Fig. 1 a, Shift in calmodulin-associated CnA isoforms in the failing human left ventricle. Protein extracts were immunoprecipitated with excess calmodulin antibody and calcineurin protein levels were analyzed by sequential western blot analysis of the same immunoprecipitated samples, using antibodies against the CnA C-terminal (Autoinhibitory) or N-terminal (Catalytic) domain. Bottom, Quantification by densitometric scanning was normalized to protein levels determined by Ponceau S staining. Calmodulin levels reflected protein levels in these samples as confirmed by separate western blot analysis on a higher percentage gel (data not shown) b, Full-length CnA protein is decreased in the failing human left ventricle. Protein extracts were analyzed for CnA protein by western blot analysis using an antibody specific for the C-terminal. Equal protein loading was confirmed by Ponceau S staining and by GAPDH immunoreactivity (middle); quantification by densitometric scanning (bottom) was normalized to GAPDH. Lane 1, Patient 1 shows normal CnA levels, perhaps related to an unusually prolonged survival with heart failure (15 years). c, CnA-β transcripts are decreased in the failing human left ventricle. Total RNA was size-fractionated on an agarose gel. Equal loading of samples was confirmed by ethidium bromide staining. a b c