The development of Raman imaging microscopy to visualize drug actions in living cells

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Ling, Jian

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Direct Raman imaging techniques are developed and applied to study the drug distribution in single living cells. From the drug distribution, its mechanism and efficacy can be evaluated. The advantages of Raman imaging are that the Raman signals are inherent to the drug, thus no external dyes, markers or labels are required during the imaging. This makes the sample preparation much simple for the experiment. At the same time, the mechanism of action of the drug is minimally disturbed during imaging. The major challenge in Raman imaging is the weak nature of the Raman signal. In this study, the quality and sensitivity of a Raman system was improved. A model was developed to describe the degradation of Raman signals by several processes: non-uniform illumination of the excitation laser, distortion by the microscope system, and the influence of additive signal-dependent Gaussian noise that arises during imaging. Using this model, special-purpose image-processing algorithms were developed to restore the Raman images. The general Raman imaging and data analysis techniques were then applied to the visualization of drug in living cells. Taxol, an important anticancer agent whose mechanisms at the cellular level have been well studied, was used to evaluate the capabilities of direct Raman imaging. Raman images were obtained from a MDA-435 cancer cell before, during, and after the drug treatment. The results clearly show how the Taxol distribution changes with time in a living tumor cell. It was also found that Taxol does not enter the cell nucleus, but is more concentrated around the cell centrosome and near the cell membrane. This finding is explained by the binding characteristics of the Taxol and its molecular target ñ the microtubules. This result demonstrated the feasibility of using direct Raman imaging to study the distribution of anticancer agents in single living cells. Direct Raman imaging can also be extended to study drug uptake, resistance, and intracellular pharmacokinetics. I believe Raman imaging will not only provide a cost-effective tool for the study of drug mechanisms at the cellular level, but also be a general molecular imaging technique to be used in many applications.