Malaria is a disease generally found in a tropical area including Thailand. It is widely known that the biological technique such as PCR normally used for an accurate detection of malaria-infected blood requiring a considerable period to repeat the process. Raman spectroscopy is considered to be an alternative method for the malaria infected blood detection. Theoretically, Raman spectroscopy is based on the scattering process that is less likely in a normal situation. Therefore, an enhancing technique known as “surface-enhanced Raman scattering (SERS)” is required for the signal augmentation. The SERS provides the enhancement of the electric field near the surface of the substrate. With the application of the technique, the main target of this research focuses on the comparison of the SER Raman spectra of the normal red blood cell and malaria infected red blood cell. However, only a single spectrum cannot provide a clear difference between the normal and the infected blood. An additional tool for even more effective discrimination was provided by using PCA analysis. In the sample preparation stage, the spin coating process was applied to spread the red blood cells uniformly on the surface. In addition, the spectra of the red blood cells including media were collected in various conditions in terms of the excitation wavelengths and the types of substrate. This additional information can be used as references for any red blood cell related investigation using Raman spectroscopy.