Non-radioactive detection of proteins synthesized in vitro. Using these systems, biotinylated lysine residues are incorporated into nascent proteins during translation, eliminating the need for labeling with [35S]methionine or other radioactive amino acids. Biotinylated lysine is added to the translation reaction as a precharged ε-labeled biotinylated lysine-tRNA complex (Transcend™ tRNA) rather than as a free amino acid. After SDS-PAGE and electroblotting, biotinylated proteins can be visualized by binding either Streptavidin-Alkaline Phosphatase (Streptavidin-AP) or Streptavidin-Horseradish Peroxidase (Streptavidin-HRP), followed either by colorimetric or chemiluminescent detection. Typically, these systems enable you to detect 0.5–5ng of protein in 3–4 hours after gel electrophoresis. This sensitivity is equivalent to that achieved with [35S]methionine incorporation and autoradiographic detection 6–12 hours after gel electrophoresis.
For more information, see the Protocols & Applications Guide.