Smolke:Protocols/Guidelines

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(Filtering Samples)
(Filtering Samples)
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**Clean-up:  
**Clean-up:  
***Mammalian samples: Soak in 10% bleach for about 1 day, then soak in water about 1 day, then dry and return sheets to bag.
***Mammalian samples: Soak in 10% bleach for about 1 day, then soak in water about 1 day, then dry and return sheets to bag.
-
***Yeast samples: Soak in water for a couple of days, then dry and return sheets to bag. (YHW: I usually spread EtOH on it while drying)
+
***Yeast samples: Soak in water for a couple of days, then dry and return sheets to bag. (YHW: I usually spread ethanol on it while drying)
*Cell strainers (BD Falcon Cat. No.: 352340)
*Cell strainers (BD Falcon Cat. No.: 352340)
**Not consistently stocked. Order if needed.
**Not consistently stocked. Order if needed.

Revision as of 19:40, 19 July 2012

Contents

Start Up

Instructions for machine start up and various other functions are on the lamenated guides near the VYB.

If machine is off:

  • Turn on machine by touching to screen.
  • Logon and put the machine into data acquisition mode.
  • Run back flush (~16 min).
  • Run clean with 1% hypochlorite (~10 min).
    • NOTE: 100% bleach = 15% hypochlorite.
    • NOTE: Clean only runs if machine is in the single tube mode.
  • Run calibration beads.

If machine is already on, assume it is not clean:

  • Run back flush (~16 min).
  • Run clean with 1% hypochlorite (~10 min).

Shut Down

  • When finished, ALWAYS make sure there is ENOUGH BUFFER and ENOUGH SPACE in the waste container. Otherwise, shut down will stall and leave sticky, dead, clog-inducing samples in the machine until someone notices.
  • Shut down the machine unless another user is signed up within a couple of hours after you finish, but check buffer and waste in case later user cancels.
  • Machine will automatically attempt to shut down ~2 hrs after its last fluidics function. However, you must still check buffer and waste before leaving the machine.

96-Well Plates

  • The VYB can only run ONE full 96-well plate if it has a FULL bottle of running buffer and an EMPTY waste bottle.
  • Make sure the running buffer bottle is full and waste bottle is empty before EACH 96-well plate.
  • NOTE: In the future, there will be larger buffer bottles & this may no longer be a problem.

Empty Buffer Bottles

  • Rinse with water.
  • Remove the label and lid.
  • Recycle (blue atrium bin).

Filtering Samples

ALWAYS FILTER YOUR SAMPLES.

  • Nylon filter sheets (http://www.amazon.com/Microns-Square-Opening-Thread-Diameter/dp/B000FN0OG2/ref=pd_sbs_indust_5)
    • Available in zip-lock bag above VYB bench.
    • Clean-up:
      • Mammalian samples: Soak in 10% bleach for about 1 day, then soak in water about 1 day, then dry and return sheets to bag.
      • Yeast samples: Soak in water for a couple of days, then dry and return sheets to bag. (YHW: I usually spread ethanol on it while drying)
  • Cell strainers (BD Falcon Cat. No.: 352340)
    • Not consistently stocked. Order if needed.
    • Disposable.

Quirks

  • The software is frozen.
    • If you are running a sample, leave the machine alone. It is probably busy recording a lot of data and will recover once it finishes (up to ~15 min).
  • I can't run 'Clean'.
    • Make sure the machine is set in the single tube mode.
  • My samples look weird in FlowJo.
    • In FlowJo, adjust the file properties to match the decade settings used on the VYB.
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