Atomic force microscopy imaging of a supported bilayer of the phospholipid DPPC revealed the presence of thin lines, which were thought to be the boundaries of domains with a different orientation. Temperature-controlled AFM showed that the melting from the gel state L b 0 to the fluid L a phase starts on these lines. The observed onset of melting at 40.3 1C compared well to reported DSC measurements.The same mechanism of melting was observed in computer simulations on a bilayer of a coarse-grained lipid model at the L b 0 phase. Two boundary lines were present in the initial configuration. It was shown that the lipid acyl tails became more disordered at the boundaries upon temperature increase. r 2004 Published by Elsevier B.V.

Temperature-controlled Atomic Force Microscopy (TC-AFM) in Contact Mode is used here to directly image the mechanisms by which melting and crystallization of supported, hydrated DPPC bilayers proceed in the presence and absence of the model peptide WALP23. Melting from the gel L(β)' to the liquid-crystalline L(α) phase starts at pre-existing line-type packing defects (grain boundaries) in absence of the peptide. The exact transition temperature is shown to be influenced by the magnitude of the force exerted by the AFM probe on the bilayer, but is higher than the main transition temperature of non-supported DPPC vesicles in all cases due to bilayer-substrate interactions. Cooling of the fluid L(α) bilayer shows the formation of the line-type defects at the borders between different gel-phase regions that originate from different nuclei. The number of these defects depends directly on the rate of cooling through the transition, as predicted by classical nucleation theory. The presence of the transmembrane, synthetic model peptide WALP23 is known to give rise to heterogeneity in the bilayer as microdomains with a striped appearance are formed in the DPPC bilayer. This striated phase consists of alternating lines of lipids and peptide. It is shown here that melting starts with the peptide-associated lipids in the domains, whose melting temperature is lowered by 0.8-2.0°C compared to the remaining, peptide-free parts of the bilayer. The stabilization of the fluid phase is ascribed to adaptations of the lipids to the shorter peptide. The lipids not associated with the peptide melt at the same temperature as those in the pure DPPC supported bilayer.

Infrared absorption spectroscopy of lipid layers was performed by combining optics and scanning probe microscopy. This experimental approach enables sub-diffraction IR imaging with a spatial resolution on the nanometer scale of 1, 2-dioleoyl-sn-glycero-3-phosphocholine lipid layers.

Sub diffraction limited infrared absorption imaging of hemoglobin was performed by coupling IR optics with an atomic force microscope. Comparisons between the AFM topography and IR absorption images of micron sized hemoglobin features are presented, along with nanoscale IR spectroscopic analysis of the metalloprotein.

IR absorption spectroscopy of hemoglobin was performed using an IR optical parametric oscillator laser and a commercial atomic force microscope. This experimental approach enables detection of protein samples with a resolution that is much higher than that of standard IR spectroscopy. Presented here are AFM based IR absorption spectra and images of micron sized hemoglobin features. IntroductionAbsorption spectroscopy is a widely applied technique for chemical characterisation. This method is able to detect both luminescent and non-luminescent materials and provide chemical specific information. An extensively used form of absorption spectroscopy is infrared absorption (IR) spectroscopy. This measures specific frequencies in the infrared region of the electromagnetic spectrum at which constituent parts of molecules corresponding to specific types of molecular bonds vibrate. This makes possible structural elucidation and compound identification of materials. As a consequence, IR absorption is extensively used as an analytical tool.