ROX Dye qPCR Calibration Solution *10,000X*

This generic protocol is suitable for the ABI7500 FAST qPCR instrument. For complete instructions on use, refer to the maintenance guide for your instrument.

1. Prepare a ROX Dye qPCR Calibration working solution by mixing 1 µL of the 10,000X ROX Dye qPCR Calibration Solution with 10 mL of nuclease-free water or qPCR buffer.

   Note: The final dye concentration must be optimized based on the calibration requirements for your instrument. Refer to the manufacturer’s maintenance guide for detailed instructions.

2. Add 20 µL of the ROX Dye qPCR Calibration working solution to all wells of a reaction plate.

   Note: The volume may vary depending on your instrument. Refer to the manufacturer’s maintenance guide for detailed instructions.

3. Seal the plate with an optical adhesive cover.

4. Centrifuge the plate for 2 minutes at < 1500 rpm.

5. Check that the liquid in each well of the plate is at the bottom of the well. If not, centrifuge the plate again at a slightly higher rpm and for a longer period of time.

6. For complete calibration instructions, refer to the maintenance guide for your instrument.