See JAX Pipeline on the IMPC website for order of tasks for all mice tested

Project protocol: Insulin blood level

• Introduction
• Study design
• Workflow
• Equipment, software and supplies
• Reagents and solutions
• Procedure
• Measurements
• Experimental and technical parameters

Introduction

The insulin concentration in the blood is an important indicator of diabetes. Note: Blood collection for Insulin Blood Level is performed as a non-fasting, terminal procedure.

Study design

Paradigm: Baseline study

Panel: Collaborative Cross

Number of strains: 18

Sex: Both

Age: 17-19 wks

Sample size: Six mice per sex from at least two litters

Number of cohorts: 1

Workflow

Step	Description	Equipment	Data collected
1	Mice are anesthetized	-	-
2	Blood is collected and plasma isolated	Microtainer tubes, Eppendorf tubes, centrifuge	-
3	Insulin levels are assessed	Imager	Plasma insulin levels

Equipment, software and supplies

• Imager: Meso Scale Discovery SECTOR Imager 2400 [See Experimental and technical parameters]
• Microtainer tubes (Lithium Heparin)
• Refrigerated centrifuge
• Eppendorf tubes
• Pipettes

Reagents and solutions

• Isofluorane

 

Procedure

1. Mice are anesthetized with isofluorane.
2. Blood is collected via retro-orbital puncture.
3. Blood is collected in lithium heparin tubes and the samples are kept on ice for a maximum of 2 hours prior to isolation of the plasma.
4. Blood samples are centrifuged at 5,000 x g for 10 minutes at 8ºC and the plasma removed and aliquoted for freezing (-70ºC).
5. Plasma samples are subsequently defrosted and the required amount of sample is used to perform the analysis (2 replicates). Samples are diluted 1:2.

 

Data QC: Plasma samples must be free of Fibrin clots in order to be analyzed. Badly hemolyzed samples should not be included.

 

Measurements

1. LIMS field name	MPD variable name	Description	Units	Series	Primary measurement	Series order	MPD computed	Calculation	Method	Data type
   Insulin	insulin	insulin (plasma)	pg/mL						hormone quantification	Float

   
   

   
   

   Experimental and technical parameters

   Equipment name	Imager
   Equipment manufacturer	Meso Scale Discovery
   Equipment model	SECTOR Imager 2400
   Anesthesia	Isofluorane
   Blood collection	Retro orbital