Protein Expression and Purification Series - Bio-Rad

Protein Expression and Purification Series
Measuring Change in Absorbance at 340 nm for Enzyme Activity Calculation
1. Turn on the SmartSpec Plus spectrophotometer and press Kinetics.
Enter reading wavelength
for kinetics assay: nm
2. Enter “340” using the numeric keypad and then press Enter.
3. Enter “150” using the numeric keypad and then press Enter.
4. Enter “15” using the numeric keypad and then press Enter.
5. Press Enter to select NO.
6. Place the cuvette containing 1x PBS (the blank) into the chamber. Close the sample compartment
door, and press Read Blank.
7. Press to accept the blank reading.
8. Remove the cuvette containing 1x PBS from the chamber. Prepare your “No Substrate Control
Reaction Sample” in the cuvette and place the cuvette into the chamber, close the sample
compartment door and press Read Sample.
Note: If you do not have printer paper in your spectrophotometer, manually record each data point for
all time points as they appear on the screen display.
Appendix D: SmartSpec Plus Setup
Enter duration of timed
assay: seconds
Enter interval of data
collection: sec
Do you want to substract
background reading? NO
Ready to read absorbance
=Options
A340= 0.000
>=continue
Ready to read absorbance
=Options
Display Absorbance: 150
A340=0.342 45
221
Example data for the 45 second time point
APPENDIX D
SETTING UP THE
SMARTSPEC PLUS