JPH0641909B2 - Microphotometric method for measuring nucleic acid content of cells using color image analyzer - Google Patents
Microphotometric method for measuring nucleic acid content of cells using color image analyzerInfo
- Publication number
- JPH0641909B2 JPH0641909B2 JP63047115A JP4711588A JPH0641909B2 JP H0641909 B2 JPH0641909 B2 JP H0641909B2 JP 63047115 A JP63047115 A JP 63047115A JP 4711588 A JP4711588 A JP 4711588A JP H0641909 B2 JPH0641909 B2 JP H0641909B2
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- nucleic acid
- color
- cells
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Links
- 150000007523 nucleic acids Chemical class 0.000 title claims description 30
- 102000039446 nucleic acids Human genes 0.000 title claims description 30
- 108020004707 nucleic acids Proteins 0.000 title claims description 30
- 238000000034 method Methods 0.000 title claims description 15
- 238000005259 measurement Methods 0.000 claims description 22
- 238000002834 transmittance Methods 0.000 claims description 12
- 238000002835 absorbance Methods 0.000 claims description 6
- 238000005375 photometry Methods 0.000 claims description 5
- 238000010186 staining Methods 0.000 claims description 5
- 238000004364 calculation method Methods 0.000 claims description 3
- 238000010191 image analysis Methods 0.000 claims description 3
- 210000004027 cell Anatomy 0.000 description 18
- 238000010521 absorption reaction Methods 0.000 description 12
- 210000004940 nucleus Anatomy 0.000 description 8
- 238000009826 distribution Methods 0.000 description 7
- 230000015654 memory Effects 0.000 description 6
- 238000009649 Feulgen staining Methods 0.000 description 4
- 206010028980 Neoplasm Diseases 0.000 description 4
- 238000004458 analytical method Methods 0.000 description 4
- 201000011510 cancer Diseases 0.000 description 4
- 230000022131 cell cycle Effects 0.000 description 3
- 210000000805 cytoplasm Anatomy 0.000 description 3
- 238000000684 flow cytometry Methods 0.000 description 3
- 238000000765 microspectrophotometry Methods 0.000 description 3
- 230000003287 optical effect Effects 0.000 description 3
- 238000002360 preparation method Methods 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 230000001086 cytosolic effect Effects 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 239000000975 dye Substances 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 230000005284 excitation Effects 0.000 description 2
- 238000000605 extraction Methods 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 239000011358 absorbing material Substances 0.000 description 1
- 230000002238 attenuated effect Effects 0.000 description 1
- 230000002902 bimodal effect Effects 0.000 description 1
- 210000003855 cell nucleus Anatomy 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 235000019441 ethanol Nutrition 0.000 description 1
- 238000012757 fluorescence staining Methods 0.000 description 1
- 239000007850 fluorescent dye Substances 0.000 description 1
- 239000011888 foil Substances 0.000 description 1
- 238000009828 non-uniform distribution Methods 0.000 description 1
- 238000003672 processing method Methods 0.000 description 1
- 210000001685 thyroid gland Anatomy 0.000 description 1
- 210000004881 tumor cell Anatomy 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
- G01N15/10—Investigating individual particles
- G01N15/14—Optical investigation techniques, e.g. flow cytometry
- G01N15/1468—Optical investigation techniques, e.g. flow cytometry with spatial resolution of the texture or inner structure of the particle
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- Chemical & Material Sciences (AREA)
- Dispersion Chemistry (AREA)
- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Image Analysis (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Investigating Or Analysing Materials By Optical Means (AREA)
- Image Processing (AREA)
- Closed-Circuit Television Systems (AREA)
Description
【発明の詳細な説明】 〔産業上の利用分野〕 この発明は、医学または生物学に用いられ、例えばがん
の早期発見などに好適なカラー画像解析装置を用いた細
胞の核酸量の顕微測光方法に関するものである。TECHNICAL FIELD The present invention is used for medicine or biology, and for example, microscopic photometry of the amount of nucleic acid in cells using a color image analyzer suitable for early detection of cancer and the like. It is about the method.
