Lipid Coated Bead Preparation

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Contents

Protocol

Lipid Preparation:
  1. In a glass container, and using lipids dissolved in chloroform, mix desired lipids to a final quantity of ~4mmol total lipid.
  2. Dry lipids under inert gas.
  3. Suspend lipids in 400ml PBS or other buffer, sonicate to suspend completely.
  • Note: if covered with inert gas, these lipids are stable for ~1 week at 4oC.
Glass Bead Preparation:
  1. Measure out 2mg of glass beads of the desired diameter.
  2. Wash 3X in 1ml ddH2O.
  3. Pellet glass beads. Remove as much water as possible.
  4. Suspend beads in 400ml of the PBS-lipid solution, rotate at room temp 20min to allow bylayers to self-assemble.
  5. Pellet beads with a brief (<5sec) cetrifugation and remove the liquid phase. This will remove unincorporated liposomes.
  6. Suspend beads in 200-400ml desired buffer or in ddH2O.
Laxex Bead Preparation:
  1. Measure out 10ml latex beads of the desired diameter.
  2. Wash 2X in 50mM carbonate-bicarbonate buffer, pH 9.6. Pellet cells with a max-speed spin in minifuge, 10min.
  3. Incubate 2hrs at 37oC, rotating constantly, with lipisome fractions prepared in “lipid preparation”.
  4. Wash 2X with carbonate-bicarbonate buffer
  5. Incubate with 5% BSA in PBS for 2+ hours to block beads.

Recipes

Carbonate-bicarbonate buffer:
  • 1.15g Anhydrous sodium carbonate
  • 4.13g Sodium bicarbonate
  • 100ml ddH2O
    Dissolve in water, pH to 9.6.


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