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Kado, Masataka; Kishimoto, Maki; Tamotsu, Satoshi*; Yasuda, Keiko*; Aoyama, Masato*; Tone, Shigenobu*; Shinohara, Kunio*
AIP Conference Proceedings 1696, p.020019_1 - 020019_4, 2016/01
Times Cited Count：3 Percentile：85.37(Microscopy)Soft X-ray microscope is a very powerful tool to observe cellular organelles of living biological cells and many works have demonstrated imaging of inner structures of the cells. However the inner structures are very complicated and it is difficult to identify the organelles obtained with the soft X-ray microscopes. We have proposed a hybrid imaging method with a soft X-ray microscope and a fluorescence microscope that is to observe the same biological cells with the both microscopes at the same time. Using the information of the cellular organelles obtained with the fluorescence microscope, inner structures obtained with the soft X-ray microscope are accurately identified. We have observed living biological cells by the hybrid imaging method. Since the soft X-ray microscope has higher spatial resolution than that of the fluorescence microscope, fine structures of the cellular organelles in the living biological cells were discussed.
Koike, Masato
Heisei 9nenndo - Heisei 12 nendo Kagaku Kennkyuhi Hojyokin, Kenkyu Seika Houkokusho (Kenkyu Kadai Bangou 09309001), p.70 - 76, 2001/03
no abstracts in English
Kado, Masataka; Ishino, Masahiko; Kishimoto, Maki; Tamotsu, Satoshi*; Yasuda, Keiko*; Shinohara, Kunio*
no journal, ,
Intense soft X-ray emissions are very useful for X-ray microscopy to observe living biological specimens. Increasing the photon flux in the range of wavelength 2.3 nm to 4.4 nm, so-called "water window", is a key issue for the soft X-ray microscopy. We have proposed to use ultra-thin foiled targets to limit the target mass to be heated by an intense pulsed laser. Thin foiled gold targets with various thicknesses from 10 nm to 50 
m were irradiated with an intense laser pulse at the wavelength of 1.053 m generated from a Nd:glass laser system with 20 J output energy in 600 ps pulse duration. The soft X-ray emissions from the laser-produced plasmas were measured with a soft X-ray spectrometer and a soft X-ray plasma camera. The X-ray flux increased as the target thickness decreased and reached the maximum at the target thickness of 20 nm. We have observed living biological cells with the soft X-ray emissions generated from the thin-foiled gold targets.
Kado, Masataka; Kishimoto, Maki; Tamotsu, Satoshi*; Yasuda, Keiko*; Aoyama, Masato*; Shinohara, Kunio*
no journal, ,
Laser-plasma soft X-ray source produced by a high power pulsed laser is extremely bright and very suitable for biological X-ray microscopy to capture an image of living specimens for which require a single flash exposure to avoid imaging any damages on the specimens. We also have invented to use a fluorescent microscope to identify the cellular organelles in the images obtained with the soft X-ray microscope. The biological cells were cultivated directly onto the PMMA photo resists and observed with the soft X-ray microscope and the fluorescent microscope at the same time. The obtained soft X-ray images and fluorescence images of the cells were directly compared and each cellular organelle such as mitochondria, actin filaments, and chromosomes in the soft X-ray images were clearly identified. Since the soft X-ray microscope has higher spatial resolution than that of the fluorescent microscope, fine structures of the cellular organelles in the hydrated biological cells were observed.
Kado, Masataka; Kishimoto, Maki; Tone, Shigenobu*; Tamotsu, Satoshi*; Yasuda, Keiko*; Aoyama, Masato*; Shinohara, Kunio*
no journal, ,
Soft X-ray microscope using water window X-ray as light sources has been developed as epoch-making technology to observe living hydrated biological cells with high spatial resolution. Controlling the thickness of water layer is critical to observe living biological cells since the absorption of the soft X-ray by the water layer degrade the cell images. It is important to keep the thickness of the water layer at 5 m to obtain clear cell images. We have designed new silicon nitride windows with spacer and groove structure to keep the water layer thickness at 5 m and have succeeded to obtain high resolution cell images.
