INTRODUCTION: Apoptosis detection methods cause the cell culture medium to be physically or chemically affected. In addition, these methods have disadvantages such as cost and cell losses. The aim of this study is to develop a method to assess apoptosis in real-time using the back-reflection spectroscopy (BRS) in breast cancer cells.
METHODS: BRS experiment set up consists of a spectrometer, a tungsten-halogen light source and a fiber optic probe. In order to assess the changes associated with apoptosis, the intensity of the reflected light was measured with a fiber-optic probe from MCF7 breast cancer cell samples in which apoptosis induced doxorubicin. Spectroscopic measurements were performed 6 hours after incubation from control and doxorubicin groups. Cell viability measurement with MTT method, imaging of cell morphology with toluidine blue staining method, ROS measurement with DCFH-DA method were performed for the evaluation of apoptosis.
RESULTS: We analyzed the back reflection spectrum and determined the signal difference in apoptotic cell samples compared to the control. A significant difference was found in ROS measurements in all groups compared to the control. We did not observe any morphological change in the cells imaged with the toluidine blue method for 6 hours. Cell viability decreased 50% in all groups compared to the control for 24 hours.
DISCUSSION AND CONCLUSION: BRS is a new approach that may detect apoptosis in real time without interfering cell culture conditions. This method has the potential to be developed as a non-invasive, objective and reproducible technique that enables time-dependent monitoring of apoptosis.