 We have synthesized and characterized a novel photoreactive Ca2-analog reagent, Azido ruthenium AzRu that binds covalently to Ca2-binding sites in proteins. The specificity of AzRu for Ca2-binding proteins, its photoactivation and its radioactive labeling pave the way for a wide use of AzRu as a powerful tool to assess Ca2-dependent cellular processes, both in vitro and in vivo. Using radiolabeled AzRu, it will be possible to identify Ca2-binding proteins in a biological sample such as whole cell, isolated mitochondria, ER or other protein containing fractions. Protein bound 103RuAzRu in biological sample can be separated by SDS gel electrophoresis, followed by Coomassie staining and exposure of the dried gel to X-ray film autoradiography. This allows the identification of the 103Ru radiolabeled proteins. The labeled protein bands can be cut from the gel and subjected to MALADI-TOF analysis and the identity of the protein can be determined by a sequence homology search. 