MATERIALS AND METHODS

The following reagents were obtained as noted: reagent grade N-butanol, sodium phosphate, sodium chloride, disodium ethylenediaminetetraacetate (EDTA), barbital, calcium chloride dihydrate, and tris (hydroxymethy1)-aminomethane (Tris) from Fisher Chemical, Fairlawn. NJ; gelatin from Difco Laboratories, Detroit, MI; diisopropylfluorophosphate (DFP), phenylmethylsulfonyl fluoride (PMSF) from Sigma Chemical Co., St. Louis, MO; diethylaminoethyl-(DEAE) Sephacel, phenyl-Sepharose, and CH-Sepharose from Pharmacia Fine Chemicals, Piscataway, NJ: Nonidet P-40 (NP-40) from British Drug House, Poole, England; DEAE-cellulose (DE-52) from Whatman, Clifton, NJ; Bio-Gel HT (hydroxylapatite) and diethylaminoethyl Affiblue from BioRad. Richmond, CA; and trypan blue from Allied Chemical, New York, NY. Trypan blue was coupled to CH-Sepharose by 1-ethyl-3-(3-diethylaminoethyl) carbodiimide hydrochloride with 50 mg of dye/ml of packed Sepharose according to the instructions of Pharmacia. The coupled matrix was washed sequentially with water, methanol, and ethanol to remove types were obtained from the American Red Cross, Washington, DC, or uncoupled dye. Fresh, human, packed red blood cells of various serologic from normal volunteers. as the standard. Discontinuous SDS-PAGE was performed with 7% gels Protein was quantitated by Folin analysis (8) with bovine serum albumin