Detecting elusive surface atoms with atomic force microscopy.

Future development of proteomics may be hindered by limitations in the concentration sensitivity of widespread technological approaches. The concentration sensitivity limit (CSL) of currently used approaches, like 2-DE/LC separation coupled with MS detection, etc., varies from 10(-9) to 10(-12) M. Therefore, proteomic technologies enable detection of up to 20% of the protein species present in the plasma. New technologies, like atomic force microscopy (AFM molecular detector), enable the counting of single molecules, whereas biospecific fishing can be used to capture these molecules from the biomaterial. At the same time, fishing also has thermodynamic limitations due to the reversibility of the binding. In cases where the fishing becomes irreversible, its combination with an AFM detector enables the registration of single protein molecules, and that opens up a way to lower the CSL down to the reverse Avogadro number.

Microscopy, Atomic Force↗

PubMed2007

Single molecule studies of protein folding using atomic force microscopy.

Atomic force microscopy (AFM) offers new insights into the ability of proteins to resist mechanical force. The technique has been opened up by the availability of easy-to-use instruments that are commercially available, so that the technique no longer relies on the need to build instruments in the lab. Indeed it may become common for AFM instruments to sit beside stopped-flow apparatus in protein folding laboratories. In this chapter, we describe the instrument set-up, the preparation of suitable protein substrate, and the collection of data. Data selection and analysis are more complex than for conventional stopped-flow ensemble studies, but offer new insights into the function of proteins in vivo.

Microscopy, Atomic Force↗

PubMed2006

Deposition of DNA rafts on cationic SAMs on silicon [100].

We demonstrate a guided self-assembly approach to the fabrication of DNA nanostructures on silicon substrates. DNA oligonucleotides self-assemble into "rafts" 8 x 37 x 2 nm in size. The rafts bind to cationic SAMs on silicon wafers. Electron-beam lithography of a thin poly(methyl methacrylate) (PMMA) resist layer was used to define trenches, and (3-aminopropyl)triethoxysilane (APTES), a cationic SAM precursor, was deposited from aqueous solution onto the exposed silicon dioxide at the trench bottoms. The remaining PMMA can be cleanly stripped off with dichloromethane, leaving APTES layers 0.7-1.2 nm in thickness and 110 nm in width. DNA rafts bind selectively to the resulting APTES stripes. The coverage of DNA rafts on adjacent areas of silicon dioxide is 20 times lower than on the APTES stripes. The topographic features of the rafts, measured by AFM, are identical to those of rafts deposited on wide-area SAMs. Binding to the APTES stripes appears to be very strong as indicated by "jamming" of the rafts at a saturation coverage of 42% and the stability to repeated AFM scanning in air.

Microscopy, Atomic Force↗