Lipidic Cubic Phase (LCP)

LCP

The detailed protocol for crystallizing membrane proteins in LCP was published by Martin Caffrey and Vadim Cherezov in: Crystallizing Membrane Proteins Using Lipidic Mesophases, Nat. Protoc. 2009; 4(5): 706–731.

More information about LCP can be found here:
> Vadim Cherezovs USC web site “The Cherezov Lab”

PCC’s standard procedure for preparing the mesophase follows closely the method described by Martin Caffrey and Vadim Cherezov by mixing two parts protein solution (40%) with three parts monoolein (60%; CAS# [111-03-5]; HR2-435). For this procedure two Hamilton syringes (25µL, 50µL or 100µL Syringe Model 1710 RN) get connected by a LCP syringe coupler (Formulatrix, SKU 209526). The mixing process by moving the syringe plungers back and forth is repeated until the cubic mesophase becomes homogenous, clear and does not show any significant birefringence. The protein sample volume required for this procedure is listed in Sample Volume.

The Gryphon-LCP dispenses a 40nl bolus of the LCP-protein mixture and an excess of 800nl precipitant solution in each of the 96 well of a LCP-sandwich plate. The volume of the LCP bolus range between 20nl – 200nl. The SWISSCI LCP sandwich plate used for all LCP-experiments consists of a hydrophobic SBS sized base plate with a 60um or 120um spacer. The experiments are sealed with a glass- or a Laminex Film (MD11-54) cover. PCC recommends for bolus volumes <50nl the sandwich plates with the 60um spacer and the glass cover. The LCP-plate will be incubated at 20°C in the Rock Imager.