Editing MuLV Psueduvirus (section)

=== 5. Infectivity Quantification by Luciferase Assay Readout ===

'''NOTE:&nbsp;'''Perform initial steps in the biosafety cabinet.

#Thaw luciferin substrate (stored at -80 °C) and 5x luciferase assay lysis buffer (stored at -20 °C) until they reach room temperature.
#Dilute luciferase assay lysis buffer to 1x with sterile water.
#Aspirate supernatants of cells infected with pseudotyped particles.
#Add 100 µL of 1x luciferase assay lysis buffer to each well.
#Place the plate on a rocker and incubate for 15 min with rocking at room temperature (from this point onwards the plate can be handled outside of a biosafety cabinet).
#Prepare microcentrifuge tubes for each well by adding 20 µL of luciferin substrate in each tube.
#Turn on the luminometer.
#Perform luciferase activity measurement one well at a time by transferring 10 µL of lysate to one tube containing 20 µL of luciferin substrate.
#Flick the tube gently to mix contents, but avoid displacing the liquid on walls of tube.
#Place the tube in device and close lid.
#Measure the luminescence value of the tube by using the luminometer.
#Record the relative light unit's measurement.
#Repeat steps 5.8–5.12 until all wells are analyzed.

'''NOTE:&nbsp;'''With the appropriate equipment such as a plate reading luminometer, this process can be performed automatically. The assay will need to be scaled to the plate format (e.g.'',''&nbsp;96-well plate format).