Confocal Raman microscopy for monitoring the membrane polymerization and thermochromism of individual, optically trapped diacetylenic phospholipid vesicles

Optical-trapping confocal Raman microscopy allows the 1, 4-addition reaction of diacetylenic functional groups in 1,2-bis(10,12-tricosadiynoyl)-sn-glycero-3-phosphocholine lipids to be monitored in individual phospholipid vesicles. Optical trapping allows a single vesicle to be observed over time, allowing the direct observation of structural changes in the vesicle membrane during polymerization. Confocal Raman microscopy excludes light collection outside the optical-trap region avoiding interferences from the surrounding solution, while chemical reactions occurring in the membrane of the trapped vesicle can be measured with high sensitivity. Individual, optically trapped liposomes (0.6 mu m in diameter) were exposed to photolysis radiation at 254nm. Upon exposure to UV light, the cross-linking polymerization reaction formed a conjugated eneyne backbone in the bilayer of the optically trapped vesicle. Polymerization produces two different polymers, red and yellow in color, which can be distinguished structurally by their Raman spectra. Rates of red and yellow polymer formation were monitored by the Raman scattering intensities from both C=C stretching vibrations at 1455cm1 and 1508cm1 and C=C stretching vibrations at 2080 and 2110cm1, respectively. Polymer formation rates depended linearly on 254-nm light intensity, consistent with a one-photon excited polymerization reacting in a photostationary state. Relative populations of red and yellow polymer in a polymerized vesicle depend sensitively on the sample temperature. From temperature-dependent Raman spectra, the enthalpy change of the red-to-yellow thermochromic response and corresponding structural changes in the polymer could be determined. Copyright (C) 2011 John Wiley & Sons, Ltd.