 |
 |
 |
 |
> I request some advice regarding sorting.
> Question: We use a FACStar for sorting. However, inspite of my best
> efforts I am not able to get the Left and Right Streams to separate much
> from the center stream. As a result the sorted cells miss the collection
> tubes. (The left and rt. stream deflection controls are turned to max).
>
> - Does the angle between the 2 charged metal plates (which is adjustable)
> have anything to do with it ?.
> - Any other suggestions would be welcome.
Yes, the angle of the deflection plates is important; in our FACStar+, we
adjust ours to about 2mm apart at the top and equally distant from the stream.
[ASIDE: We have made a pair of curved deflection plates for our FACS II with
curvature well matched to the deflected droplet trajectory. Deflection is at
least 50% better and we can run quite a bit faster with no other modifications
(not necessarily relevant here but may be of general interest).]
Other sugestions are:
1. B_D nozzle holders are all slightly different. Sometimes the wire feeler
contact for the deflection pulses does not make good connection with the
stainless steel sample inlet tube. You may try unscrewing the nozzle holder,
dropping it down and then *GENTLY* deflecting the wire feeler down *a little*.
After screwing the nozzle holder back in, wiggle the feeler back and forth
along the inlet tube a little to make a good contact.
2. The contact's insulation may be wet or covered with a salt film which
leaks away the deflection pulses. *Carefully* (the contact is quite fragile)
wash around the insulation with distilled water to remove the salt and then
alcohol to remove the water.
3. The deflection plates can also be wet or covered with a salt film which
leaks away the high voltage. Remove them and clean them and also clean around
the contacts they plug into. Replace them without splashing saline on them.
4. If your sheath pressure is too high, the droplets will deflect less. With
a 70micron nozzle, we use about 12psi gas pressure with a 0.2micron
syringe-type filter in the saline line (you would need a lower gas pressure
without the sheath filter resisting the sheath flow). You should be able to
get your deflected streams to go down the middle of the collection tubes.
\ / < Flow Systems Laboratory
Frank Battye \__/ <<<<< The Walter & Eliza Hall Institute
Robyn Muir ----------!!<<<<<<<< Victoria 3050, AUSTRALIA
Dora Kaminaris /!!\ <<<<< ph: 61_3_9345 2540 fax: 61_3_9347 0852
o !! \ < in:: "facs_copy@wehi.edu.au"
|
|
|
|