The Pressure Not At Setpoint error can occur at two different time points during the partitioning.
1) If the error occurs within the first 3-5 seconds of the run, the pressure applied by the manifold that sits on top of the gasket, cannot be established properly. Take out the chip and check the gasket for debris and other impurities. Carefully remove the gasket and gently wipe with isopropanol. Alternatively, apply a new gasket to the chip. Once the chip is loaded into the instrument, confirm that it is firmly mounted to the tray.
2) If the error occurs with ~2-3 min left (for Single Cell Assay) of the run, either the chip ran out of partitioning oil before the completion of the run or a wetting failure in >3 channels occurred during partitioning. Because of the improper channeling of reagents in the chip the oil in the channels with the wetting failure runs out early, causing air bubbles to occur in the outlet wells. At this point the pressure sensor measurement sees that the pressure across all wells of the chip is “out of the acceptable pressure window” and the system stops with an error. Because the system stops, you also lose the other channels that may have been running perfectly, have formed emulsions, but have stopped early without the pushback step at the end of the run where the excess oil in the outlet well is forced back into the chip, enabling an easier extraction of the emulsion from the chip.
Firmware version 1.2.1 and above have a parameter change to stop sensing the pressure soon after the run begins, allowing the run to continue and samples that are free of wetting failures to complete emulsion formation and recovery. To update the firmare to the latest version available see the Support pages of the 10x Genomics website.
When running the Readiness Test after a firmware update confirm that the test that was done was the “Test Readiness” using the Test chip and not a Training chip. Note also that the test chip is meant to be used without any reagents (i.e. dry).