Actin/Tm ratios. (A) Relative staining intensity of actin and Tm in SDS-PAGE gels stained with Coomassie blue was determined. 1 µg actin from two separate preps (lanes 1–6) and 1 µg Tm (lanes 7–9) were compared as were 0.2 µg actin (lanes 10–15) and 0.2 µg Tm (lanes 16–18). The concentrations of actin and Tm were determined using their extinction coefficients. For actin, absorbance at 290 nm 0.1% (1 g/liter) was 0.63. For Tm absorbance at 280 nm 0.1% (1 g/liter) was 0.274. The average actin/Tm staining intensity ratio was 1.55. (B) Mixtures of 3.5:1 actin/Tm were made (total, first six lanes). Following centrifugation (TLA 100 rotor, 100,000 rpm, 30 min, 4°C) actin and bound (cosedimenting) Tm in the pellet were examined (pellet, lanes 7–12). Supernatant (unbound Tm) was also examined (supernatant, lanes 13–18). The relative intensities of the actin and Tm bands was 3:1 for the Total and 7.5:1 for the Pellet following correction for staining differences determined in A and differences in molecular weights (actin, 42,000 D; Tm dimer, 65,300 D). Total and Supernatant can be directly compared as the same volume was examined for each. 40% of the input Tm remained unbound indicating that there was excess Tm in the mixture. All of the actin was found in the pellet.