User:Samiye Yaman/Notebook/Lab 581/2013/03/20

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Part A: Prepare tris-HCl Buffer

  • 25mmol/L x 1mol/1000mmmol x 1L/1000mL x 300mL x 121.14g/mol

= 0.90855g tris

  • Add 250ml of distilled water to the beaker with 0.9094g of tris.
  • Add HCl drop by drop until the pH was at 8.07.
  • 47ml of more distilled water was added to have 300ml of 25mM tris-HCl buffer.

Part B: Preparing hydrogels for liposome coating

  • 100mg of transfection reagent 1 was dissolved in 5ml choloroform.
  • 1 ml of the choloroform with transfection reagent 1 was placed under the hood and let it evaporate.
  • After the evaporation of the choloroform, 1 ml of phosphate buffer was added to the vial.
  • 3 freeze-thaw vortex cycles were applied to the vial in order to create multi lamellar vesicles. (30sec freeze with liquid nitrogen, 30 sec thaw in water bath at room temperature, and vortex for 30sec)
  • By using Nuclepore Track-Etched Membrane, multi lamellar were converted to uni lamellar vesicles.
  • Add 4mg lipid with 2 mg hydrogels.(1. 0.5g PVOH/3mL buffer/6μL R6G/50mL oil 2. 0.5g PVOH/4mL buffer/35mL oil)
  • Gently stir for 2 hours.