Ultrastable Atomic Force Microscopy for Biophysics

<p>Atomic force microscopy (AFM) is a multifunctional workhorse of nanoscience and molecular\&nbsp;<span style="font-size: 13px; line-height: 1.6em;">biophysics, but instrumental drift remains a critical issue that limits the precision and duration\&nbsp;</span><span style="font-size: 13px; line-height: 1.6em;">of experiments. We have signicantly reduced the two most important types of drift: in position\&nbsp;</span><span style="font-size: 13px; line-height: 1.6em;">and in force. The first, position drift, is defined as uncontrolled motion between the tip and the\&nbsp;sample, which occurs in all three dimensions. By scattering a laser off the apex of a commercial\&nbsp;AFM tip, we locally measured and thereby actively controlled its three-dimensional position above\&nbsp;a sample surface to \&lt;0.4\r A(Δf = 0.01\textendash10 Hz) in air at room temperature. With this enhanced\&nbsp;stability, we demonstrated atomic-scale (~1 \r A) tip-sample stability and registration over tens of\&nbsp;minutes with a series of AFM images. The second type of drift is force drift. We found that the\&nbsp;primary source of force drift for a popular class of soft cantilevers is their gold coating, even though\&nbsp;they are coated on both sides to minimize drift. When the gold coating was removed through\&nbsp;a simple chemical etch, this drift in defl</span><span style="font-size: 13px; line-height: 1.6em;">ection was reduced by more than an order of magnitude\&nbsp;</span><span style="font-size: 13px; line-height: 1.6em;">over the first 2 hours after wetting the tip. Removing the gold also led to 10-fold reduction in\&nbsp;</span><span style="font-size: 13px; line-height: 1.6em;">refl</span><span style="font-size: 13px; line-height: 1.6em;">ected light, yet short-term (0.1\textendash10 s) force precision improved. With both position and force\&nbsp;</span><span style="font-size: 13px; line-height: 1.6em;">drift greatly reduced, the utility of the AFM is enhanced. These improvements led to several new\&nbsp;</span><span style="font-size: 13px; line-height: 1.6em;">AFM abilities, including a five-fold increase in the image signal-to-noise ratio; tip-registered, label-free\&nbsp;</span><span style="font-size: 13px; line-height: 1.6em;">optical imaging; registered tip return to a particular point on the sample; and dual-detection\&nbsp;</span><span style="font-size: 13px; line-height: 1.6em;">force spectroscopy, which enables a new extension clamp mode. We have applied these abilities\&nbsp;</span><span style="font-size: 13px; line-height: 1.6em;">to folding of both membrane and soluble proteins. In principle, the techniques we describe can\&nbsp;</span><span style="font-size: 13px; line-height: 1.6em;">be fully incorporated into many types of scanning probe microscopy, making this work a general\&nbsp;</span><span style="font-size: 13px; line-height: 1.6em;">improvement to scanning probe techniques.</span></p>
Year of Publication
Ph. D.
Number of Pages
Date Published
University of Colorado Boulder
Boulder, CO
Advisors - JILA Fellows