JPH0259942B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a biological sample pretreatment device for a scanning electron microscope.

To observe a biological sample with a scanning electron microscope (hereinafter referred to as a scanning electron microscope), it is necessary to
Secondary electrons are irradiated to generate secondary electrons with an energy of 100 eV or less, and these generated secondary electrons are used as an image information source, but since the primary electrons are charged on the sample surface, causes a defect, and 2.
High resolution cannot be obtained because secondary electrons are difficult to emit.

For this reason, conventionally, after fixing a biological sample and dehydrating and drying it, metal atoms such as gold are deposited on the surface to make it easier to emit secondary electrons, prevent charging, and obtain high resolution. However, when metal is deposited to a thickness of several hundred angstroms on the surface of a sample, the situation is similar to observing rocks on a riverbank with a thin layer of snow, making it impossible to accurately observe minute details. In addition, when using gold or platinum for vapor deposition, it is expensive, and when using tungsten, the emissivity of secondary electrons is high and high resolution can be obtained, but due to its high melting point, biological samples that are sensitive to heat may be Easily altered by heat.

It is an object of the present invention to provide a biological sample pretreatment device that eliminates these problems and enables correct observation of minute parts of a biological sample.

The present invention provides a plasma discharge chamber that has an inert gas inlet and an exhaust port, can maintain a constant pressure inside the container, and has a capacitively coupled electrode connected to a high frequency power source on the outside; A plasma extraction chamber that extends in the axial direction of the discharge chamber and has a plasma extraction electrode; and a plasma extraction chamber that is removably attached to the end of the plasma extraction chamber, and that allows the surface of a biological sample placed inside the plasma extraction chamber to be introduced through the plasma extraction chamber. The first feature is that the processing chamber is irradiated with an inert gas plasma, and the second feature is that it has an inductively coupled electrode in place of the capacitively coupled electrode.

That is, the present invention is provided with an inlet through which an inert gas such as argon gas flows at a constant flow rate, and an exhaust port through which inert gas such as argon gas can be discharged at a constant rate, so that the inside of the container can be maintained at a constant pressure of, for example, 0.3 to 5 Torr. A process chamber made of glass, for example, which is removably attached to one end of the discharge chamber through a rubber gasket, and a capacitive or inductive coupling connected to a high-frequency power source outside the discharge chamber. A biological sample is placed inside the processing chamber, and a part of the inert gas plasma generated by the discharge due to high frequency energy is extracted and processed by the extraction electrode. This device can be introduced indoors and irradiated the surface of biological samples.
Part of the plasma generated in the discharge chamber is introduced into the removable processing chamber by an extraction electrode, so the sample is separated from the heat source and
In addition, it has become possible to irradiate the sample with low-temperature plasma, making it possible to observe minute parts using a scanning electron microscope without depositing metal on the biological sample.

Examples will be described below.

FIG. 1 is a perspective view of one embodiment, in which 1 is a glass discharge chamber provided with an inert gas inlet 2 and an exhaust port 3, and 4 is a rubber gasket 5 at the lower end of the discharge chamber 1. 71 and 72 are a pair of semi-cylindrical capacitively coupled electrodes attached along the outside of the discharge chamber 1, and 8 is a capacitor. Combined electrode 71
and a high frequency power supply connected to 72, 9 is the discharge chamber 1
The lower plasma extraction chamber 10 is a ring-shaped plasma extraction electrode wound around the plasma extraction chamber 9, and is connected to the ground side of the high frequency power source 8.

