Materials and Reagents

1. 6-8 weeks old mouse
   
2. PBS without Ca²⁺ and Mg²⁺ (Wisent, catalog number: 311-01-CL )
   
3. Dulbecco's Modified Eagle's Medium High glucose with stable L-glutamine (DMEM) (Wisent, catalog number: 319-015-CL )
   
4. Fetal bovine serum (FBS) (Life Technologies, Gibco^®, catalog number: 12483 )
   
5. Penicillin/Streptomycin solution (Wisent, catalog number: 450-201-EL )
   
6. Trypan blue (Life Technologies, Gibco^®, catalog number: 15250-061 )
   
7. Trypsin 0.05%/EDTA 0.53 mM (Wisent, catalog number: 325-042-EL )
   
8. APC-conjugated anti-CD31 antibody (clone MEC13.3) (BD Biosciences, catalog number: 551262 )
   
9. FITC-conjugated anti-CD45 antibody (clone 30-F11) (BD Biosciences, catalog number: 553080 )
   
10. APC-conjugated anti-CD44 antibody (clone IM7) (BD Biosciences, catalog number: 559250 )
    
11. PE-conjugated anti-CD105 antibody (clone MJ7/18) (BD Biosciences, catalog number: 562759 )
    
12. FITC-conjugated anti-CD90 antibody (clone 5E10) (BD Biosciences, catalog number: 555595 )
    

Equipment

1. Scissors and forceps
   
2. Syringe 1cc with 27 Gauge x 1-½ needle (BD, catalog numbers: BD-309659 and BD-305109 )
   
3. Petri dishes 100 x 20 mm (BD, catalog number: DL-353003 )
   
4. 50 ml conical tubes (Progene^®, catalog number: 71-5000-B )
   
5. Table top centrifuge
   
6. Culture hood
   
7. Hemocytometer
   
8. T-25 flask (BD, catalog number: 353108 )
   
9. 37 °C, 5% CO₂ Cell culture incubator
   
10. Flow cytometer (e.g. BD LSRFortessa)
    

Procedure