Polydiacetylene Liposomes Attached to Glass Fibers for Fluorescent Bioassays

Abstract

ABSTRACTPolydiacetylene (PDA) is a conjugated polymer that changes its fluorescent state in response to environmental changes and can act as a transducer to convert molecular interactions into a discernable output measurable in the macroscopic world. Energy transfer to fluorophores can further enhance the fluorescent signal. Diacetylene liposomes can be prepared with phospholipids and other cell membrane components in the liposomes, and photopolymerized. PDA liposomes have been used for absorbance based detection of biological targets; however, moving to fluorescence detection gives increased sensitivity and also allows sensing from PDA structures deposited on opaque membranes. Unfortunately, PDA liposomes are prone to aggregation, particularly in the presence of divalent cations. Many enzymes require divalent cations such as Mg2+, Mn2+, Ca2+, etc., as co-factors; the tendency of PDA liposomes to aggregate in the presence of these cations limits their use as a platform for detection of enzymatic activity. We have developed methods for attaching PDA liposomes to glass fiber membranes, via thiol-epoxide coupling chemistry, for use in bio-assays and have seen that these materials can be used in place of PDA liposome solutions. We present here the attachment of PDA liposomes to glass fiber membranes in 96-well format, cryogenic TEM scans of the attached liposomes and the use of these materials in fluorescence assays to detect the activity and inhibition of phospholipase A2.