Abstract
The efficacy of a number of different methods for depositing a dimyristoylphosphatidylcholine (DMPC) lipid bilayer or DMPC–cholesterol (3 : 1) mixed bilayer onto a silicon substrate has been investigated in a quantitative manner using atomic force microscopy (AFM) image analysis to extract surface coverage. Complementary AFM-IR measurements were used to confirm the presence of the lipids. For the Langmuir–Blodgett/Schaefer deposition method at temperatures below the chain-melting transition temperature (Tm), a large number of bilayer defects resulted when DMPC was deposited from a water subphase. Addition of calcium ions to the trough led to smaller, more frequent defects, whereas addition of cholesterol to the lipid mixture led to a vast improvement in bilayer coverage. Poor coverage was achieved for deposition at temperatures above Tm. Formation of the deposited bilayer from vesicle fusion proved a more reliable method for all systems, with formation of near-complete bilayers within 60 seconds at temperatures above Tm, although this method led to a higher probability of multilayer formation and rougher bilayer surfaces.
Highlights
We present a quantitative approach for determining the coverage extent for lipid bilayers formed on silicon surfaces using atomic force microscopy (AFM) and on gold using AFM-IR
The monolayer of pure DMPC at 18 C collapsed at 53 mN mÀ1 at an area per lipid molecule of $37 A2 and extrapolation of the steeper portion of the curve to the abscissa gives a limiting area of 49 A2, in good agreement with previously reported values[36,37]
The addition of cholesterol to the DMPC lipid solution (3 : 1 DMPC : cholesterol) signi cantly decreased the area per molecule, to $27 A2 at the point of collapse, with a limiting area of 34 A2; this, and the loss of the expanded-condensed phase transition, indicates that the cholesterol causes a change in packing structure
Summary
Solid-supported lipid bilayers are widely used for studying fragile model cell membrane structures using techniques such as neutron re ectometry,[1,2,3,4,5] X-ray photoelectron spectroscopy,[6] quartz crystal microbalance methods,[7,8,9] surface-based vibrational spectroscopic techniques,[10,11,12,13,14,15] atomic force microscopy[16,17,18,19] (AFM) and electrochemical measurements,[20,21,22,23] amongst others. For some systems it can be difficult to achieve full bilayer coverage, with patches of the substrate surface remaining exposed. This can be disadvantageous when attempting to extract values for parameters such as the bilayer resistance or capacitance. A thicker lm is o en seen for deposited bilayers of DMPC that include cholesterol than for pure DMPC bilayers, arising from more efficient headgroup packing and a lower chain tilt angle.[29]
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