A novel negative imaging technique for accurate localization of stainable proteins on complex two-dimensional...

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.