If a fresh probe cannot be filled despite using the protocol given here it can have several reasons.
To find the cause fill fill the reservoir of the cantilever and apply 1 bar of pressure for 30 s. Now observe the water meniscus between the reservoir and the small channel of the CYTOCLIP:
- Does the meniscus not enter the channel at all?
- Does it move in upon increasing the pressure and move back when releasing the pressure?
- Does it move in a bit, but doesn't move back when releasing the pressure?
- Does it fill the channel but seemingly nothing exits the probe?
Case 1: There is a leak between pressure controller and probe. Typically the leak is at the connector. This can be tested by mounting the CONNECTOR to a dead-end CYTOCLIP, where the old chip is removed and the opening is blocked with glue. You can then immerse this test setup in a water-filled beaker and apply high pressure with a syringe. Any leaks will reveal themselves through bubbles.
Case 2: The probe opening or the channel is blocked. Discard the probe, write us a mail and we will replace it for free with your next order.
Case 3: The probe is open, but the pressure and time were not sufficient to fill the channel yet. Either wait for a few minutes with 1 bar applied or use a syringe as pressure source to achieve a higher pressure.
- If the opening is smaller than 1 um this is expected as the surface tension at the opening is higher than back pressure. Fluorescent tracer in the liquid can be used to reassure filling of probes with such small openings. The probe is ready to use as soon as the liquid reaches the tip.
- If the opening is larger than 1 um case 4 indicates that the probe is clogged. Either by particles in the liquid or due to fabrication residues in the very end of the channel. By adding fluorescent tracer to the liquid this two situations can be distinguished. A fabrication failure would show as dark area, while clogging due to particles gives a fully fluorescent cantilever.
- Fluorescent tracer can even be added after a first filling attempt. Simply add 5 ul tracer to the reservoir and wait until it diffuses through the probe channel, a waiting period of 1 hour should be enough.
Air in the probe
Even a filled probe will suck in air, if left without applied overpressure in air. Gas in the probe shows as compressible domain, whenever the pressure is changed.
- To get rid of the air, dry the probe by touching the FluidFM chip (avoid the probe) with a kimwipe.
- Apply 1 bar pressure to the probe while it is still in air
- Make sure the liquid reaches the opening of the probe
- Immerse the probe into your liquid medium/petridish while the pressure is still set to 1 bar
- Release the pressure