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CELL OBSERVATION DEVICE, CELL OBSERVATION METHOD AND PROGRAM THEREOF UPDATE 新技術説明会

外国特許コード F150008129
整理番号 AF30-01WO
掲載日 2015年2月12日
出願国 世界知的所有権機関(WIPO)
国際出願番号 2013JP081894
国際公開番号 WO 2014084255
国際出願日 平成25年11月27日(2013.11.27)
国際公開日 平成26年6月5日(2014.6.5)
優先権データ
  • 特願2012-259880 (2012.11.28) JP
発明の名称 (英語) CELL OBSERVATION DEVICE, CELL OBSERVATION METHOD AND PROGRAM THEREOF UPDATE 新技術説明会
発明の概要(英語) A cell observation device comprising: an outline extracting section for extracting edge pixels from a cell image in a captured image of single layered cells and synthesizing an edge image consisting of the edge pixels thus extracted; a plastid region extracting section for extracting pixels of plastid of the cell image in the captured image and synthesizing a plastid image consisting of the plastid pixels thus extracted; and an image synthesizing section for, in a synthesized image obtained by overlapping the edge image and the plastid image together, detecting a cell image region and a background image region in the captured image on the basis of the pixel luminance value distribution and thus detecting the cell image region in the captured image.
従来技術、競合技術の概要(英語) BACKGROUND ART
In recent years, when detecting the status of a cell, a live state cells can be observed by a polarization microscope over a long period of experimental apparatus has been put to practical use. By using such an experimental apparatus, such as cell growth or cell division can be observed in real time the course. Then, the progress of the change of these cells to capture image data of the resulting cells by analyzing the time series, and detail analysis changes occurring in the cell becomes possible.
For example, fermentation was performed using a unicellular yeast, beer, alcoholic beverages having a variety of shochu can be manufactured. Is for maintaining the quality of an alcoholic beverage, the yeast used for fermentation such physiological states determined before fermentation, it is possible to predict the effect of subsequent to the fermentation is performed (for example, see Patent Document 2). That is, yeast fermentation, brewing substance production and the like, from which the production of a biological condition of contemplated for use in yeast cells is ascertained in advance, foreseeing the success or failure of the fermentation, and stabilized products having high quality is required in order to obtain.
Microalgae, mainly refers to unicellular photosynthetic organisms. This microalgae is, by photosynthesis, light energy is converted into chemical energy, self-energy conversion of the figure is used for survival and growth. In addition, the microalgae is, depending on the type, the hydrocarbon and the essential unsaturated fatty acid as a useful component (for example DHA (Docosa Hexaenoic Acid), such as EPA (Eicosa Pentaenoic Acid)), starch, and is combined with the raw pigment and the like, function in the biosynthesis of these industrial application has been expected.
The microalgae using a highly efficient production of a useful component in future, the physiological state of the microalgae cells it is important to properly grasp. That is, the physiological state of the microalgae cells, media composition, carbon dioxide concentration, light intensity, culture temperature, cell density of the surrounding environment such as changes greatly depending on the growth conditions. Then, the microalgae cells, the production of a useful component in accordance with the physiological conditions and the stored quantity also changes in some cases.
Therefore, a useful component by cells of microalgae for the production of a high-efficiency, optimization of the conditions of the growth of cells in the course of, a useful component in the course of the production by the cells also also, microalgae in culture to grasp the state of the metabolic, useful component is necessary to know the amount of production. In addition, by culturing the cell of the organism are mixed into the medium of the other type of microalgae influence the physiological state of cells is. Other types of physiological conditions due to the intrusion of the influence on the organism, a useful component by the cells are not uncommon problem in the production run. Therefore, the above-described culture into the culture medium for the detection of other types may be mixed, with the cells of the microalgae is a highly efficient for the production of a useful component becomes important.
As an example of the microalgae cells, and a kind of microalgae Haematococcus pluvialis. Is this Haematococcus pluvialis, is also used as a health food red anti-oxidant astaxanthin biosynthesis can be industrially very useful. Is Haematococcus pluvialis, reflects the physiological state of cells according to the various cell lines. The amount of accumulation of astaxanthin, varies depending on the culture conditions (for example, see non-patent document 1).
To this end the production of astaxanthin by Haematococcus pluvialis, the use of high-producing strain or culture condition optimization has been carried out (for example, see Patent Document 1). However, in the present situation is as not considered to have a high, a further improvement in productivity is expected. Further, in recent years, other types of parasitic Haematococcus pluvialis as an example of a fungal organism Paraphysoderma sedebokerensis is found. Haematococcus pluvialis Paraphysoderma sedobokerensis is infected, a dark brown color change from green cells, leading to death (for example, see non-patent document 2).
