Difference between revisions of "HighPoint/CannonLab:Lipid Vesicle Preparation"

From OpenWetWare
Jump to: navigation, search
(New page: {{CannonLabMenu}} ==Lipid Film Preparation== ===Materials=== * CHCl<sub>3</sub> * MeOH * dry lipids * screw top glass vial with teflon seal ===Procedure=== # Make 8mL of 3:1 CHCl3:MeO...)
(No difference)

Revision as of 08:50, 16 February 2012

<owwmenu font="arial, helvetica, sans-serif" bold="1"

       color="white" bgcolor="indigo" hovercolor="white"
       bghovercolor="gray" topfontsize="10" fontSize="8" pagewidth="750"
       image="CannonLabIMG01.JPG" lab="MGSC/CannonLab">
   Lab Members=#, People=People, Contact=Contact Information, Associates=Collaborators and Former Members
   Research=#,Protein Membrane=Protein Membrane Interactions, Osmolytes=Osmolyte Effects
   Protocols=#, Vesicles=Lipid Vesicle Preparation, Solutions=Buffer and Osmolyte Solutions, Fluorescence=Fluorescence Spectrometry, UV-Vis=UV-Vis Spectrometry, Pipetting=Pipetting Exercise
   Notebooks=#, Master List=Notebook, Dr Cannon=Notebook/Jonathan Cannon
   Links=#, Courses=Course Pages


Lipid Film Preparation


  • CHCl3
  • MeOH
  • dry lipids
  • screw top glass vial with teflon seal


  1. Make 8mL of 3:1 CHCl3:MeOH (6mL CHCl3/2mL MeOH)
  2. Add 2.5mL of CHCl3:MeOH solution to 25 mg lipids
  3. Take 1mL (10 mg lipid) of lipid/CHCl3:MeOH solution and place in 4mL vial (2 dram?)
  4. Dry using rotovap
    • Attach vial to rotovap with a rubber septum (silicone septa only work 2-3 times before shrinking)
    • Insert needle through septum to allow solvent to evaporate
    • Evaporation takes <<30 min if conditions are right
  5. Place vial in vacuum desicator overnight to remove remainder of solvent
  6. Close tightly and store aliquot in freezer

Lipid Film Hydration

  • Prepare the buffer appropriate for your experiments
  • Determine the concentration of lipids you wish to use in your experiments
  • Prepare a hydrated lipid suspension much more concentrated than the desired experimental concentration
    • A concentration of 10 mg/mL is often a good starting point
      1. add 1 mL buffer to 10 mg lipid
      2. tape vial to vortexer and shake for 1 h
      3. store hydrated lipids in freezer until you are ready to make LUVs