The QUEST system for quantitative analysis of two-dimensional gels

Garrels, J. I. (March 1989) The QUEST system for quantitative analysis of two-dimensional gels. J Biol Chem, 264 (9). pp. 5269-82. ISSN 0021-9258

URL: http://www.ncbi.nlm.nih.gov/pubmed/2925692

Abstract

The strategies and methods used by the QUEST system for two-dimensional gel analysis are described, and the performance of the system is evaluated. Radiolabeled proteins, resolved on two-dimensional gels and detected using calibrated exposures to film, are quantified in units of disintegrations per minute or as a fraction of the total protein radioactivity applied to the gel. Spot quantitation and resolution of overlapping spots is performed by two-dimensional gaussian fitting. Pattern matching is carried out for groups of gels called matchsets, and within each matchset every gel is matched to every other gel. During the matching process, spots are automatically added to each pattern at positions where unmatched spots were detected in other patterns. This results in enhanced accuracy for both spot detection and for matching. The spot fitting procedure is repeated after matching. Tests show that up to 97% of spots in each pattern can be matched and that fewer than 1% of the spots are matched inconsistently. Approximately 2000 proteins are detected from typical gels. Of these 1600 are high quality spots. Tests to measure the coefficient of variation of spot quantitation versus spot quality show that the average coefficient of variation for high quality spots is 21%. The intensities of the detected proteins range from 4 to 20,000 ppm of total protein synthesis. The QUEST analysis system has been used to build a quantitative database for the proteins of normal and transformed REF52 cells, as presented in the accompanying reports (Garrels, J., and Franza, B. R., Jr. (1989) J. Biol. Chem. 264, 5283-5298, 5299-5312).

Item Type: Paper
Uncontrolled Keywords: Algorithms Animals Electrophoresis, Gel, Two-Dimensional/instrumentation/*methods/standards Image Processing, Computer-Assisted Information Systems/instrumentation/organization & administration/standards Pattern Recognition, Automated Proteins/*isolation & purification/standards Rats Reproducibility of Results Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. Software/instrumentation/*methods/standards
Subjects: bioinformatics > genomics and proteomics > databases > databases
Investigative techniques and equipment > assays > gel electrophoresis
CSHL Authors:
Communities: CSHL labs
Depositing User: Gail Sherman
Date: 25 March 1989
Date Deposited: 28 Jul 2017 16:55
Last Modified: 28 Jul 2017 16:55
Related URLs:
URI: http://repository.cshl.edu/id/eprint/34850

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