Haematococcus pluvialis have been identified and a plurality of the culture, a plurality of types of ordered from all over the world, organism other than the mixed Haematococcus pluvialis was examined. As a result, it has been surprisingly found, industrially used mixed in culture and contains all of the strains was confirmed. Therefore, the physiological state of cell Haematococcus pluvialis, a useful component of astaxanthin accumulation amount, to a mixing rate of the organism from other species can be grasped, a critical issue in industrial use.
As a method to grasp the state of the metabolic, for budding yeast microorganism, such as the evaluation method of discrimination techniques of killing by methylene blue method (for example, see non-patent document 3) have been known. However, in the method according to this Non-Patent Document 3, the physiological state of cells cannot be determined various aspects. In addition, the patent document 2, the quantitative value of the cell morphology of a physiological condition of yeast used evaluation method is shown. Specifically, the contour of the object of the yeast cell, nucleus, and fluorescent staining of the actin cytoskeleton and the image is subjected to image analysis, yeast cell morphological feature set form in advance based on the parameter, a value obtained by quantitative analysis of cell morphology, a database prepared in advance the value by comparing the, for the purpose of assessing a biological condition of yeast.
However, in this method cells fixed, coloring treatment and which require observation by fluorescence microscopy, such as in a production site for field and is not suitable for real-time of a physiological condition of grasp. The microalgae cells to incorporate not suggest no consideration. In addition, as a method for detecting an accumulation amount of the useful component, in patent document 1, the amount of astaxanthin Haematococcus pluvialis, extracted from the cells by the plastid dimethyl sulfoxide, 750 nm 492 nm and quantitated by measuring the absorbance of the method is shown. However, a large number of microalgae cells can be measured with the extraction of the color bodies from the required, time is required for the extraction of the plastid.
In addition, for the detection of other types, diatoms microalgae cells in the parasitic fungus to the pot of fungi, a component of the pot cell walls of fungi calcofluor white chitin binding specifically to a method of coloring is shown (non-patent document 4). In addition, according to another prior study, the capsular bag of a plant parasitic Haematococcus pluvialis Paraphysoderma sedebokerensis and stained with FITC-WGA(non-patent document 5).
However, any of these, and the processing and staining of the cells, cells in culture is not directly determined. Also, such as Patent Document 2 and non-patent document 5 in the method, the need for a fluorescence microscope, such as for field and is not suitable for the determination at a production site. Or more as mentioned above, the growing condition of the cells of the microalgae, the microalgae cells in the culture medium of other contaminating species such as biological (for example flora) mixed state, the amount of production of microalgae cell by the cells of each of the useful substance to monitor in real time is not a simple method.
  • 出願人(英語)
  • ※2012年7月以前掲載分については米国以外のすべての指定国
  • JAPAN SCIENCE AND TECHNOLOGY AGENCY
  • 発明者(英語)
  • OHYA Yoshikazu
  • KAWANO Shigeyuki
  • NOGAMI Satoru
  • OHNUKI Shinsuke
  • OTA Shuhei
  • WATANABE Koichi
国際特許分類(IPC)
指定国 National States: AE AG AL AM AO AT AU AZ BA BB BG BH BN BR BW BY BZ CA CH CL CN CO CR CU CZ DE DK DM DO DZ EC EE EG ES FI GB GD GE GH GM GT HN HR HU ID IL IN IR IS JP KE KG KN KP KR KZ LA LC LK LR LS LT LU LY MA MD ME MG MK MN MW MX MY MZ NA NG NI NO NZ OM PA PE PG PH PL PT QA RO RS RU RW SA SC SD SE SG SK SL SM ST SV SY TH TJ TM TN TR TT TZ UA UG US UZ VC VN ZA ZM ZW
ARIPO: BW GH GM KE LR LS MW MZ NA RW SD SL SZ TZ UG ZM ZW
EAPO: AM AZ BY KG KZ RU TJ TM
EPO: AL AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HR HU IE IS IT LT LU LV MC MK MT NL NO PL PT RO RS SE SI SK SM TR
OAPI: BF BJ CF CG CI CM GA GN GQ GW KM ML MR NE SN TD TG
参考情報 (研究プロジェクト等) CREST Creation of Basic Technology for Improved Bioenergy Production through Functional Analysis and Regulation of Algae and Other Aquatic Microorganisms AREA
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