Kado, Masataka; Kishimoto, Maki; Ejima, Takeo*; Tamotsu, Satoshi*; Yasuda, Keiko*; Aoyama, Masato*; Shinohara, Kunio*
no journal, ,
Soft X-ray microscope has a potential to observe live biological cells. But the cells had to be frozen in order to avoid radiation effects onto the biological cells or to be made a short pulse imaging with an intense X-ray source. Otherwise live cell imaging was not realized. Laser-plasma soft X-ray source was extremely bright and had short pulse duration. Hence it makes possible to capture soft X-ray images of biological cells before the cells damaged. Since the spatial resolution of the soft X-ray microscope depends on the photon flux irradiated onto the samples, it is important to increase brightness of the soft X-ray sources to observe inner structures of biological cells. We have increased the brightness of the soft X-ray source and succeeded in observing inner structures of live biological cells.
Kado, Masataka; Kishimoto, Maki; Tamotsu, Satoshi*; Yasuda, Keiko*; Aoyama, Masato*; Shinohara, Kunio*
no journal, ,
We have developed a laser-plasma soft X-ray microscope combining an intense short pulsed soft X-ray source to a contact microscopy irradiating the soft X-rays onto cells directly cultivated on an X-ray photo resist and succeeded to observe inner structures of living cells directly. Using a fluorescent microscope with the soft X-ray microscope to observe the same cells at the same time, accurate identification and high resolved observation of cellular organelles have been achieved. Using the laser-plasma soft X-ray microscope inner structure of Leydig cells of mouse testis and structural deformation of apoptotic HeLa S3 nuclei have been observed. Immune cells of mouses have been observed and important structural change at the activation of immune function was found. We have found several important features by observing live cells which were unable by ordinal microscopes.
Kado, Masataka; Kishimoto, Maki; Tamotsu, Satoshi*; Yasuda, Keiko*; Aoyama, Masato*; Tone, Shigenobu*; Shinohara, Kunio*
no journal, ,
Soft X-ray wavelengths between absorption K-edges of Oxygen and Carbon (2.3 nm and 4.4 nm) are so called "water window" and the X-ray were well absorbed by Carbon and less absorbed by water. Soft X-ray microscope using the water window X-ray as the light source has advantage to be able to observe live biological cells without any artifacts and can observe fine structures of cells compared to the light microscope. Combining with bright and short-pulsed laser-plasma soft X-ray the soft X-ray microscope which named a laser-plasma soft X-ray microscope can observe live biological cells in situ without radiation damages. We have generated bright water window soft X-ray irradiating a high power laser with 1053 nm in wavelength, 20 J in pulse energy and 600 ps in pulse duration onto thin foiled gold targets. Cultivating biological cells directly on the PMMA photoresists in situ observation of live biological cells with the laser-plasma soft X-ray microscope has been realized.
Kado, Masataka
no journal, ,
Soft X-ray microscope using laser-plasma soft X-ray as the light source has advantage to be able to observe inner structures of live biological cells with the spatial resolution higher than 100 nm and provides powerful observation method to understand various life functions. In the seminar I present development of the laser-plasma soft X-ray microscope and examples of observation of live biological cells. I also present observation results of apoptotic nuclei which is one of the most important life functions.
Kado, Masataka; Kishimoto, Maki; Ejima, Takeo*; Tamotsu, Satoshi*; Yasuda, Keiko*; Aoyama, Masato*; Tone, Shigenobu*; Shinohara, Kunio*
no journal, ,
Although soft X-ray microscope has an ability to observe live biological cells, in order to avoid radiation damages onto the biological cells the cells had to be frozen and live cell imaging was not realized. Since laser-plasma soft X-ray source was extremely bright and had short pulse duration, it makes possible to capture soft X-ray images of biological cells before the cells damaged. Since the spatial resolution of the soft X-ray microscope depends on the photon flux irradiated onto the samples, it is important to increase brightness of the soft X-ray sources to observe inner structures of biological cells. We have increased the brightness of the soft X-ray source and succeeded to observe inner structures of live biological cells.
Kado, Masataka; Kishimoto, Maki; Ishino, Masahiko; Tamotsu, Satoshi*; Yasuda, Keiko*; Aoyama, Masato*; Tone, Shigenobu*; Shinohara, Kunio*
no journal, ,
We have developed a laser-plasma soft X-ray microscope composed of an intense short pulsed soft X-ray source and a contact microscopy system in which soft X-rays are irradiating onto biological cells directly cultivated on a recording media, an X-ray photo resist, and succeeded in observing inner structures of living biological cells. Using a fluorescence microscope with the soft X-ray microscope to observe the same biological cells at the same time, accurate identification and high resolved observation of cellular organelles have been achieved such as inner structure of Leydig cells from mouse testis, structural deformation of apoptotic HeLa S3 nuclei and mouse immune cells. In the case of the immune cells we have found structural changes possibly attributed to the activation of immune function.
