Dandekar & Chandler:AHL Extraction: Difference between revisions

Revision as of 14:26, 22 January 2014

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In a large test tube containing the sample you wish to test:

Add 5mL of ethyl acetate and vortex on high for 30seconds
Let the sample sit for 10 minutes to allow the two phases to separate out.
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.
Deposit the clear acetate layer in a 10mL glass tube with a sealable air tight cap.
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.

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-== N-Acyl homoserine lactone (AHL) Extraction ==
+== AHL Signal Extraction  ==
-===Principle===
+In a large test tube containing the sample you wish to test:
+# Add 5mL of ethyl acetate and vortex on high for 30seconds
-===Materials Needed===
+# Let the sample sit for 10 minutes to allow the two phases to separate out.
+# 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.
-* Spectrophotometer capable of reading 520 nm
+# Deposit the clear acetate layer in a 10mL glass tube with a sealable air tight cap.
-* Vortex
+# Repeat steps 1-4 and add acetate supernatant to collection tube from step 4.
-* 10mL test tube
+# Place acetate filled tubes into N2 evaporator until all acetate has evaporated off. (keep on low level to prevent splashing)
-* Sterile Cuvets
+# Suspend dried signal in 100uL of ethyl acetate, screw sealable cap on tightly, wrap in parafilm and place in -20 freezer.
-* 5ml glass pipet
-* Nanopure H20
-* 0.2m HCl
-* Chloroform
-===Protocol===