An anti-2-phenyloxazolone single-chain antibody was expressed in Esche
richia coli as a lipoprotein fusion in order to generate a biosyntheti
cally lipid-tagged molecule [Laukkanen et al. (1993) Protein Eng. 6, 4
49-454]. For purification, a hexahistidinyl tag was introduced to the
C-terminus of the protein. The resulting antibody, termed Ox lpp-scFv-
H6, was membrane-bound, displayed hapten-binding activity, and contain
ed the lipoprotein-specific lipid modification as indicated by metabol
ic [H-3]palmitic acid labeling. The Ox lpp-scFv-H6 was purified by imm
obilized metal affinity chromatography followed by hapten-based affini
ty chromatography to essential homogeneity with a yield of 0.4-1.6 mg/
L of culture. In detergent dialysis, the purified antibody partitioned
quantitatively into phospholipid liposomes. The immunoliposome prepar
ation consisting of a homogeneous population of unilamellar 100-200 nm
vesicles displayed specific hapten-binding activity as measured by us
ing ELISA and surface plasmon resonance (SPR)-based realtime biospecif
ic interaction analysis. In SPR experiments, the immunoliposomes exhib
ited virtually irreversible binding to immobilized hapten compared to
soluble antibody fragments, consistent with the predicted multivalent
binding. Biosynthetic lipid-tagging of antibodies may prove useful for
immunoliposome-based diagnostic and therapeutic applications.