FACS Canto II not showing events - (Apr/03/2014 )
We are having some troubles with our FACSCanto II cytometer: it is not showing any event in wells where we positively know there's a big amount of cells. We performed several primes to solve possible needle obstructions, followed by a long clean and all the possible cleaning modes existing in the machine. We also filled the fluidics filters, checked the waste tanks and cubitainers etc. we did everything we thought of, but still any event was being detected (no acquisition light was blinking). The lasers were checked about a month ago and we don't really think it is a laser problem...
Any idea where is the problem? We are going mad!
Thanks a lot
What you did would have been the exact things I'd have done too, especially obstruction is the first thing to think of. Four additional thoughts:
1.On our Calibur, we once had to solve an obstruction by carefully (!!) inserting a needle because priming and other forms of cleaning did not help.
2.When you run simple water and start acquisition, does it show some occasional dots or is there absolutely nothing at all ?
3.This may be basic, but is the flow rate high enough ? Once had the trouble that low flow rates on Canto did for some reason not draw any fluid at all, while it worked with medium or high flow rate.
4. It may be an internal hardware problem, in which case you should call the technician.
My reply may be redundent with Tabaluga's, yet i'd answer with my experience on the Canto2.
If the issue comes from the fluidics, you may need to clean it thoroughly (especially if, like we did, you use unfiltered "dirty" samples with lots of debris i.e.: spleen extracts) and/or remove bubbles.
For hardware problems, the best solution is to ask your BD technician who'll - of course - charge you big times (ours stopped working out of the blue because a software update needed some kind of new wires to plug to the computer).
Of course, checking your lasers calibrations wouldn't hurt
Sorry for up this topic! I know i'm a little bit late, but if I can't help you, maybe I can help other person.
In this situation is very important to do a CST for check the overall performance of the equipment. If you don't have events in the CST mode, you probably have a clog or hardware problem (in this last, you'll need to call the BD assistance).
But still are something very simple to do: you can check the threshold settings of your experiment (if you don't did this yet). Obviously, high levels of threshold (e.g. on FCS) will cut all the events and nothing will be detectable. On the bottom left of aquisition window, you can see the ignored events by threshold while aquiring the sample (threshold count). It is one simple difference of normal situation for the clog situation.
Perhaps this tip may seem foolish for you, but happened with me some times, when I lost time, FACS Clean/ Flow/ Shutdown, Rinse, Contrad and patience =P.
You mentioned that lasers LEDs was not blinked. That suggest the problem is not related with threshold. But anyway, check it! =D
Hope I helped someone!