Páncreas murino aislamiento Islet

Protocol

Preparación colagenasa Pesar de la colagenasa en un búfer de 50 ml y agregar aislamiento cónico para que la concentración final, como se indica en la tabla de abajo. Disolver la colagenasa por agitación. (12 + 2 ratones ratones extra = 14 ratones x 5 ml / ratón = 70 ml de solución de colagenasa total) (70 ml x 0,5 mg / ml o 0,3 mg / ml = 35 mg o 21 mg de colagenasa total P) <table style="BORDER-RIGHT: #ddd 1px soli…

Materials

Material Name	Tipo	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