Scanning Ion Conductance Microscopy

Abstract

The subject invention concerns methods for interrogating a surface using scanning ion conductance microscopy (SICM). In one embodiment, a method of the invention comprises the steps of: a) repeatedly bringing a SICM probe into proximity with the surface at discrete, spaced locations in a region of the surface and measuring surface height at each location; b) estimating surface roughness or other characteristic for the region based upon the surface height measurements; and c) repeatedly bringing the probe into proximity with the surface at discrete, spaced locations in the region, the number and location of which is based upon the estimated surface roughness or other characteristic in the region, and obtaining an image of the region with a resolution adapted to the surface roughness or other characteristic.

1 . A method for interrogating a surface using scanning ion conductance microscopy (SICM), comprising the steps of:
 a) repeatedly bringing a SICM probe into proximity with the surface at discrete, spaced locations in a region of the surface and measuring surface height at each location;   b) estimating surface roughness or other surface characteristic for the region based upon the surface height measurements; and   c) repeatedly bringing the probe into proximity with the surface at discrete, spaced locations in the region, the number and location of which is based upon the estimated surface roughness or other surface characteristic in the region, and obtaining an image of the region with a resolution adapted to the surface roughness or other surface characteristic.   
     
     
         2 . The method according to  claim 1 , wherein steps b) and c) are repeated recursively for sub-regions according to the required image resolution. 
     
     
         3 . The method according to  claim 1 , wherein the step of bringing the probe into proximity with the surface at each location is performed by approaching each location from a distance greater than the height of the surface at that location. 
     
     
         4 . The method according to  claim 1 , wherein lateral movement of the probe occurs only when the probe is distant from the surface. 
     
     
         5 . The method according to  claim 1 , wherein, during the step of bringing the scanning probe into proximity with the surface, the approach is terminated when a measured probe current reaches a threshold value. 
     
     
         6 . The method according to  claim 5 , wherein the threshold value is based upon the probe current measured when the probe is distant from the surface. 
     
     
         7 . The method according to  claim 5 , wherein the approach is terminated when probe current is reduced by 0.25% to 1%. 
     
     
         8 . The method according to  claim 6 , wherein for each measurement, the distance travelled by the probe, from the position distant from the surface to the position at the threshold value, is greater than 1 μm. 
     
     
         9 . The method according to  claim 1 , wherein, during the step of bringing the scanning probe into proximity with the surface, the approach rate or speed is constant. 
     
     
         10 . The method according to  claim 1 , wherein a local relationship between probe current and distance from probe to surface is determined for each measurement location. 
     
     
         11 . The method according to  claim 1 , wherein a differential map of the surface is obtained by obtaining a scanning measurement when the probe is distant from the surface and when the probe is in proximity to the surface and subtracting the second measurement from the first, to obtain the differential map. 
     
     
         12 . The method according to  claim 11 , wherein an agent or other stimulus is applied at the tip of the probe and measurements of response to the agent or stimulus is made at positions distant and proximal to the surface, wherein subtraction of the second measurement from the first provides a differential map of the surface. 
     
     
         13 . The method according to  claim 11 , carried out in the presence of a fluorophore which is activated by a surface structure, wherein a laser beam is focused at the tip of the probe to measure fluorescence, wherein a scanning measurement is obtained together with a fluorescence measurement at positions distant and proximal to the surface and wherein subtraction of the second fluorescence measurement from the first reveals local changes in fluorescence. 
     
     
         14 . The method according to  claim 11 , wherein, during the step of bringing the probe into proximity with the surface, the approach is terminated, and an image is obtained, at different threshold values. 
     
     
         15 . The method according to  claim 14 , wherein the approach is terminated when probe current is reduced by 1%, 5% and 10%. 
     
     
         16 . The method according to  claim 1 , wherein steps (b) and (c) are carried out using estimated surface roughness. 
     
     
         17 . The method according to  claim 1 , wherein steps (b) and (c) are carried out by measuring the presence of a fluorescence signal. 
     
     
         18 . The method according to  claim 5 , wherein an image is obtained at multiple different threshold values for probe current, where differences in the results obtained provide information on the mechanical properties of the surface or reveals information on structures underneath the surface. 
     
     
         19 . The method according to  claim 1 , wherein step (c) is carried out at different voltages, where differences in the results obtained provide information on the mechanical properties of the surface or reveals information on structures underneath the surface. 
     
     
         20 . An apparatus for carrying out scanning probe microscopy, comprising:
 (i) a scanning probe;   (ii) means for measuring and/or controlling the distance of the tip of the probe from a surface to be scanned;   (iii) means for moving the probe laterally relative to the surface.   
       wherein the means for measuring and/or controlling the distance of the tip of the probe comprises two piezo actuators of different response times. 
     
     
         21 . The apparatus according to  claim 20 , which is a scanning ion conductance microscope. 
     
     
         22 . The apparatus according to  claim 20 , wherein the first piezo actuator has a travel range of at least 100 μm, and the second piezo-actuator has a travel range less than 50 μm. 
     
     
         23 . The apparatus according to  claim 20 , wherein the second piezo-actuator has a travel range of no more than 25 μm.