Before data acquisition, the Wavelength and and Raman shift of the scattered light needs to be calibrated. A gas-discharged neon lamp needs to be used for wavelength calibration.

Wavelength Calibration:

Step 1: Taking spectrum data using neon lamp and then making plot of neon spectrum: Intensity vs. pixel.

Step 2: Using theoretical neon spectrum plot ( Intensity vs. wavelength) to identify neon peaks on the plot in Step 1 and making plot: wavelength vs. pixel.

Step 3: Making a third-order polynomial regression of the known wavelengths of neon on the corresponding pixel numbers.

Now, pixel numbers are able to be converted to wavelength by using the third-order polynomial relation.

Raman shift calibration:

Step 1: Taking spectrum data from any sample with known Raman shift spectrum(like Tylenol)

Step 2: Converting pixel to wavelength for Tylenol data and then plotting Tylenol spectrum: wavelength vs. pixel

Step 3: Using theoretical spectrum of Tylenol to identify peaks on the plot of Step 2 and save these peak points

Step 4: Using the equation: 1/laser wavelength = Raman shift +1/ wavelength of tylenol vibration to estimate the laser wavelength for each Tylenol vibration. Then take the average of laser wavelength.

Step 5: By using the equation in step 4 and the average laser wavelength, the wavelength is able to be converted into Raman shift