Kado, Masataka; Kishimoto, Maki; Tamotsu, Satoshi*; Yasuda, Keiko*; Aoyama, Masato*; Tone, Shigenobu*; Shinohara, Kunio*
no journal, ,
A laser-plasma soft X-ray microscope which is combination of a highly intense laser-plasma soft X-ray source and contact microscopy has been developed. We have proposed a correlative microscopy with the laser-plasma soft X-ray microscope and a fluorescent microscope that is to observe the same biological cells with the both microscopes at the same time. Live hydrated biological cells and various cellular organelles of them have been observed with the correlative microscopy. Using the information of the cellular organelles obtained with the fluorescence microscope, inner structures obtained with the soft X-ray microscope are exactly identified. Since the spatial resolution of the soft X-ray microscope is much higher than that of the fluorescent microscope, fine structures of the cellular organelles of the live biological cells have been visualized with the correlative microscopy.
Kado, Masataka; Kishimoto, Maki; Tone, Shigenobu*; Tamotsu, Satoshi*; Yasuda, Keiko*; Aoyama, Masato*; Shinohara, Kunio*
no journal, ,
A soft X-ray microscope using a highly intense laser-plasma soft X-ray source (laser-plasma soft X-ray microscope) has an ability to observe live biological cells with a spatial resolution higher than 100 nm in a very short time period. Using the laser plasma soft X-ray microscope inner structures of various live biological cells such as cellular organelles of Leydig cells of mouse testis and structure change caused by immunological functions of immunological cells have been observed. We used the laser-plasma soft X-ray microscope to observe structural deformation of nucleus due to apoptotic process which is one of important life functions. We have found the structural deformation of nuclei from ring structure to necklace and also found important structure inside of the nuclei for the first time.
Kado, Masataka; Kishimoto, Maki; Tamotsu, Satoshi*; Yasuda, Keiko*; Aoyama, Masato*; Tone, Shigenobu*; Shinohara, Kunio*
no journal, ,
A laser-plasma soft X-ray microscope which is combination of a highly intense laser-plasma soft X-ray source and contact microscopy has been developed. We have proposed a correlative microscopy with the laser-plasma soft X-ray microscope and a fluorescent microscope that is to observe the same biological cells with the both microscopes at the same time. Live hydrated biological cells and various cellular organelles of them have been observed with the correlative microscopy. Using the information of the cellular organelles obtained with the fluorescence microscope, inner structures obtained with the soft X-ray microscope are exactly identified. Since the spatial resolution of the soft X-ray microscope is much higher than that of the fluorescent microscope, fine structures of the cellular organelles of the live biological cells have been visualized with the correlative microscopy.
Kado, Masataka
no journal, ,
Soft X-ray wavelengths between absorption K-edges of Oxygen and Carbon (2.3 nm and 4.4 nm) are so called "water window" and the X-ray were well absorbed by Carbon and less absorbed by water. Soft X-ray microscope using the water window X-ray as the light source has advantage to be able to observe live biological cells without any artifacts and can observe fine structures of cells compared to the light microscope. Combining with bright and short-pulsed laser-plasma soft X-ray the soft X-ray microscope which named a laser-plasma soft X-ray microscope can observe live biological cells in situ without radiation damages. We have generated bright water window soft X-ray irradiating a high power laser with 1053 nm in wavelength, 20 J in pulse energy and 600 ps in pulse duration onto thin foiled gold targets. Cultivating biological cells directly on the PMMA photoresists in situ observation of live biological cells with the laser-plasma soft X-ray microscope has been realized.
Kado, Masataka; Kishimoto, Maki; Tamotsu, Satoshi*; Yasuda, Keiko*; Aoyama, Masato*; Tone, Shigenobu*; Shinohara, Kunio*
no journal, ,
Laser-plasma soft X-ray sources have advantage of brightness and short pulse duration and a soft X-ray microscope using them can observe live biological cells with a high spatial resolution. We have succeeded to observe fine structures of cellular organelles of live biological cells and also structural deformations of apoptotic nuclei.
