Gernot Hamann presents a combination of methods for high-precision optical, topographic and molecular analyses
In pharmaceutical research, living cell investigations, nanophotonics or analyses in photovoltaics or semiconductor technology, classical microscopic methods are often no longer sufficient in terms of optical resolution or information content. To obtain more extensive and more accurate measurement data of a sample, it is now possible to combine different microscopic methods with one another. However, this combination of methods not only makes high demands on the individual components of the microscopes, but also on the systems used for sample positioning.
For example, piezo-based positioning stages (Fig. 1.) achieve a position resolution in the sub-nanometer range and response times of less than 1ms. With this, dynamic operation at scanning frequencies of up to 1,000Hz is possible. The high dynamics in the Z axis for focusing processes or topography scans allow high-speed scanning of the sample in the X and Y directions. This shortens measuring times, increases throughput and reduces time-dependent impacts on the measurement. Moreover, they can be readily integrated into microscopes.
An application example of piezo-based scanning stages is a microscopic system of modular design, combining a Raman microscope with atomic force microcopy (AFM), see Fig. 2. Raman microscopy is based on a confocal, optical microscope and a Raman spectrometer. The sample is scanned point by point and line by line. The lateral resolution is about 200nm with green excitation light. During the measurement, a complete Raman spectrum is recorded for each pixel. These Raman spectra are like a specific fingerprint for each type of molecule, so that the chemical components of a sample can be identified for each pixel and their distribution in the sample can be displayed.
The scanning stage used for the sample positioning is designed for working distances of 100 or 200µm in the axes of the scanning plane and 20µm in the direction of the Z axis. Position resolution is better than 2nm.
With the AFM method, a measuring tip is moved over the sample surface line by line in a defined grid. It measures the forces between a pointed measuring tip and the surface of the object. It gives information on the topography of the surface at a lateral resolution capacity of 10nm.
Since the distance between the measuring tip and the surface has to be kept constant, the position of the sample must be readjusted in the Z direction. This task is performed by the scanning stage. The variation of the Z position together with the relevant X and Y coordinates for the spatial resolution then provides the high-precision topography information on the samples.
The AFM and Raman images are recorded in succession and then superimposed and related to one another.
Precise positioning in all three axes and path accuracy during the scan are indispensable for achieving high-quality images.
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Gernot Hamann is business development manager for microscopy at PI (Physik Instrumente), based in Karlsruhe, Germany.