Evaluating the performance of new approaches to spot quantification and differential expression in 2-dimensional gel electrophoresis studies

Jeffrey S. Morris, The University of Texas M.D. Anderson Cancer Center
Brittan N. Clark, The University of Texas M. D. Anderson Cancer Center
Wei Wei, The University of Texas M. D. Anderson Cancer Center
Howard B. Gutstein, The University of Texas M. D. Anderson Cancer Center

Abstract

2-DE is an important method for proteomics. Accurate spot detection and quantification on the resulting images is a challenging task, but must be done effectively for the technology to fulfill its potential. Traditional analytical methods have significant weaknesses, including spot mismatching and missing data. These problems require time-consuming manual editing to correct, which dramatically decreases throughput and compromises the objectivity and reproducibility of the analysis. To address this issue, we developed Pinnacle, a new method that markedly improves spot detection and quantification precision. Another new method implemented in Progenesis SameSpots, a commercial 2-DE analysis product, has also been touted as an improvement upon traditional approaches. In this study, we compared Pinnacle and SameSpots in spot detection and precision of quantification using two different dilution series, and evaluated the detection of differentially expressed proteins in two differential expression experiments. We found that SameSpots at times had problems with spot delineation, while Pinnacle did not. While both Pinnacle and SameSpots showed marked improvement in precision of spot quantifications over conventional methods, Pinnacle yielded spot quantifications with greater validity and reliability than SameSpots. Pinnacle detected more differentially expressed proteins than SameSpots, which may be a result of its increased precision and improved spot delineation.