Quantification of drug in live cells using label-free multi-bounce attenuated total reflection Fourier Transform Infrared (MB-ATR FTIR) spectroscopy

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In the process of developing a new medicine, it is important to know how much of the medicine is absorbed by a human cell. This is usually achieved by adding a dye to label the medicine in order to visualise the amount of the medicine absorbed into the cell using visible or ultra-valet light. However, the added labelling dye may have an unknown effect to the overall results, therefore there is a need to develop and apply a new method that does not require any labelling of the medicine. This project will investigate on how to use a method called Fourier Transform Infrared (FTIR) spectroscopy, which is a label-free while chemically specific (this method can distinguish a large range of chemicals without labelling) analytical technique, to study the amount of medicine in living cell without any added labelling dye. FTIR spectroscopy is a well-established method for the analysis of medicine. However, it has not been used for measuring the amount of medicine in living human cell because, first, it is normally not sensitive enough to detect the small amount of medicine in a living cell. Second, it is not suitable to measure samples that are surrounded by a large amount of water such as live cells. These obstacles, fortunately, can be overcome by using a special measurement mode called the multi-bounce attenuated total reflection (MB-ATR) which can increase the sensitivity by at least 10 times as well as allowing the measurement of live cells bathing in aqueous medium. The project will use a medicine for treatment of cancer (doxorubicin) to demonstrate, for the first time, the possibility of using the FTIR technique to measure the amount of the medicine in living cells at concentration levels that is relevant to the treatment of the disease (~10 micro molar). This can be achieved by optimising the MB-ATR measurements through using the optimal number of bounces, ATR materials and measurement parameters so that the signal is maximised while the noise is minimised. The result of this development will then be compared to the results from the traditional methods to demonstrate that the FTIR technique is a reliable tool for studying medicine in living cell.