Liberase dh enzyme

Day 3 ovaries were harvested and dissected into 18 to 20 pieces using 27-gauge needles. The ovary pieces were digested using Liberase DH enzyme (Roche Diagnostics Australia Pty Limited, North Ryde, NSW, Australia) at 50 ug/ml for 75 min at 37 °C with gentle agitation. The enzyme reaction was stopped using foetal calf serum (final concentration 5%). Primordial follicles were isolated using stripper tips and transferred to vials and snap frozen in liquid nitrogen and stored at −80 °C prior to RNA extraction.

Adult rat cardiomyocytes were isolated after heart perfusion with collagenase type II (1 mg/ml, Worthington), then purified and cultured, as described previously46 (link). Primary cultures of cardiomyocytes were incubated with different sugar concentrations for the indicated periods of time, as detailed in the figure legends.
Adult mice were anesthetized with a mixture of ketamine (80 mg/kg) and xylazine (10 mg/kg). Cardiomyocytes were isolated, as described34 (link), with the following modifications: hearts were perfused with Liberase DH enzyme (0.625 mg/heart, Roche) and trypsin (2.1 mg/heart, Life Technologies 15090–046). The cells were then purified, cultured and plated on laminin-coated dishes, and incubated for 1 h in fresh minimum essential medium (MEM) with Hank’s salts (Life Technologies 11575–032) supplemented with L-glutamine (2 mM), bovine serum albumin (BSA) 100 μg/ml, penicillin (100 U/ml) and streptomycin (100 μg/ml). Primary cultures of mice cardiomyocytes were treated as adult rat cardiomyocytes.