従来、細胞の核酸量解析を行なう方法として、フローサ
イトメトリー（Flow Cytometry）による方法が知られて
いる。これは核酸に螢光色素を標識した細胞が均一に浮
遊する溶液状の試料を細い管状ないし滴状の流れとして
高速で流し、この流れにレーザ光を照射して、細胞から
発する螢光強度と散乱光の角度を測定し、これらの測定
値に基づいて細胞の核酸量を速やかに解析するものであ
る。この方法は測定精度が高く、細胞周期（cell cvcl
e）の解析にも利用可能であり、最も信頼性のある方法
とされている。しかし、設備が高価なこと、比較的大量
の細胞が均一に浮遊する溶液状の試料でしか計測できな
いこと、螢光の減衰などから再度計測ができないことな
どから、現在のところその利用範囲は研究所等の特殊な
施設に限られ、臨床応用には到っていない。Conventionally, as a method for analyzing the amount of nucleic acid in cells, a method using flow cytometry has been known. This is because a solution-like sample in which cells labeled with a fluorescent dye on nucleic acid are uniformly suspended is made to flow at high speed as a thin tubular or droplet-like flow, and this flow is irradiated with laser light, and the fluorescence intensity emitted from the cells is The angle of scattered light is measured, and the amount of nucleic acid in cells is rapidly analyzed based on these measured values. This method has high measurement accuracy, and cell cycle (cell cvcl
It can be used for e) analysis and is considered to be the most reliable method. However, because the equipment is expensive, it is possible to measure only a solution type sample in which a relatively large amount of cells are uniformly suspended, and it is not possible to measure again due to fluorescence attenuation etc. It is limited to special facilities such as offices and has not reached clinical application.
細胞の核酸量の解析を行なう別な方法としては顕微鏡を
用いる顕微螢光測光および顕微分光測光法がある。顕微
螢光測光法は核酸螢光染色を施した顕微鏡プレパラート
上の細胞が励起光を受けて発する螢光を顕微鏡を通して
光電子増倍管（フォトマル）で測るものである。理論的
にはフローサイトメトリーと類似するが、計測に時間を
要し、その間に励起光を受けた螢光が減衰すること、計
測する領域に顕微鏡を通過する螢光を絞り込む必要があ
るが、絞りを小さくするには限界があることなどから誤
差が生じ、測定精度に問題がある。As another method for analyzing the amount of nucleic acid in cells, there are microfluorescence photometry using a microscope and microspectrophotometry. The microfluorescence photometric method is a photomultiplier tube (photomultiplier) that measures the fluorescence emitted by cells on a microscope preparation which has been subjected to nucleic acid fluorescence staining upon receiving excitation light. Although theoretically similar to flow cytometry, it takes time to measure, the fluorescence that receives excitation light during that time is attenuated, and it is necessary to narrow down the fluorescence that passes through the microscope to the measurement area, Since there is a limit to how small the aperture can be made, an error occurs, which causes a problem in measurement accuracy.
一方、顕微分光測光法は、吸光度と溶質の濃度との間に
は比例関係が成立するという吸光光度計の理論（Lamber
t-Beerの法則）に基づいており、単色光を光源とする顕
微鏡を用いて核酸染色したプレパラート上の細胞の吸光
度を計測し、核酸を解析するものである。Lambert-Beer
の法則は対象が透明であり且つその中に吸光物質が均一
に分布していることを前提としている。しかし、顕微鏡
プレパラート上の細胞の核酸量を計測する場合、目的と
する核を取り囲む細胞質が必ずしも透明でないために、
非特異的に光が失われる（非特異的光損失）こと、また
核内の吸光物質（核酸）が不均一に分布することによる
誤差（分布誤差： Distributional Error）が生じるため、やはり精度に問
題が残る。このため現在では核酸染色色素に最も良く吸
収される波長と、ほとんど吸収されない波長の２波長の
光の吸光度を、微小な点で計測（焦点測光）し、それを
走査して集計算出する「２波長走査法」を用いて誤差を
なくす工夫がなされている。On the other hand, in microspectrophotometry, the theory of an absorptiometer that a proportional relationship is established between the absorbance and the concentration of solute (Lambertian).
Based on t-Beer's law), the absorbance of cells on a preparation stained with nucleic acid is measured using a microscope using monochromatic light as a light source to analyze the nucleic acid. Lambert-Beer
The law of (1) assumes that the object is transparent and that the light absorbing material is uniformly distributed therein. However, when measuring the amount of nucleic acid in cells on a microscope preparation, the cytoplasm surrounding the target nucleus is not necessarily transparent,
Since the light is lost non-specifically (non-specific light loss) and the non-uniform distribution of the light-absorbing substance (nucleic acid) in the nucleus causes an error (distribution error), accuracy is also a problem. Remains. For this reason, at present, the absorbance of light of two wavelengths, that is, the wavelength most absorbed by the nucleic acid staining dye and the wavelength hardly absorbed by the nucleic acid staining dye, is measured (focus photometry) at a minute point, and the result is scanned and tabulated for calculation. The wavelength scanning method is used to eliminate the error.
このように、顕微分光測光法はその測定誤差を少なくす
るために２波長走査法によって計測する必要があるが、
それでも計測走査スポット（焦点）を小さくするには限
界があり、どうしても多少の分布誤差の問題が残る（Re
sidual Distributional Error）。また、２波長の光の
吸光度を別々に２度しかも同一部位で計測することの技
術的な問題、計測点を走査しなければならないために時
間がかかるなどの解決すべき問題がある。As described above, in the microspectrophotometry, it is necessary to perform measurement by the two-wavelength scanning method in order to reduce the measurement error.
