6. Eye protection.
7. Cryogloves (face mask or goggles).
8. Double impulse heat sealer (American International Electric,
Inc.).
9. Surgery tools (scissors, scalpels, forceps, etc.).
10. Liquid nitrogen.
11. Liquid nitrogen dewar.
12. Conical centrifuge tubes, 50 mL, sterile.
13. Conical centrifuge tubes, 20 mL, sterile.
14. Cryovials (Nalgene).
15. Cryostraws, 1/2 cc (TS Scientific).
16. Falcon 40-μm cell strainers (Becton Dickinson).
17. MidiMACS separator (Miltenyi Biotec).
18. MACS LD columns (Miltenyi Biotec).
19. MACS anti-FITC microbeads (Miltenyi Biotec).
2.2 Reagents
and Media
1. Hank’s Balanced Salt Solution containing 1.26 mM calcium
and 0.90 mM magnesium without phenol red (HBSS).
2. Dulbecco’s calcium- and magnesium-free phosphate-buffered
saline (DPBS).
3. Versene (Gibco).
4. Recombinant trypsin EDTA solution (Biological Industries).
5. Defined trypsin inhibitor.
6. Collagenase A type I (see Note 1).
7. Ficoll-Paque Premium 1.073 (GE Healthcare).
8. 100Â antibiotic-antimycotic mix (Gibco).
9. 100Â glutamax (Gibco).
10. 100Â non-essential amino acids (NEAA; Gibco).
11. 50Â essential amino acids (EAA; Gibco).
12. Ethylenediaminetetraacetic acid tetrasodium salt (EDTA).
13. Ethylene glycol tetraaceticacid (EGTA).
14. Dimethyl sulfoxide (DMSO).
15. Ethylene glycol (EG).
16. Trehalose.
17. Ficoll.
18. Polyvinyl alcohol (PVA).
19. Glutathione.
20. L-Ascorbic acid 2-phosphate magnesium salt.
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Melany Lo ´ pez and Ali Eroglu