Induction and Detection of Antibodies to Squalene
WALTER REED ARMY INST OF RESEARCH SILVER SPRING MD DEPT OF MEMBRANE BIOCHEMISTRY
Pagination or Media Count:
An enzyme-linked immunosorbent assay ELISA utilizing antigen coated on hydrophobic polyvinyldiene fluoride PVDF membranes is described for detecting antibodies that bind to squalene SQE. Because of the prior lack of availability of validated antibodies to SQE, positive controls for the assay were made by immunization with formulations containing SQE to create monoclonal antibodies mAbs that reacted with SQE. Among eight immunogens tested, only two induced detectable murine antibodies to SQE liposomes containing dimyristoyl phosphatidylcholine, dimyristoyl phosphatidylglycerol, 71 SQE, and lipid A L71 SQE1LA, and, to a much lesser extent, an oil-in-water emulsion containing SQE, Tween 80, Span 85, and lipid A. In each case, lipid A served as an adjuvant, but neither SQE alone, SQE mixed with lipid A, liposomes containing 43 SQE and lipid A, nor several other emulsions containing both SQE and lipid A, induced antibodies that reacted with SQE. Monoclonal antibodies produced after immunizing mice with L71 SQE1LA served as positive controls for developing the ELISA. Monoclonal antibodies were produced that either recognized SQE alone but did not recognize squalane SQA, the hydrogenated form of SQE, or that recognized both SQE and SQA. As found previously with other liposomal lipid antigens, liposomes containing lipid A also induced antibodies that reacted with the liposomal phospholipids. However, mAbs were also identified that reacted with SQE on PVDF membranes, but did not recognize either SQA or liposomal phospholipid.
- Medicine and Medical Research