Liposomes as adjuvants with immunopurified tetanus toxoid: influence of liposomal characteristics

Abstract

The effect of various manipulations of liposomes composed of equimolar phospholipid and cholesterol on immune responses to the incorporated immunopurified tetanus toxoid was investigated in BALB/c mice. In studies designed to establish proper dosage for immunization and to reveal the roles of the liposomal phospholipid to toxoid mass ratio, gel-liquid crystalline transition temperatures (Tc) of the liposomal phospholipids and mode of antigen incorporation into liposomes on immune responses, animals were injected intramuscularly with various amounts of the toxoid, free, entrapped in multilamellar dehydration-rehydration vesicles (DRV) or covalently linked to the surface of multilamellar vesicles (MLV) prepared by the classical procedure. Two identical injections separated by 4 weeks were given and IgG1 and IgG2b antibodies specific for the toxoid assayed in sera by an enzyme-linked immunosorbent assay. Results suggest that the adjuvant effect of liposomes is improved considerably when liposomal phospholipid to toxoid mass ratios are as high as 2049:1; however, adjuvanticity is reduced to reach very low levels for much higher ratios (e.g. 90,361:1); antibody responses are similar for liposomes (phospholipid to toxoid mass ratios: 14.3-33.1) made of a variety of phospholipids with Tcs ranging from -32 degrees to 41.5 degrees but are low or non-existent when liposomes are made of distearoyl phosphatidylcholine (Tc, 54 degrees) (however, see Discussion); there are no differences in antibody responses between liposomes with entrapped and surface-linked toxoid.