The Worthington Papain Dissociation System is a set of reagents intended for use in the neural cell isolation method of Huettner and Baughman (J. Neurosci., 6, 3044, 1986). The materials are designed for convenience and simplicity and are useful to the occasional user as well as the more experienced and frequent user. Each lot is use tested for performance in rat spinal neural cell isolation and this kit provides freshly prepared enzyme solutions for each dissociation.
Stability/Storage: The reagents are stable at ambient temperatures for the periods of time expected in normal shipping procedures, but the package should be refrigerated upon arrival. Contents may be stored at 2-8°C for 4-6 months before use. Store at 2-8°C.
• Vial 1: Sterile Earle's Balanced Salt Solution (EBSS) with calcium, magnesium, bicarbonate and phenol red, one vial per package, 100ml. Aliquots of this vial are used to reconstitute other vials and to prepare dilute inhibitor solution. Refrigerate between uses and equilibrate with sterile O2:CO2 before each use.
• Vial 2: Papain containing L-cysteine and EDTA, 5 x 100 unit single-use vials per package. The material is 0.22 micron membrane filtered and lyophilized in autoclaved vials. A vial reconstituted with 5ml of EBSS (Vial 1) yields a solution at 20 units of papain per ml in 1mM L-cysteine with 0.5mM EDTA. Brief incubation at 37°C is needed to insure full solubility and activity.
• Vial 3: Deoxyribonuclease I (DNase), 5 x 2000 unit single use vials per package.
• Vial 4: Ovomucoid protease inhibitor with bovine serum albumin, one vial per package, 32ml upon reconstitution.
Also included is a card correlating color with pH for use as a guide in O2:CO2 equilibration.