Materials and Reagents

1. 50 ml centrifuge tubes
   
2. 70 µm cell strainer (BD Biosciences, Falcon^®, catalog number: 352350 )
   
3. Male mouse (e.g., C57BL/6J) 8-20 weeks old
   Note: Generally, male mice are more severely affected by type 2 diabetes than female mice, and they are used exclusively in diet-induced diabetes studies (www.jax.org).
   
4. Collagenase from Clostridium histolyticum type IV (Sigma-Aldrich)
   
5. Fetal Bovine Serum (FBS) (Pan biotech GmbH)
   
6. Trypan blue
   
7. RPMI 1640 (with L-glutamine; 25 mM Hepes; 2.2 g/L NaHCO₃) (Pan Biotech GmbH)
   
8. MilliQ water
   
9. 0.83% NH₄Cl
   
10. 0.168% Na₂CO₃
    
11. 1 mM EDTA (pH 7.3)
    
12. 1x PBS (pH 7.3)
    
13. 0.2% BSA
    
14. 3% RPMI 1640 (see Recipes)
    
15. 3% RPMI + 1 mg/ml Collagenase D (or IV) (see Recipes)
    
16. Erylysis buffer (see Recipes)
    
17. FACS wash buffer (see Recipes)
    

Equipment

1. Soft wood tablet and pins
   
2. Disinfectant
   
3. Small thin surgical scissors and tweezers
   
4. Thermostatic shaker
   
5. Vortex
   
6. Vacuum pump
   
7. Centrifuge
   

Procedure