UV Vis
UV-Visible Spectoscopy
UV-Vis is used to measure absorption spectroscopy in the ultraviolet-visible spectral region. We are using the UV-Vis to measure the activity of peroxidases, starting with HRP to develop a standard protocol. We used the kinetics mode of the UV-Vis to gather our data.
Steps to Run a Peroxidase through the UV-Vis
1) Turn on UV-Vis
2) Open the UV-Vis software on the computer monitor
3) Turn on Kinetics Mode
a) Set wavelength to the desired absorption (ours was 436 nm for Guaiacol)
b) Set a cycle time - the lower the cycle time the more data collected (we used 0.5 seconds)
c) Set a run time - HRP does not show significant data past a minute (so we used 60 seconds)
4) Select time-based measurements and create a name for each set of data before running the sample
5) Run a blank with DI water before running a sample
6) Fill a cuvette with the assay and place the cuvette in the UV-Vis
7)Add desired amount of peroxidase to the assay and mix with a plastic transfer pipette
8)Select the sample button on the monitor