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Instruments / Technical Area	Publication Details	Published	Actions
B22-Multimode InfraRed imaging And Microspectroscopy	Live single cell analysis using synchrotron FTIR microspectroscopy: development of a simple dynamic flow system for prolonged sample viability James Doherty , Alan Raoof , Ansaf Hussain , Magda Wolna , Gianfelice Cinque , Michael Brown , Peter Gardner , Joanna Denbigh Analyst DOI: 10.1039/C8AN01566J Diamond Proposal Number(s): [18968, 19084] Show Abstract ▼ Abstract: Synchrotron radiation Fourier transform infrared microspectroscopy (SR-microFTIR) of live biological cells has the potential to provide far greater biochemical and morphological detail than equivalent studies using dehydrated, chemically-fixed single cells. Attempts to measure live cells using microFTIR are complicated by the aqueous environment required and corresponding strong infrared absorbance by water. There is also the additional problem of the limited lifetime of the cells outside of their preferred culture environment. In this work, we outline simple, cost-effective modifications to a commercially available liquid sample holder to perform single live cell analysis under an IR microscope and demonstrate cell viability up to at least 24 hours. A study using this system in which live cells have been measured at increasing temperature has shown spectral changes in protein bands attributed to α-β transition, consistent with other published work, and proves the ability to simultaneously induce and measure biochemical changes. An additional study of deuterated palmitic acid (D31-PA) uptake at different timepoints has made use of over 200 individual IR spectra collected over ~4 hours, taking advantage of the ability to maintain viable cell samples for longer periods of time in the measurement environment, and therefore acquire greatly increased numbers of spectra without compromising on spectral quality. Further developments of this system are planned to widen the range of possible experiments, and incorporate more complex studies, including of drug-cell interaction.	Oct 2018
B22-Multimode InfraRed imaging And Microspectroscopy	Increased optical pathlength through aqueous media for the infrared microanalysis of live cells James Doherty , Zhe Zhang , Katia Wehbe , Gianfelice Cinque , Peter Gardner , Joanna Denbigh Analytical And Bioanalytical Chemistry 99 DOI: 10.1007/s00216-018-1188-2 Diamond Proposal Number(s): [15375, 17811] Open Access Show Abstract ▼ Abstract: The study of live cells using Fourier transform infrared spectroscopy (FTIR) and FTIR microspectroscopy (FT-IRMS) intrinsically yields more information about cell metabolism than comparable experiments using dried or chemically fixed samples. There are, however, a number of barriers to obtaining high-quality vibrational spectra of live cells, including correction for the significant contributions of water bands to the spectra, and the physical stresses placed upon cells by compression in short pathlength sample holders. In this study, we present a water correction method that is able to result in good-quality cell spectra from water layers of 10 and 12 μm and demonstrate that sufficient biological detail is retained to separate spectra of live cells based upon their exposure to different novel anti-cancer agents. The IR brilliance of a synchrotron radiation (SR) source overcomes the problem of the strong water absorption and provides cell spectra with good signal-to-noise ratio for further analysis. Supervised multivariate analysis (MVA) and investigation of average spectra have shown significant separation between control cells and cells treated with the DNA cross-linker PL63 on the basis of phosphate and DNA-related signatures. Meanwhile, the same control cells can be significantly distinguished from cells treated with the protein kinase inhibitor YA1 based on changes in the amide II region. Each of these separations can be linked directly to the known biochemical mode of action of each agent.	Jul 2018

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