The Preparation of γ-Globulin from Placental Blood by Ethanol Fractionation¹

Abstract

A procedure is described for the separation and purification of 7-globulin from human placental extracts and postpartum blood. Frozen placentas together with the postpartum blood are thawed, chopped and extracted with isotonic saline. The initial separation of a fraction containing 7-globulin is accomplished by the addition of ethanol in the cold. A large part of the -globulins are separated from the 7-globulin fraction by the precipitation of the former at pH 4.8, and ethanol 8%. This is followed by the precipitation of a 7-globulin-rich (70%) fraction at pH 7.0-7.2 and ethanol 25%. A fraction rich (91%) in a-and /3-globulins is separated at pH 5.1 and ethanol 17%, leaving a supernatant fluid from which the 7-globulin is again precipitated with a final purity of 96%.

IgG Subclass Composition and Immunochemical Characteristics of Plasmin-Treated Human γ-Globulin

“…It was a product of the Central Laboratory of the Swiss Red Cross, Blood Transfusion Service, Berne (Switzerland). (2) Three preparations of placental origin (VG-1, VG-2, VG-3) were produced in the Insti tut Mérieux, Lyon (France) by the procedure of Taylor et al [32], (3) Anti tetanus and anti-smallpox /-globulins (GG-Tet, GG-Vac) fractionated from hy perimmune donor plasma were outdated commercial lots of the Swiss Institute of Sera and Vaccines, Berne. (4) /-Globulin with high anti-streptolysin 0 activity (GG-Stn) was isolated by ammonium sulphate precipitation of the serum at 40% saturation followed by purification on a Sephadex G-200 column.…”

The Purification of Human Placental Plasminogen (Profibrinolysin)

“…Human placental fractions were obtained from normal human placentas and intrapartum blood by the cold ethanol techniques [9,10] developed in this laboratory for the routine pre paration of serum albumin and immunoglobulin G.…”

“…Freezing Precipitate II at -20°C caused the formation of a relatively Ppt. II [9,10] insoluble complex between plasminogen and other proteins and thus made assay and purification difficult.…”

The Fractionation of Heparinized Blood

“…(a) method 6 of Cohn, et al [4] ; (b) method 9 of Oncley, et al [12], and (c) zinc-ethanol procedures developed in this laboratory for frac tionation of placental extracts [7,17,18].…”

“…In a further experiment, a pool of heparinized plasma was processed using the zinc-ethanol system [7,17,18] developed and currently in use at the Michigan Department of Health for the preparation of both gamma globulin and albumin from placental extracts and intrapartum blood. By this method, the gamma globulin and albumin were both recovered in highly purified form (98.4 and 99 %, respectively, by electrophoretic analysis).…”