Note: DNase-free RNase should be used at a concentration such that no more than 10 L contains the full number of units needed to treat a sample. For example, the RNase Cocktail from Ambion has 500 U/ mL activity, so 4 L per well equals two units per well.
Step 3. Add 100 L of 2X CyQUANT GR dye/ cell-lysis buffer to each of the microplate wells, including the control wells without cells.
Step 4. Prepare the DNA standard curve by transferring the previously prepared DNA serial dilutions into the appropriate wells of the microplate, as defined in the template editor. The plate should contain two controls: a digest control prepared following the same steps as the cellular samples, but without cells (no DNA), and a standard curve control/plate blank containing only 200 L 1X CyQUANT GR dye/cell-lysis buffer (no DNA and no RNase).
Step 5. Incubate the samples for 2-5 minutes.
Set up the instrument and software
Step 1. Turn on the microplate reader. Launch the SoftMax Pro software and use the Assays drop-down menu to locate and open the Basic endpoint protocol in the Basic protocols folder.
Step 2. Set up the Instrument Settings dialog box as shown in Table 2.
Step 3. Create a template of the assay showing where standards,
unknowns (cells treated with RNase), and plate blanks will be located
on the microplate. To access the Template Editor, click the Template