The Intrinsic Resolution Limit in the Atomic Force Microscope: Implications for Heights of Nano-Scale Features

Abstract

BackgroundAccurate mechanical characterization by the atomic force microscope at the highest spatial resolution requires that topography is deconvoluted from indentation. The measured height of nanoscale features in the atomic force microscope (AFM) is almost always smaller than the true value, which is often explained away as sample deformation, the formation of salt deposits and/or dehydration. We show that the real height of nano-objects cannot be obtained directly: a result arising as a consequence of the local probe-sample geometry.Methods and FindingsWe have modeled the tip-surface-sample interaction as the sum of the interaction between the tip and the surface and the tip and the sample. We find that the dynamics of the AFM cannot differentiate between differences in force resulting from 1) the chemical and/or mechanical characteristics of the surface or 2) a step in topography due to the size of the sample; once the size of a feature becomes smaller than the effective area of interaction between the AFM tip and sample, the measured height is compromised. This general result is a major contributor to loss of height and can amount to up to ∼90% for nanoscale features. In particular, these very large values in height loss may occur even when there is no sample deformation, and, more generally, height loss does not correlate with sample deformation. DNA and IgG antibodies have been used as model samples where experimental height measurements are shown to closely match the predicted phenomena.ConclusionsBeing able to measure the true height of single nanoscale features is paramount in many nanotechnology applications since phenomena and properties in the nanoscale critically depend on dimensions. Our approach allows accurate predictions for the true height of nanoscale objects and will lead to reliable mechanical characterization at the highest spatial resolution.

Trypanosoma brucei 20 S Editosomes Have One RNA Substrate-binding Site and Execute RNA Unwinding Activity

“…3E). These numbers deviate from the published EM-based measurements (27) and are likely influenced by two phenomena: (i) flattening of the complexes during the AFM analysis (39, 42, 43) and (ii) shrinking of the particles caused by dehydration on the mica surface (44,45). Nevertheless, the dimensions can be used to calculate apparent volumes (V app ) for the complexes, which revealed Gaussian distributions with an average volume of 3-4 ϫ 10 3 nm 3 for the 20 S editosome and 6 -7 ϫ 10 3 nm 3 for the 35-40 S editosome (Fig.…”

The nature of the air-cleaved mica surface

“…The well-defined diameter of the dsDNA molecule could be used as a "molecular dip stick" since the apparent height of the molecule is influenced by the adsorbed water thickness surrounding the DNA [208]. The kinetics of water layer formation were influenced by the incubation time of the mica in Ni 2+ solution (30 s or 5 min) as well as the time the Ni-mica was left at ambient humidity (hours to days).…”

Silk Reconstitution Disrupts Fibroin Self-Assembly

“…37,53 We note that the lateral dimensions of the imaged protein structures are of the same order as the size of the AFM probes used, and consequently, the apparent widths are most likely overestimated and heights underestimated relative to their true values. 52,56 Therefore, precise determinations of dimensions and volumes are very challenging. Nevertheless, relative comparisons to other AFM data, as well as coarse general assessments are feasible and useful.…”