Isolation and Growth of Adult Mouse Dorsal Root Ganglia Neurons

Materials and Reagents

1. 6 to 9 weeks old female mouse
   
2. Poly-L-lysine hydrobromide (100 μg/ml in water) (Sigma-Aldrich, catalog number: P1524 )
   
3. Laminin (1 mg/ml) (Thermo Fisher Scientific, Invitrogen^TM, catalog number: 23017-015 )
   
4. Collagenase from Clostridium histolyticum (10 mg/ml in Ca²⁺, Mg²⁺ free PBS) (Sigma-Aldrich, catalog number: C-9891 )
   
5. Dispase II (10 mg/ml in Ca²⁺, Mg²⁺ free PBS) (Roche Diagnostics, catalog number: 04-942-078-001 )
   
6. Deoxyribonuclease I from bovine pancreas (4 mg/ml in PBS) (Sigma-Aldrich, catalog number: DN25 )
   
7. Leibovitz's L-15 Medium (Thermo Fisher Scientific, Gibco^TM, catalog number: 11415-064 )
   
8. Neurobasal^®-A Medium (Thermo Fisher Scientific, Gibco^TM, catalog number: 10888-022 )
   
9. B-27^® supplement (50X), serum free (Thermo Fisher Scientific, Invitrogen^TM, catalog number: 17504-044 )
   
10. Percoll^® (Sigma-Aldrich, catalog number: P1644 )
    
11. Hank’s Balanced Salt Solution (HBSS, calcium, magnesium, no phenol red) (Thermo Fisher Scientific, Gibco^TM, catalog number: 1425092 )
    
12. 15 mm Glass coverslips (Neuvitro, catalog number: GG-15 )
    
13. Pasteur pipette
14. Falcon^® 35 mm Not TC-Treated Easy-Grip Style Bacteriological Petri Dish, 20/Pack, 500/Case, Sterile (Corning Incorporated, Falcon^®, catalog number: 351008 )
15. Falcon^® 60 mm x 15 mm Not TC-Treated Treated Bacteriological Petri Dish, 20/Pack, 500/Case (Corning Incorporated, Falcon^®, catalog number: 351007 )
16. Falcon^® 14 ml Round Bottom High Clarity PP Test Tube, Graduated, with Snap Cap, Sterile, 25/Pack, 500/Case (Corning Incorporated, Falcon^®, catalog number: 352059 )
17. Falcon^® 14 ml Round Bottom PP Test Tube, without Cap, Sterile, 125/Pack, 1000/Case (Corning Incorporated, Falcon^®, catalog number: 352018 )
    

Equipment

1. #10 scalpel blade (Fine Science Tool, catalog number: 10010-00 ) (Figure 1)
   
2. Standard scalpel handle #3 (Fine Science Tool, catalog number: 10003-12 ) (Figure 1)
   
3. Spring Scissors-Angled to Side (Fine Science Tool, catalog number: 15006-09 ) (Figure 1)
   
4. Small curved scissors (Fine Science Tool, catalog number: 14095-11 ) (Figure 1)
   
5. Standard Pattern Forceps (Fine Science Tool, catalog number: 11000-12 ) (Figure 1)
   
6. Dumont #5 forceps (Fine Science Tool, catalog number: 11252-20 ) (Figure 1)
   
7. Dissecting microscope
   
8. Swinging bucket rotor (Sorvall instrument, model: HB-4 )
   
9. Centrifuge (Sorvall instrument, model: RC5 plus )
   
   
   Figure 1. The dissection tools for mouse dissection and DRG isolation. A. Standard scalpel handle #3; B. Spring scissors; C. Small curved scissors; D. Standard forceps; E. Dumont #5 forceps.
   

Procedure