Dandekar & Chandler:AHL Extraction: Difference between revisions

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== AHL Signal Extraction  ==  
== AHL Signal Extraction  ==  


In a large test tube containing the sample you wish to test:
Use only glass tubes to reduce loss of signal during extraction and storage!


* Add 5mL of ethyl acetate and vortex on high for 30seconds
In a large test tube (18 mm) containing the sample you wish to test:
* Let the sample sit for 10 minutes to allow the two phases to separate out.   
 
* After the the 10 minute rest, pipette off the clear acetate layer only. Be careful to not draw up any of the bottom cell debris layer.  
# Add 5mL of ethyl acetate and vortex on high for 30seconds
* Deposit the clear acetate layer in a 10mL glass tube with a sealable air tight cap.   
# Let the sample sit for 10 minutes to allow the two phases to separate out.   
* Repeat steps 1-4 and add acetate supernatant to tube from first cycle.  
# After the 10 minute rest, pipette off the clear acetate layer only being careful to not draw up any of the bottom cell debris layer.  
* Place acetate filled tubes into N2 evaporator on medium until all acetate has evaporated off.  
# Deposit the clear acetate layer in a 10mL glass tube with a sealable air tight cap.  [https://www.fishersci.com/shop/products/fisherbrand-disposable-borosilicate-glass-tubes-polypropylene-screw-cap-6/p-196656 This type of vial] works well. (Fisher 14-962-26F)
* Suspend dried signal in 100uL of ethyl acetate, screw sealable cap on tightly, wrap in parafilm and place in -20 freezer.
# Repeat steps 1-4 and add acetate supernatant to collection tube from step 4.  
# Place acetate filled tubes into N2 evaporator until all acetate has evaporated off. (keep on low level to prevent splashing)
# Suspend dried signal in 100uL of ethyl acetate, screw sealable cap on tightly, wrap in parafilm and place in -20 freezer.

Revision as of 12:21, 11 August 2015

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AHL Signal Extraction

Use only glass tubes to reduce loss of signal during extraction and storage!

In a large test tube (18 mm) containing the sample you wish to test:

  1. Add 5mL of ethyl acetate and vortex on high for 30seconds
  2. Let the sample sit for 10 minutes to allow the two phases to separate out.
  3. After the 10 minute rest, pipette off the clear acetate layer only being careful to not draw up any of the bottom cell debris layer.
  4. Deposit the clear acetate layer in a 10mL glass tube with a sealable air tight cap. This type of vial works well. (Fisher 14-962-26F)
  5. Repeat steps 1-4 and add acetate supernatant to collection tube from step 4.
  6. Place acetate filled tubes into N2 evaporator until all acetate has evaporated off. (keep on low level to prevent splashing)
  7. Suspend dried signal in 100uL of ethyl acetate, screw sealable cap on tightly, wrap in parafilm and place in -20 freezer.