., Colulombic) and electrodynamic (hydrophobic and other nonpolar based) interactions. Theseprotocols appear to be generic with potential applications in several areas of analytical biotechnology.