Collagenase Preparation
Strain | Age | Collagenase P (mg/ml) | Time (min) |
---|---|---|---|
C3H, Balb/C, B6 | > 12 weeks | 0.8 | 17 |
C3H, Balb/C, B6 | <= 12 weeks | 0.5 | 13 |
NOD, NOR, NON | > 12 weeks | 0.8 | 17 |
NOD, NOR, NON | <= 12 weeks | 0.5 | 17 |
Distension
Digestion
Ficoll
Perform quickly. Long-term Ficoll exposure is TOXIC to islets.
Material Name | Type | Company | Catalogue Number | Comment |
---|---|---|---|---|
Dithizone | reagent | Dissolve 5mg Dithizone in 5 ml of DMSO in a 50 ml conical tube. Add 45 ml of normal saline and mix gently. Filter with a 60 cc syringe and a 0.22 “m syringe filter into a new 50 ml tube. Store at 4 OC, until use. Add 1 ml to each 35 mm counting Grid-plate, when needed. | ||
PBS | Buffer | |||
HBSS | Buffer | |||
Isolation soln. | Buffer | HBSS containing 10 mM of HEPES, 1 mM of MgCl2, 5 mM of Glucose, pH 7.4 | ||
Wash soln. | Buffer | Isolation buffer containing 1mM CaCl2 | ||
Collagenase P | Boehringer Mannheim-Roche | 1213873 | ||
Dissection tools | Hemostat clamp (n=2), Forceps (n=2), Scissors (for skin incision an for taking out pancreas from adjacent tissue) | |||
Syringe | sterile, 5ml, 1/mouse. | |||
30 G needles | sterile, 1/syringe | |||
siliconized glass vials w/Teflon cap | Fisher | 213-018-54 | sterile, 1/mouse | |
large sink strainer | sterile | |||
1000 ml Beaker | sterile | |||
Gauze pads | 4×4, 2/mouse | |||
plastic eye dropper | sterile | |||
Mice | Animal |