Even so, there is a limit to reducing the measurement scanning spot (focus), and the problem of some distribution error remains (Re
sidual Distributional Error). In addition, there are technical problems of separately measuring the absorbance of light of two wavelengths twice and at the same site, and there are problems to be solved such as it takes time because a measurement point must be scanned.
（課題を解決するための手段〕 このような問題を解決するためにこの発明は、カラーテ
レビジョンカメラと画像解析装置の組み合わせによっ
て、「カラー画像」測光の手段を用いるものである。(Means for Solving the Problem) In order to solve such a problem, the present invention uses a “color image” photometric means by a combination of a color television camera and an image analysis device.
カラーテレビジョンンカメラによって撮影されたデータ
がメモリに記憶され、順次読みだされて解析される。The data captured by the color television camera is stored in a memory, which is sequentially read and analyzed.
第１図はこの発明の一実施例を示すブロック図である。
図において、１はフォイルゲン染色した細胞の顕微鏡標
本であり、チャンバースライドで腫瘍細胞を単層培養後
７０％エチルアルコールで固定し、フォイルゲル（Feul
gen）反応で核酸染色を施したものである。この標本１
は光源２、反射鏡３、コンデンサレンズ４、対物レンズ
５、接眼レンズ６等からなる光学式顕微鏡７に載置され
る。顕微鏡７の光学像はカラーテレビジョンカメラ８で
撮影され、縁（Ｇ）および青（Ｂ）のカラーマルチ画像
としてＡ／Ｄ変換器９および１０にそれぞれ入力され、
デジタル画像データに変換されてＧおよびＢごとに設け
た画像メモリ１１および１２に記憶される。この実施例
においては、画像メモリ１１および１２をそれぞれ５１
２×４８０の画素で２５６階調の濃淡が計測できるよう
に構成した。なお、顕微鏡７とカラーテレビジョンカメ
ラ８は一体に構成された３管式顕微鏡カラーテレビジョ
ンカメラを用いている。顕微鏡の光源、コンデンサレン
ズには顕微測光用に開発されたものを使用し、テレビジ
ョンカメラのＧおよびＢの撮像管にはそれぞれフォイル
ゲン染色の特異吸収光（緑色光565ｎｍ）および非吸収
光（青色光450ｎｍ）に合うように干渉フィルタが挿入
され調整されている。画像メモリ１１および１２のＧお
よびＢのデジタル画像データは読み出されて特異透過率
画像抽出処理部１３に入力され、フォイルゲン染色の特
異吸光域にある色成分のＧ画像と、非特異吸光域のある
色成分のＢ画像とについて、それぞれ次式に基づいてＧ
成分透過率画像TgとＢ成分透過率画像Tbを作成する。FIG. 1 is a block diagram showing an embodiment of the present invention.
In the figure, 1 is a microscope specimen of Feulgen-stained cells. Tumor cells were monolayer-cultured on a chamber slide, fixed with 70% ethyl alcohol, and subjected to foil gel (Feulgel).
gen) reaction and subjected to nucleic acid staining. This specimen 1
Is mounted on an optical microscope 7 including a light source 2, a reflecting mirror 3, a condenser lens 4, an objective lens 5, an eyepiece lens 6 and the like. The optical image of the microscope 7 is photographed by the color television camera 8 and input to the A / D converters 9 and 10 as color multi-images of edge (G) and blue (B), respectively.
It is converted into digital image data and stored in the image memories 11 and 12 provided for each of G and B. In this embodiment, the image memories 11 and 12 are respectively 51
It is configured such that the gradation of 256 gradations can be measured with 2 × 480 pixels. The microscope 7 and the color television camera 8 use a three-tube microscope color television camera that is integrally configured. For the light source and condenser lens of the microscope, those developed for microscopic photometry are used, and the specific absorption light (green light 565 nm) and non-absorption light (blue light) of Feulgen staining are used for the G and B image pickup tubes of the television camera, respectively. An interference filter is inserted and adjusted to match the light (450 nm). The G and B digital image data of the image memories 11 and 12 are read out and input to the specific transmittance image extraction processing unit 13, and the G image of the color component in the specific absorption region of Feulgen staining and the non-specific absorption region are detected. For a B image of a certain color component, G
A component transmittance image Tg and a B component transmittance image Tb are created.
