HighPoint/CannonLab:Lipid Vesicle Preparation: Difference between revisions

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(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...)
 
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{{CannonLabMenu}}
{{CannonLabMenu}}
Avanti Polar Lipids has detailed protocols and information on lipids and lipid handling
[http://avantilipids.com/index.php?option=com_content&view=article&id=1383&Itemid=371 Liposome Preparation]


==Lipid Film Preparation==
==Lipid Film Preparation==
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**# tape vial to vortexer and shake for 1 h
**# tape vial to vortexer and shake for 1 h
**# store hydrated lipids in freezer until you are ready to make LUVs
**# store hydrated lipids in freezer until you are ready to make LUVs
==Large Unilamellar Vesicle Extrusion==
Read through the following links on the Avanti Polar Lipids website
* [http://avantilipids.com/index.php?option=com_content&view=article&id=1384&Itemid=372 Liposomes]
* [http://avantilipids.com/index.php?option=com_content&view=article&id=1600&Itemid=381 LUVET]
''Follow these protocols carefully and you will be successful at producing LUV''

Latest revision as of 15:25, 22 February 2012

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Avanti Polar Lipids has detailed protocols and information on lipids and lipid handling

Liposome Preparation

Lipid Film Preparation

Materials

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

Procedure

  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

Large Unilamellar Vesicle Extrusion

Read through the following links on the Avanti Polar Lipids website

Follow these protocols carefully and you will be successful at producing LUV

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