Microfluidics is an expanding research area for analytical chemistry and the biomedical industry. Microfludic devices have been used for protein and DNA sorting, early detection techniques for cancer and other disease, and a variety of other analytical techniques. Dielectrophoresis is a technique is often used to control particles within microfluidic devices however the non-uniform electric field can affect the interior of the device. In order to expand the applications of microfluidic devices and to make it easier to work with techniques such as dielectrophoresis, it is essential to understand as much as possible about how the internal environment of the device will affect the sample. A significant part of this is being able to non-invasively determine the temperature inside the microfluidic device in the both the channel and reservoir regions. Several other research group have successfully used temperature sensitive dyes and fluorescence to measure the temperature within microfluidic devices so research began with understanding their techniques and trying to optimize them for the chosen microfluidic channel. Results from calibration and reservoir tests show that there is a linear relationship between the temperature of the channel and the ratio between the dyes Rhodamine 110 and Rhodamine B. Results within the channel showed that the calibration may be difficult to apply directly as absorption from the PDMS continues to be a problem but several coatings can be used to improve the results.