If you’re running an experiment with a custom cycling protocol, use the “Create an Experiment” option to set up your run. For test kits with integrated Open qPCR software such as a Chai Coronavirus Test Kit or PIKA kit, use the “Run a Test Kit” option. The following workflow walks through the “Create an Experiment” option. Additionally, here's a Create a New Experiment video explanation.
Step 1: Create the Experiment. Select the “Create an Experiment” button on the home page to create and name your experiment.
Step 2: Edit the Cycling Protocol. Modify the cycling protocol as necessary from the default protocol. The default protocol includes one holding stage (Stage 1) and one 40 cycle Cycling Stage (Stage 2). Additional holding, cycling, or melt curve stages can be included by selecting “Add Stages” under “Edit Steps and Stages.” Additional steps within existing stages can be included by selecting “Add Step” under “Edit Steps and Stages.”
The example requires a 25 °C hold for 5 minutes, then a 62 °C hold for 5 minutes, then a 95 °C hold for 30 seconds. Add steps to the current Holding stage and modify the time and temperature details directly on the orange screen or on the bottom of the page under “Temperature Details”.
Finish modifying the remainder of the cycling protocol by adding, deleting, or modifying the cycling protocol as necessary. This example includes 40 PCR cycles in its cycling protocol. The cycling protocol is 95 °C for 10 seconds, followed by 62 °C for 40 seconds. Ensure that the “Gather Data” option is toggled on for the data collection step.
Step 3: Assign Samples + Targets. Use the toolbar on the left side to navigate to “Samples + Targets.” Input the name of your samples and targets and whether the targets are detected in Channel 1 (FAM) or Channel 2 (HEX/VIC/JOE, dual channel Open qPCRs only). For this example, we will add three samples and two targets.
Step 4: Assign Well Layout. Navigate to “Well Layout” using the toolbar on the left and assign individual samples and targets to each well as desired for the run. Sample names do not need to be set for positive and negative control wells. Select the Well Type for each target (S for Standard, U for Unknown, + for Positive Control, and – for Negative Control/NTC).
For this example, we will include a Positive Control in Well A1, a Negative Control in Well A2, and samples in Wells A3, A4, and A5.
Step 5: Start the Experiment. Ensure that the lid of the Open qPCR is closed, then select “Start Experiment” in the upper right hand corner. Select “Confirm Start” and the experiment will begin.
Step 6: Review Results. View the progress of the experiment during or after the run by navigating to the Amplification Curve. Amplification curve data will only be available once the machine starts taking fluorescence measurements. You can also access your Standard Curve or Melt Curve if you included either of these for your run.