Collecting Data to Determine the Ca²⁺-Binding Properties of DM-Nitrophen and Proteins

Abstract

Conventional techniques used to measure Ca²⁺ binding are too slow to determine accurately the fast binding kinetics of most molecules such as Ca²⁺-binding proteins (CBPs). We have developed an ultra-fast in vitro technique for measuring the Ca²⁺-binding properties of CBPs following flash photolysis of caged Ca²⁺. Although the details of the setup, the mathematics, and the analysis involved in this technique have been published elsewhere, many of the practical details regarding the actual measurements have, until now, only been described minimally. Here, we present a protocol to gather the data necessary to determine the kinetic properties of a caged-Ca²⁺ compound and a CBP.