Method for the determination of antibiotic susceptibility through stimulated Raman metabolic imaging

A hyperspectral stimulated Raman scattering imaging apparatus for collecting images for antibiotic susceptibility testing, the apparatus comprising:

• a dual output femtosecond pulse laser, the dual out femtosecond pulse layer including:
• a pump beam;
• a Stokes beam;
• the Stoke beam path modulated by an acousto-optical modulator;
• the pump beam path having a translational stage to tune a delay between the pump beam and the Stokes beam;
• a combiner, the combiner adapted to combine the pump beam and the Stokes beam;
• a laser scanning microscope, the laser scanning microscope having an objective adapted to focus the pump beam and the Stokes beam on a sample, and an oil condenser, the oil condenser adapted to collect the laser from the sample;
• a filter, the filter positioned after the condenser and adapted to filter out the Stokes beam;
• a photodiode, the photodiode positioned after the filter, the photodiode detecting the pump beam;
• and
• a lock in amplifier, the lock in amplifier adapted to extract the pump beam signal loss.