Three separate preparations of actin and C0–C2 were used (1, 2, and 3) to run mock plate reader screens for calculating Z′ score using Eq. 2 (see Materials and methods) based on mean lifetime (nanoseconds) and SD. For binding experiments using IAEDANS–actin, 2.5 µM C0–C2 was added. 1.25 µM C0–C2 was used for IAEDANS–actin–Tm. Binding was done with (+) or without (−) PKA treatment to phosphorylate cMyBP-C. Within each preparation, actin alone was compared to actin with bound C0–C2 for one comparison (actin ± C0–C2; first row versus second row for each preparation). Actin bound to unphosphorylated versus phosphorylated C0–C2 was also compared (actin C0–C2 ± PKA; second row versus third row for each preparation). See Fig. 2, C–E, for a representative example of the mock screen.