Isothermal titration microcalorimetry. Titration microcalorimetry results for binding of Me-α-Man (3.804 mM) to S23A (295 μM) in 0.1 M Tris buffer (pH = 7.5) with 30 μM CaCl2. A) Data obtained from 29 automatic injections (10 μL) of Me-α-Man each into the S23A-containing cell. B)Plot of the total heat released as a function of ligand/protein molar ratio for the titration shown in panel A. The solid represents the best least-squares fit for the obtained data.