Most of the issues relating to partial blocking of nanopores can be minimised with thorough care during sample preparation - using clean tubes, filtered fresh electrolyte and Izon qEV columns to isolate EVs. However, bubbles and contamination may lead to partial blocking of the nanopore.
A good measurement should consist of a run that has a stable baseline current which meets these requirements:
- Signal trace largely horizontal (current drift is 1% per minute)
- RMS noise of < 15 pA (blockade magnitude should lie above the orange RMS noise line, ideally in the green zone)
- Additionally, a minimum particle count of 200 for calibrations and 400 for samples.
Should you come across a partial blockage during a measurement run:
- Pause the measurement when a blockage occurs to avoid measurement inaccuracies.
- Follow the flowchart to recover baseline current:
- Once the baseline current has been recovered, apply the same pressure and resume recording.
- If the above steps don't recover the baseline current, start the nanopore setup sequence again either with the same or a new nanopore.
- Consider changing the sample's concentration or make up a brand new sample.
- For some particles that are prone to blocking (more than 3 times per measurement/pressure), consider increasing nanopore stretch or use a larger nanopore.
If the above suggestions don't resolve the issue, contact Izon Support
Watch Emma demonstrate these techniques: