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Protocol for in vivo and ex vivo assessments of glucose-stimulated insulin secretion in mouse islet β cells.

STAR protocols (2021-08-20)
Yun-Xia Zhu, Yun-Cai Zhou, Yan Zhang, Peng Sun, Xiao-Ai Chang, Xiao Han
ABSTRACT

Pancreatic islet β cells secrete insulin in a biphasic manner when sensing high blood glucose level. This protocol describes the evaluation of different phases of insulin secretion, as well as basal, glucose-stimulated and total insulin secretion abilities, thereby enabling precise assessment of β cell function both in vivo and ex vivo. The in vivo assay consists of intravenous tube imbedding surgery and hyperglycemic clamp. The ex vivo assay consists of islet isolation, dynamic perfusion and static immersion. For complete details on the use and execution of this protocol, please refer to Sun et al. (2021).

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Dithizone, Practical Grade
Sigma-Aldrich
Calcium chloride, anhydrous, BioReagent, suitable for insect cell culture, suitable for plant cell culture, ≥96.0%
Sigma-Aldrich
Magnesium sulfate, anhydrous, ReagentPlus®, ≥99.5%
Sigma-Aldrich
Ammonium hydroxide solution, ACS reagent, 28.0-30.0% NH3 basis
Sigma-Aldrich
Magnesium chloride hexahydrate, ACS reagent, 99.0-102.0%
Sigma-Aldrich
Collagenase from Clostridium histolyticum, release of physiologically active rat pancreatic islets tested, Type V, ≥1 FALGPA units/mg solid, >125 CDU/mg solid
Sigma-Aldrich
α-D-Glucose, anhydrous, 96%
Sigma-Aldrich
Histopaque®-1077, sterile-filtered, density: 1.077 g/mL
Sigma-Aldrich
Sodium chloride, ACS reagent, ≥99.0%
Sigma-Aldrich
Potassium chloride, ACS reagent, 99.0-100.5%