Scanning Ion Conductance Microscopy

2.1.1 Principles

A single-channel nanopipette (typically 10’s nm opening) filled with electrolyte and equipped with a Ag/AgCl quasi-reference counter electrode (QRCE) is approached towards a surface in a bath of electrolyte solution containing a second QRCE. Then, an ion current is generated between both electrodes when a bias voltage is applied. The magnitude of the ion current is sensitive to the probe-surface distance, the nature of the surface and the voltage applied between both QRCEs, changing from the bulk value when the probe-surface distance is less than one tip diameter.105-107 This change in the ion current is used as the feedback signal to detect the surface, such that topographical and functional maps can be obtained without establishing physical probe-surface contact.94,95,108,109

2.1.2 Feedback Types and Scanning Modes

SICM can operate in various modes that use different feedback signals to probe a surface. SICM feedback modes include: (i) direct current (DC) mode,105 _{in which a}

constant bias is applied between both QRCEs. The ion current decreases rapidly as a function of probe-surface distance as a result of the increased resistance in the region at the end of the nanopipette. This current drop is used as a feedback signal. (ii) Distance modulation (DM) mode,110 which involves vertically oscillating the probe whilst applying a fixed bias, generating an alternating current (AC) signal upon approach to the surface. The AC signal is more stable than the DC signal and can be described by both its amplitude and phase angle component, with respect to the applied oscillation of the voltage. In this mode, an increase in the AC amplitude is used as feedback. (iii)

Bias modulation (BM) mode,111 _{in which a harmonically oscillating bias between both}

QRCEs is applied to induce an AC signal. In this case, both a decrease in the AC amplitude and an increase in the AC phase can be used for feedback. A schematic illustration of the different feedback types used in SICM is shown in Figure 1.11. BM- SICM was first demonstrated relatively recent and has several advantages over the traditional DC-SICM and DM-SICM modes that make it more suitable for multifunctional imaging.75,94,111

Figure 1.11 Comparison between different modes of SICM feedback. (a) Direct current mode in which the surface is detected via a drop in ionic current. (b) Distance modulation mode, in which the probe is oscillated in the vertical direction to produce an AC signal at small tip– substrate separation. (c) Bias modulation mode, in which a sinusoidal bias is applied between the two electrodes and the AC phase is monitored for feedback. Illustrative approach curves are shown in the insets on the images, revealing the feedback response as a function of the relative probe-to-substrate distance d/r (physical probe–substrate separation, d, and probe opening radius, r). In b and c, “set” indicates a set-point value for a particular quantity that is used to stop the tip moving closer to the surface and can be used for feedback during imaging. Reproduced with permission from reference 72. Copyright (2016) American Chemical Society.

SICM maps can also be acquired using different scanning modes. Constant distance

and hopping mode are the most commonly used in SICM imaging. In constant distance mode, a constant DC or AC signal is maintained by a feedback loop that moves the probe vertically while it scans laterally.105,112 By maintaining a constant DC or AC signal, a constant probe-surface distance is maintained and a topographical image is formed by tracking the Z-position throughout the scan (Figure 1.12a). Although constant distance mode has proven useful for a wide range of applications, it is difficult to image complex surface features of high aspect ratio.113

On the other hand, hopping mode consist of a series of approach curves to determine the surface topography. The probe approaches the surface to a certain probe-surface distance determined by the value of the DC or AC feedback threshold and the Z- position is recorded.114 Then, the probe is withdrawn and moved to a new location before starting a new approach (Figure 1.12b). The number of data points and the retraction distance determines the image resolution and the imaging time. Generally, constant distance enables faster scan rates than hopping mode, but hopping mode allows high-resolution imaging of more complex features.

Figure 1.12. Trajectory of the SICM probe during (a) a constant distance scan and (b) a hopping mode scan.

2.1.3 Applications

Since SICM was introduced in 1989 by Hansma et al.,105 it has been used primarily in molecular biology and materials science for high resolution topographical imaging of soft samples such as living cells.113-115 An important aspect of this technique is that allows the characterization of mechanical responses or morphological transformations of cells caused by external stimuli.110 SICM is sensitive to local heterogeneities in the ionic atmosphere, and has been used to study the transport properties of ion channels in cells and porous membranes via conductance measurements.116 _{Moreover, the}

nanopipette probe has been used as a delivery system to introduce materials to localized areas of a substrate such as DNA117 _{and fluorescent dyes}118 _{for further analysis.}

In the last five years, the technique has gone beyond these applications as a result of a better understanding of the ion flux processes and electric effects that constitute the current response,106,107,119 enabling the simultaneous topographical and functional analysis of surfaces.95 This new capability has been used to establish relationships between surface structure and surface charge in cells108,109 and obtain topography- reactivity maps of UMEs.94

Although it has not been practically applied to the crystal growth and dissolution field, only one study has been found in the literature,89 there are significant prospects for this technique to constitute a suitable tool for the investigation of the mechanism and kinetics of crystal dissolution. SICM constitutes a powerful means of probing dynamic processes at interfaces, particularly when correlating changes in the local ion concentration with variations in the surface topography.