The present invention relates to a sample holder for holding a sample to be observed for research purposes, and more particularly to a sample holder for holding a sample to be observed in an electron microscope, such as a transmission electron microscope (TEM), and which has the capability of delivering and accurately directing a light beam to the sample held by the sample holder and/or collect a light beam and transport it outside the TEM for analysis.
Structural evaluation using an electron microscope has been conventionally employed as one of the methods for examining and evaluating samples in the fields of micro- and nanotechnology. The electron microscopes used in these fields mainly include the scanning electron microscopes (SEM) and the transmission electron microscopes (TEM). In the SEM, a beam of electrons is applied to a cleavage plane or an FIB (Focused Ion Beam) processed plane of the sample being observed (observed sample) and secondary electrons etc. obtained from the sample form an image for observation.
In the TEM, a beam of electrons is transmitted through a very thin, (e.g., 1 μm thick or less), observed sample and transmitted electrons and scattered electrons (e.g., elastically scattered electrons) form an image for observation of the internal structure of the sample. The image formed from the electrons transmitted through the specimen is typically magnified and focused by an objective lens and appears on an imaging screen, (i.e., a fluorescent screen in most TEMs), plus a monitor, or on a layer of photographic film, or to be detected by a sensor such as a CCD camera.
Modern TEMs are often equipped with specimen holders that allow the user to tilt the specimen to a range of angles in order to obtain specific diffraction conditions, and apertures placed above the specimen allow the user to select electrons that would otherwise be diffracted in a particular direction from entering the specimen. By carefully selecting the orientation of the sample, it is possible not just to determine the position of defects but also to determine the type of defect present. If the sample is orientated so that one particular plane is only slightly tilted away from the strongest diffracting angle (known as the Bragg Angle), any distortion of the crystal plane that locally tilts the plane to the Bragg angle will produce particularly strong contrast variations. However, defects that produce only displacement of atoms that do not tilt the crystal to the Bragg angle (i.e. displacements parallel to the crystal plane) will not produce strong contrast.
The TEM is used heavily in both material science/metallurgy and the biological sciences. In both cases the specimens must be very thin and able to withstand the high vacuum present inside the instrument. For biological specimens, the maximum specimen thickness is roughly 1 micrometer. To withstand the instrument vacuum, biological specimens are typically held at liquid nitrogen temperatures after embedding in vitreous ice, or fixated using a negative staining material such as uranyl acetate or by plastic embedding. Typical biological applications include tomographic reconstructions of small cells or thin sections of larger cells and 3-D reconstructions of individual molecules via Single Particle Reconstruction.
In material science/metallurgy the specimens tend to be naturally resistant to vacuum, but must be prepared as a thin foil, or etched so some portion of the specimen is thin enough for the beam to penetrate. Preparation techniques to obtain an electron transparent region include ion beam milling and wedge polishing. The focused ion beam (FIB) is a relatively new technique to prepare thin samples for TEM examination from larger specimens. Because the FIB can be used to micro-machine samples very precisely, it is possible to mill very thin membranes from a specific area of a sample, such as a semiconductor or metal. Materials that have dimensions small enough to be electron transparent, such as powders or nanotubes, can be quickly produced by the deposition of a dilute sample containing the specimen onto support grids. The suspension is normally a volatile solvent, such as ethanol, ensuring that the solvent rapidly evaporates allowing a sample that can be rapidly analyzed.
In certain applications, analysis of a sample subjected to light is desirable. Specifically, it is often desirable to analyze the optical properties of a sample under light conditions within a TEM. In this regard, attempts have been made to modify conventional TEMs by providing a window to the TEM housing to allow light from an external source to enter the interior chamber of the TEM in the area of the sample. Thus, prior solutions have involved modifications of the TEM column to provide an optical path to the sample position. As can be appreciated, such solutions are very complicated and expensive and involve major modifications of the microscope column.