A novel negative imaging technique for accurate localization of stainable proteins on complex two-dimensional autoradiograms.
This paper describes a fast, simple, and accurate method for localization of protein antigens in complex two-dimensional (2-D) autoradiograms where the precise position and identity of the protein is required. The method involves the creation of negative images on an autoradiogram through arrangement of image-intensifying screens. By placing a chromogenically stained 2-D gel or blot between the intensifying screen and the film, photons emitted from the intensifying screen are obstructed by the stained spots, thus creating a negative image on the film. The technique can be used in autoradiograms of proteins labeled with either 32P or (125)I radioisotopes. The technique permits analysis of radiolabeled, gold-stained and immunoreacted proteins on a single film and offers versatility by combining analysis of total protein patterns with specific identification of radiolabeled and/or immunoreacted protein spots. The technique is especially useful when selecting a subset of specifically radiolabeled proteins from the total protein pattern in 2-D gels or membranes for microsequencing.
Naaby-Hansen
S
S
Department of Cell Biology, Center for Recombinant Gamete Contraceptive Vaccinogens, University of Virginia, Charlottesville 22908, USA.
Roof
R W
RW
Flickinger
C J
CJ
Grafer
C M
CM
Parsons
S J
SJ
Herr
J C
JC
D000818:Animals; D001345:Autoradiography / Q000379:methods*; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D005347:Fibroblasts / Q000737:chemistry; D006801:Humans; D015151:Immunoblotting; D007457:Iodine Radioisotopes; D007525:Isoelectric Focusing; D008297:Male; D051379:Mice; D010750:Phosphoproteins / Q000032:analysis; D010761:Phosphorus Radioisotopes; D011506:Proteins / Q000032:analysis*; D013094:Spermatozoa / Q000737:chemistry; D014404:Tubulin / Q000032:analysis
D007457:Iodine Radioisotopes; D010750:Phosphoproteins; D010761:Phosphorus Radioisotopes; D011506:Proteins; D014404:Tubulin