Computer analysis of double-labeled two-dimensional electrophoresis gels.
A computerized process for the automatic analysis of double-label autoradiography after two-dimensional polyacrylamide gel electrophoresis has been developed. Matching fluorographs and autoradiographs produced from gels containing 3H- and 14C-labeled proteins are digitized by a rotating drum densitometer and analyzed by the Man-computer Interactive Data Analysis System III. This system locates corresponding protein spots in the films with edge-detection algorithms, converts spot density readings to isotopic disintegrations by reference to standard curves, and computes a 3H:14C ratio for each spot in the gels. On the average, calculated ratios are accurate to approximately 9% for test strips of polyacrylamide gel containing uniform mixtures of 3H and 14C. Values obtained for two-dimensional gels containing n protein spots with a known 3H:14C ratio of 8.6 +/- 0.1 are as follows: 8.1 +/- 1.4 (n = 268), 8.8 +/- 2.1 (n = 278), 9.1 +/- 1.7 (n = 245), and 8.8 +/- 2.2 (n = 223). The computer process greatly reduces the time required to precisely compare two complex protein mixtures and has sufficient precision to detect a doubling in the biosynthesis of any individual protein.
D000818:Animals; D001331:Automation; D001345:Autoradiography / Q000295:instrumentation*; D002250:Carbon Radioisotopes / Q000032:analysis; D002642:Chick Embryo; D003201:Computers; D004591:Electrophoresis, Polyacrylamide Gel / Q000295:instrumentation*; D014176:Protein Biosynthesis; D011506:Proteins / Q000032:analysis*; D014316:Tritium / Q000032:analysis; D014962:X-Ray Film
D002250:Carbon Radioisotopes; D011506:Proteins; D014316:Tritium