The behavior of the solubilized receptor for immunoglobulin E in polyethylene glycol-detergent solutions: characterization and potential applications.
We studied the effect of polyethylene glycol (PEG) on the solubility of the receptor for immunoglobulin E (IgE) in non-ionic detergent extracts of rat basophilic leukemia (RBL) cells. We found that the precipitation patterns of free and IgE-bound receptor were identical but differed from that of unbound IgE. Thus, 85 to 95% of the free receptor and the IgE-receptor complexes precipitated at 13% PEG in the presence of 0.5% Nonidet P-40, whereas 95% of the unbound IgE remained soluble. A similar degree of differentiation between the precipitation of receptor-bound and unbound IgE was found when we used extracts and PEG solutions prepared with several non-ionic and/or neutral detergents. The intact IgE-receptor complex with the full complement of subunits (alpha, beta, gamma) precipitated more efficiently than the IgE-alpha-chain-complex. The presence of phospholipids, which were previously shown to be important for preservation of the association between the receptor subunits, enhanced the efficiency of precipitation of the IgE-receptor complex. The presence of PEG also had an effect on the solubility of cellular phospholipids and some of the detergents, although the effect of PEG on either could not be directly related to its effect on the solubility of the IgE-receptor complex. The radioiodinated receptor for IgE, much like other radioiodinated RBL cell membrane proteins, was soluble (greater than or equal to 95%) at approximately 7% PEG but could be specifically and efficiently precipitated from crude cell extracts, in the presence of 7% PEG upon the addition of anti-receptor immunoglobulins alone. Using mouse anti-dinitrophenyl IgE antibody, we found that unlike unbound antigen (DNP-BGG) or the IgE-receptor complex, the detergent-solubilized DNP-BGG-IgE-receptor complex was insoluble at 7% PEG. Consequently, PEG can be employed in assays to quantitate the soluble receptor, and to immunoprecipitate it specifically and directly. Moreover, the use of PEG can facilitate the distinction between unbound antigen and antigen-IgE-receptor complex as well as between the latter and IgE-receptor complex.
D000818:Animals; D001491:Basophils / Q000378:metabolism; D011232:Chemical Precipitation; D003902:Detergents / Q000494:pharmacology*; D007073:Immunoglobulin E / Q000378:metabolism*; D007942:Leukemia, Experimental / Q000276:immunology / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D011092:Polyethylene Glycols / Q000494:pharmacology*; D051381:Rats; D017455:Receptors, IgE; D011971:Receptors, Immunologic / Q000032:analysis / Q000187:drug effects / Q000378:metabolism*; D012995:Solubility; D012996:Solutions; D013501:Surface-Active Agents / Q000494:pharmacology*; D005719:gamma-Globulins / Q000032:analysis
D003902:Detergents; D008565:Membrane Proteins; D017455:Receptors, IgE; D011971:Receptors, Immunologic; D012996:Solutions; D013501:Surface-Active Agents; C030730:dinitrophenyl gamma globulin; D005719:gamma-Globulins; D007073:Immunoglobulin E; D011092:Polyethylene Glycols