The separation of rat T lymphocytes was investigated on anti-IgIg columns. A simple and efficient method for the purification of rat Ig by precipitation of rat serum with sodium sulfate is presented. Protein binding characteristics of glass and plastic beads, as solid support of affinity columns, are described, as well as optimal parameters for coating beads with rat Ig (with BSA, ribonuclease and lysozyme, as comparison). Binding of Ig was primarily dependent on the concentration of the Ig solution. Maximal strong binding of Ig (6.2 × 103 molecules per μ2 of bead surface) was reached at 400 μg per ml concentration of purified Ig solution during 20 min of incubation. Higher concentrations increased only the amount of loosely bound Ig on the surface of beads whereas the amount of firmly bound Ig remained unchanged. Fractionation of lymphoid cell suspensions on anti-IgIg affinity columns prepared at optimal conditions resulted in highly purified T-cell suspensions containing less than 1% of lymphocytes bearing surface Ig receptors.
ASJC Scopus subject areas
- Immunology and Allergy