Signal Generation from Switchable Polydiacetylene Fluorescence
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Signal Generation from Switchable Polydiacetylene Fluorescence Mary A. Reppy Analytical Biological Services Inc. Wilmington, DE 19801, U.S.A. ABSTRACT Chemical and biological sensors require a material component to act as a transducer from the molecular level event of interest to a discernable output measurable in the macroscopic world. One such material is polydiacetylene (PDA), a conjugated polymer that can switch from a non-emitting to a fluorescent state in response to environmental changes. This attribute can be harnessed to provide signal generation for bio-sensors and assays as a more sensitive alternative to the previously reported monitoring of PDA colorimetric shifts. While providing a more sensitive transduction mechanism the fluorescence behavior of PDA is also more complicated than the absorbance, in particular the emission profile of PDA in liposomes is strongly affected by the extent of polymerization. Incorporating small molecule fluorophores into the PDA materials further increases the overall emission of fluorescent PDA materials. The fluorophores accept energy from the excited state of the polymer and fluoresce, leading to both an overall increase in the quantum yield of the system and an increase in the Stokes shift. Basic photophysical properties of fatty acid PDA liposomes and a model assay for phospholipase A2 are presented. The model assay results show that the fluorescence response is greater than the colorimetric, and is further enhanced by addition of fluorophores. INTRODUCTION Liposomes containing polydiacetylene (PDA) can be used for colorimetric bio-assays in an approach that exploits the changeable absorbance properties of the conjugated polymer backbone of PDA and also takes advantage of the biomimetic aspect of surfactant liposomes. The liposomes are formed from diacetylene surfactants and the diacetylene tails are photopolymerized to form PDA in the alkyl regions of the liposomes. Changes in the conjugation state of the double and triple bonds of the PDA backbone due to external perturbations lead to shifts in the absorption maxima, typically causing a color change from blue to red. Charych et al photopolymerized diacetylene fatty acid liposomes with ligands or enzymatic substrates incorporated, to form PDA liposomes [1]. Exposure of the liposomes to solutions of a target analyte caused a gradual change in the color from blue to purple or red. Charych used PDA liposomes and films with sugars, gangliosides and phosphotidylcholines incorporated as colorimetric sensors for influenza virus [2], cholera toxin [3], and phospholipases [4]. Subsequently, other researchers have developed similar colorimetric assays using PDA liposomes and films [5]. For all these colorimetric assays the liposomes or films were prepared in the blue state and upon interaction with the analyte turned red. The change in the absorption properties is quantified by calculation of the “colorimetric ratio” (CR); the ratio of the intensity of the “blue” maximum absorbance peak (650nm) over the sum of the intensities o
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