Low-pH association of proteins with the membranes of intact red blood cells. I. Exogenous glycophorin and the CD4 molecule.
Glycophorin and CD4 proteins are tightly associated with intact human erythrocyte membranes after a short-time incubation at low pH (1-2 min, pH lower than 5, 37 degrees C). Flow cytometry and epifluorescence microscope observations showed that after incubation of red cells with fluorescein isothiocyanate (FITC) labeled glycophorin at pH values lower than 5, the erythrocyte membrane and subsequently formed ghost membranes were fluorescent. Unlabeled glycophorin was reacted with mouse erythrocytes using the same low-pH conditions. Flow cytometry and fluorescence microscopy showed that anti-glycophorin monoclonal antibodies were able to recognize the epitopes of glycophorin associated with the mouse erythrocytes. Kinetic experiments showed that the interaction of FITC-glycophorin with red cell membranes can be monitored by a decrease in the fluorescence intensity. Erythrocyte associated glycophorin was not removed from the membranes after 24 h incubation in human plasma (in vitro, 39 degrees C). A glycoprotein extract containing CD4 was isolated from a T4-lymphoma cell line (CEM). This protein extract was incubated with erythrocytes using the same low-pH conditions. Fluorescently labeled monoclonal antibodies against CD4 stained the red cells after association of CD4 with the membranes. Electron microscopy showed 10 nm immunoglobulin G-coated gold beads associated with CD4-bearing erythrocyte membranes after incubation with anti-CD4 antibodies and then with the gold beads. The potential use of the CD4-erythrocyte complex as a therapeutical agent against acquired immune deficiency syndrome (AIDS) is suggested.
Arvinte
T
T
Biophor Corporation, Texas A & M University Research Park, College Station 77840.
Schulz
B
B
Cudd
A
A
Nicolau
C
C
D000163:Acquired Immunodeficiency Syndrome / Q000097:blood; D000818:Animals; D000945:Antigens, Differentiation, T-Lymphocyte / Q000032:analysis*; D002450:Cell Communication; D004910:Erythrocyte Membrane / Q000276:immunology / Q000502:physiology* / Q000648:ultrastructure; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate; D005452:Fluoresceins; D005454:Fluorescence Polarization; D005614:Freeze Fracturing; D006021:Glycophorins / Q000276:immunology / Q000502:physiology* / Q000648:ultrastructure; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008565:Membrane Proteins / Q000097:blood* / Q000502:physiology / Q000648:ultrastructure; D051379:Mice; D008855:Microscopy, Electron, Scanning; D017830:Octoxynol; D011092:Polyethylene Glycols; D012795:Sialoglycoproteins / Q000502:physiology*; D013696:Temperature; D013861:Thiocyanates
D000945:Antigens, Differentiation, T-Lymphocyte; D005452:Fluoresceins; D006021:Glycophorins; D008565:Membrane Proteins; D012795:Sialoglycoproteins; D013861:Thiocyanates; D011092:Polyethylene Glycols; D017830:Octoxynol; D016650:Fluorescein-5-isothiocyanate