Using Label-Free Quantification to Assess Sample Preparation: Application to Protein Expression Dynamics of Marek's Disease Viral Infection

摘要

Predicated on absolute quantification, the SDS-based lysis buffer improves protein extraction (based on protein mass) by approximately 30percent over the NP-40-based buffer. When considering the number of proteins identified, the SDS-based lysis buffer is approximately 20percent more efficient than the NP-40-based buffer in extracting/solubilizing CEF proteins for trypsin digestion. Based on absolute quantification, acetone precipitation reduces protein recovery by approximately 60percent and reduces the number of proteins identified by about 35percent. The SDS-no precipitation method is the best of four methods used to analyze the CEF proteome using LC/MSE and is amenable to downstream sample preparation to study MDV transformed cell lines. Absolute quantification by LC/MSE provides another figure of merit for quantitative evaluation of sample preparation strategies.