Ion conductance microscopy (ICM) is an extensively used technique to obtain contact-free images of cell surface topography and to image cultured cells under liquid conditions. A new modality of ICM is presented here: the approach-retract scanning (ARS) mode. This modality is highly powerful and enable imaging of samples having steep slopes. The case of dense collagen fibril network studied via ICM is presented in order to test whether the ARS mode is suitable for imaging biological samples with large height gaps.
Collagen fibrils were derived from the tail tendon of adult Wistar rats and stored in physiological saline with 1-10 % tymol (2-isopropyl-5-methylphenol) at 4 °C for a minimum of 1 day. A tendon piece was subjected to stretching and air-dried overnight. The sample was then immersed again in physiological saline. ICM imaging was performed in the ARS mode using the XE-Bio System.
An example of ICM imaging of collagen fibril networks using the ARS mode is shown in figure 1. The width of the individual fibrils ranged from 50 to 470 nm. As per the section profile, the collagen fibril width indicated by the green arrows in figure 1 was 277 nm and the height difference between the fibril and the glass surface was 2606 nm. This result shows that certain fibrils are suspended over the glass substrate during the ICM image scanning. Hence, the ARS mode of ICM has the benefit of minimizing the loading force, which is unavoidable in conventional atomic force microscopy (AFM).
Figure 1. Dense collagen fibril networks imaged by the ARS/hopping ICM (top panel). The profile analysis (bottom panel) refers to the red line indicated in the ICM image.
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