- Ringer solution: 120 mM NaCl, 25 mM NaHCO 3 , 3.3 mM
KH 2 PO 4 , 0.8 mM K 2 HPO 4 , 1.2 mM MgCl 2 , 1.2 mM CaCl 2 ,
10 mMD-glucose, pH 7.4, equilibrated with 5% CO 2 (seeNote
5 ). - Alternatively, normal Hank’s Balanced Salt Solution can be
used: 110 mM NaCl, 5 mM KCl, 1.2 mM MgSO 4 , 1.8 mM
CaCl 2 , 4 mM NaAcetate, 3 mM Na 3 Citrate, 6 mM D-glucose,
6mML-alanine, 1 mM NaH 2 PO 4 , 3 mM Na 2 HPO 4 ,25mM
NaHCO 3 , pH 7.4 equilibrated with 5% CO 2 (seeNote 6). - Mouse vs. rat surgery tools such as hemostats, forceps, scissors.
- 8G needle (21G for smaller animals such as mice).
- Paper laboratory tissues/wipes (dry) and/or cotton
applicators. - Clean disposable razor blades or scalpels.
- Clean disposable plastic petri dishes.
- Stir bars and stir plate.
- Transfer pipets.
- Sterile water in squeeze bottle and two waste buckets.
- P1000 pipettor and tips.
- Perfusion setup with a pump, preferably peristaltic (alterna-
tively via gravity;seeNote 7). - CO 2 impermeant tubing (seeNote 8).
- Manifolds.
- Vacuum setup (with safety trap to capture liquids from the
perfusion procedure). - Laboratory tape or labeling adhesive tape.
- Several timers with multiple channels.
- Flowchart summarizing experimental conditions, additives,
times for addition of reagents, etc. - Water-resistant markers.
- Parafilm or plastic wrap.
- Stadie-Riggs tissue slicer.
- Stadie-Riggs tissue slicer blade handle.
- Stadie-Riggs tissue slicer disposable blades (for slices of approx-
imately 0.5 mm in thickness). Use a new clean disposable blade
for each separate experiment (Thomas Scientific). - Hemostat for handling of the blade as it is placed and removed
from the handle. - Blunt forceps for handling the slices from the tissue slicer.
- Plastic disposable transfer pipettes to dispense Ringer buffer
intermittently during slicing.
452 Renee Rao et al.