In the current study, we investigated the feasibility of Fourier transform infrared (FTIR) microscopy as a rapid diagnosis tool for bacterial infection by taking the example of bactericidal effect of ampicillin on Staphylococcus aureus at different doses-and time- manners. The overall data follow similar but distinct pattern with wavelength. Taking closer look at the wavelength between 600-1800 cm^-1, we found that FTIR can easily differentiate wild type and ampicillin-treated S. aureus even at low concentration (0.25ug/ml) of ampicillin. Particularly, in section of 1010-1070 cm^-1, the optical density (OD) vs. dosage and period of ampicillin demonstrated correlation of linear regression demonstrated by the regression models. Principal component analysis can group the OD values into two distinguishable classes, further subclass into 7 different groups based on their culture period. As a conclusion, FTIR can discriminate the slightly differentiated OD values of S.aureus treated with ampicillin in a time and dose dependent manner.