In order to pre-process a biological sample using the biological sample pre-processing device of this embodiment, after installing the biological sample 6 in the removed processing chamber 4, the processing chamber 4 is connected to the plasma extraction chamber 9 through the packing 5. When the pressure is reduced from the exhaust port 3 using an oil rotary vacuum pump or the like, the processing chamber 4 is adsorbed to the gasket 5 due to the pressure difference between the atmospheric pressure outside the room and the reduced internal pressure. Next, when an inert gas such as argon is flowed in from the inert gas inlet 2 at a rate of 10 ml/min to 150 ml/min, the inside of the discharge chamber 1 is maintained at 0.3 to 5 Torr. Here, when a high frequency voltage is applied to the capacitively coupled electrodes 71 and 72 by the high frequency power supply 8, an inert gas plasma is generated, and a part of this plasma is transferred by the plasma extraction electrode 10 of the plasma extraction chamber 9. It is taken out and jumped into the processing chamber 4, and the surface of the biological sample 6 in the processing chamber 4 is irradiated. At this time, the biological sample 6 is atomized by the ions of the inert gas, creating a surface on which secondary electrons can easily be discharged. It becomes possible to observe minute parts of the sample.

Figures 2 and 3 are scanning electron micrographs of rat tracheal mucosal epithelial cells (magnification: ×6700 ×
12500), the degree of vacuum was 2 to 1 Torr, the frequency output of the high frequency power supply was 3 to 5 W, and the processing time was 10 minutes. The surface of the sample was treated to observe the fine structure of cells that make up the mucosal epithelium. can do.

FIG. 4 is a perspective view of another embodiment, in which the same parts as in FIG. 1 are given the same reference numerals. This device differs from the device shown in FIG. 1 in that a coiled inductively coupled electrode is used as the electrode provided around the outer circumference of the discharge chamber 1. It operates in the same manner as the embodiment shown in the figure, and the same effects can be obtained.

In this way, the biological sample pretreatment device of the example
Since the processing chamber where the sample is installed is connected to the discharge chamber via the plasma extraction chamber, the surface of the sample can be irradiated with low-temperature plasma without altering the biological sample due to high-temperature heat. Therefore, even without metal vapor deposition, primary electrons do not charge the biological sample during observation with a scanning electron microscope, and secondary electrons are easily generated, making it possible to obtain high-resolution images. .

As described above, the biological sample pretreatment device for a scanning electron microscope of the present invention makes it possible to provide a pretreatment device that enables accurate observation of minute parts of biological samples, and has great industrial effects. It is what it is.

[Brief explanation of drawings]

FIG. 1 is a perspective view of an embodiment of the biological sample pretreatment device for a scanning electron microscope of the present invention, and FIGS. 2 and 3 are scans of a biological sample processed using the pretreatment device of FIG. 1. FIG. 4 is a perspective view of another embodiment of the pretreatment apparatus of the present invention. 1... Discharge chamber, 2... (Inert gas) inlet, 3... Outlet, 4... Processing chamber, 5... Packing, 6... Biological sample, 71, 72... Capacitively coupled electrode , 8... High frequency power supply, 9... Plasma extraction chamber, 10... Plasma extraction electrode, 11...
Inductively coupled electrode.

Claims (1)

[Claims] 1. A plasma discharge chamber having an inert gas inlet and an exhaust port, capable of maintaining a constant pressure inside the container, and having a capacitively coupled electrode connected to a high frequency power source on the outside. , a plasma extraction chamber that extends in the axial direction of the plasma discharge chamber and has a plasma extraction electrode; and a plasma extraction chamber that is detachably attached to the end of the plasma extraction chamber, and that connects the surface of a biological sample placed inside the plasma extraction chamber through the plasma extraction chamber. A biological sample pretreatment device for a scanning electron microscope, comprising a processing chamber that is irradiated with plasma of the inert gas introduced by the plasma. 2. A plasma discharge chamber having an inert gas inlet and an exhaust port, capable of maintaining a constant pressure inside the container, and having an inductively coupled electrode connected to a high frequency power source on the outside; A plasma extraction chamber that extends in the axial direction and has a plasma extraction electrode; and a plasma extraction chamber that is removably attached to the end of the plasma extraction chamber, and that allows the surface of the biological sample placed inside the plasma extraction chamber to be exposed to the plasma that is introduced through the plasma extraction chamber. A biological sample pretreatment device for a scanning electron microscope, comprising a treatment chamber for irradiation with active gas plasma.