Primary Hepatocyte Isolation
Primary hepatocyte isolation 1. MATERIALS & SOLUTIONS REQUIRED A. Collagen I coating plate - 30% ethanol + collagen I 6well plate(450ul each), 12well plate(200ul each), dry it and PBS washing. Click to Details B . media - M199 with 10% FBS + 1% antibiotics. C. Collagenase D(Roche) Click to Details or Collagenase type 1 or 4 (Worthington) Click to Details 1X EGTA solution and Enzyme solution After stir well (about 1h) and make to x1 solution, you can dissolve CaCl2, if not, solution will precipitate. And then adjust pH between 7.35 and 7.4 by 10N NaOH. 1) Hepatocyte isolation for 1 mice EGTA Solution : 50ml/mice Enzyme Solution: 100ml + Collagenase D 50mg (Collagenase Type 1 55mg) + Collagenase P 5mg or Collagenase Type 1 only 65mg ------------dissolve by stirring in bottle 2) ...