Nonradioactive Methods for Visualization of Protein Blots

The ability to detect specific proteins that have been immobilized on nitrocellulose or polyvinylidene difluoride (PVDF) membranes has become an invaluable technique in all areas of the biological sciences. In the past, visualization of low-abundance proteins required the use of radioactive markers, such as ¹²⁵ Iodine, but with the advent of enhanced chemiluminescent technology and amplified colorimetric detection, this is no longer the case (1 –3 ). This chapter details two of the more popular nonradioactive methods for immobilized protein detection, both of which rely on the indirect labeling of specific proteins with enzyme-linked antibodies. The location and abundance of these enzyme markers are then realized in a reaction producing a colored end product or by the generation of light in a chemiluminescent reaction. These techniques can identify <20 pg of protein.