An improved method of loading pH-sensitive liposomes with soluble proteins for class I restricted antigen presentation.
We have recently shown that ovalbumin (OVA) entrapped in pH-sensitive liposomes could sensitize mouse thymoma cells for lysis by MHC class I-restricted cytotoxic T lymphocytes (CTL) (Reddy et al. (1991) J. Immunol. Methods, 141, 157-163). The present studies were designed to optimize the antigen delivery system. A simple freeze-thaw method was developed to load OVA into pH-sensitive liposomes, and the protocol was optimized in terms of the choice of buffer, pH and ionic strength of the medium, lipid composition, lipid and OVA concentrations and the number of freeze-thaw cycles. Under optimized conditions, approximately 25% of OVA could be entrapped in pH-sensitive liposomes at 172 micrograms protein/mg lipid. This compares to only about 5% entrapment (70 micrograms protein/mg lipid) using the previous method. OVA loaded to pH-sensitive liposomes using the improved method led to a sensitive measure of CTL activity. The approach promises to be suitable to measure CTL against less available soluble antigens such as viral proteins.
Zhou
F
F
Department of Biochemistry, University of Tennessee, Knoxville 37996-0840.
Rouse
B T
BT
Huang
L
L
D000818:Animals; D000938:Antigen-Presenting Cells / Q000276:immunology*; D000941:Antigens / Q000008:administration & dosage*; D002021:Buffers; D002784:Cholesterol / Q000737:chemistry; D003602:Cytotoxicity, Immunologic; D005615:Freezing; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D008081:Liposomes / Q000737:chemistry*; D051379:Mice; D008810:Mice, Inbred C57BL; D009994:Osmolar Concentration; D010047:Ovalbumin / Q000008:administration & dosage / Q000737:chemistry / Q000276:immunology; D010714:Phosphatidylethanolamines / Q000737:chemistry; D012995:Solubility; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology; D013945:Thymoma / Q000276:immunology; D014280:Triglycerides / Q000737:chemistry; D014407:Tumor Cells, Cultured
D000941:Antigens; D002021:Buffers; D008081:Liposomes; D010714:Phosphatidylethanolamines; D014280:Triglycerides; D010047:Ovalbumin; D002784:Cholesterol