Ｔｇ＝Ｉｓ（ｇ）／Ｉｏ（ｇ）・・・・（１） Ｔｂ＝Ｉｓ（ｂ）／Ｉｏ（ｂ）・・・・（２） ここで、Is(g)，Is(b)は標本通過後の光量（特異吸光域
のある画像および吸光域のない画像）Io(g)，Io(b)は標
本への入射光量（背景画像）である。Tg = Is (g) / Io (g) ... (1) Tb = Is (b) / Io (b) ... (2) where Is (g) and Is (b) are samples The amount of light after passing (images with and without specific absorption region) Io (g) and Io (b) are the amounts of light incident on the sample (background image).
ついで、得られたＧ成分透過率画像TgとＢ成分画像Tbを
次式に基づいて除算するとともに、得られた特異透過率
画像を対数濃度変換してフォイルゲン染色の特異吸光画
像Ａとした。Next, the obtained G component transmittance image Tg and B component image Tb were divided based on the following equation, and the obtained specific transmittance image was subjected to logarithmic density conversion to obtain a specific absorption image A of Feulgen staining.
Ａ＝Ａｇ−Ａｂ・・・・・・・・・・（３） ＝−logＴｇ＋logＴｂ・・・・・・（４） ＝log（Ｔｂ／Ｔｇ）・・・・・・（５） ここでＡｇはＧ成分の吸光画像、ＡｂはＢ成分の吸光画
像である。A = Ag-Ab (3) = -logTg + logTb (4) = log (Tb / Tg) (5) where Ag is Absorption image of G component, Ab is an absorption image of B component.
以上の画像処理で得たフォイルゲン染色特異吸光画像Ａ
には、細胞質と細胞核が共に含まれている。従ってこの
細胞質領域と核領域を区分して目的とする核に計測領域
を限定するために、特異透過率画像を領域区分処理部１
４に入力し、各画素毎にその画素と周囲の画素との濃淡
差の和を求めその差分和を当該画素の重みとして画像全
体の濃度分布を算出して各画素の濃度差に基づく重み付
けをする。即ち、細胞質の領域が広くてもその領域内の
濃度差がなければ重みが小さく、その分布は小さな値と
なり、細胞質と核との境界に対応する濃度には濃度差が
あるため重みが大きく、例えその領域はせまくても大き
な値となる。したがって、この濃度差和分布におけるピ
ークに対応する濃度値を閾値として、フォイルゲン染色
の特異透過率画像を２値化区分して核領域画像を得た。
このようにして得た核領域画像と特異吸光画像は濃淡値
計測処理部１５に入力されて、細胞核領域内の濃淡値が
計測される。計測にあたっては、核領域画像中の任意の
核を自動的に指定して、特異吸光画像中の指定した核領
域に含まれる各画素点（１個の核あたり1000個程度含ま
れる）の濃淡値を計測集計し、第２図に示すヒストグラ
ムを作成した。Feulgen-stained specific absorption image A obtained by the above image processing
Contains both the cytoplasm and the cell nucleus. Therefore, in order to limit the measurement region to the target nucleus by dividing the cytoplasmic region and the nuclear region, the specific transmittance image is processed by the region division processing unit 1
4, the sum of the shade differences between the pixel and its surrounding pixels is calculated for each pixel, the difference sum is used as the weight of the pixel to calculate the density distribution of the entire image, and the weighting based on the density difference of each pixel is performed. To do. That is, even if the cytoplasmic region is wide, if there is no difference in concentration within that region, the weight is small, the distribution has a small value, and since the concentration corresponding to the boundary between the cytoplasm and the nucleus has a concentration difference, the weight is large, Even if the area is limited, it will be a large value. Therefore, the nuclear transmittance image was obtained by binarizing the specific transmittance image of Feulgen staining using the density value corresponding to the peak in the density difference sum distribution as a threshold value.
The nuclear region image and the specific absorption image thus obtained are input to the gray value measurement processing unit 15, and the gray value in the cell nuclear region is measured. In the measurement, an arbitrary nucleus in the nuclear region image is automatically designated, and the gray value of each pixel point included in the designated nuclear region in the specific absorption image (including about 1000 per nucleus) Was measured and tabulated, and the histogram shown in FIG. 2 was created.
第２図は培養した甲状腺未分化がん細胞株ＴＡ−１の各
核酸を前述の方法によって顕微鏡的に計測解析して求め
た核酸ヒストグラムであり、第３図は同じ細胞株につい
てフローサイトメータによって求めた核酸ヒストグラム
である。両図から明らかなように、今回提案する方法に
よって得られたヒストグラムは、細胞周期におけるG1お
よびG2＋Ｍ期において２峰性を示し、フローサイトメー
タによって得られた核酸ヒストグラムと略等しかった。FIG. 2 is a nucleic acid histogram obtained by microscopically measuring and analyzing each nucleic acid of the cultured thyroid undifferentiated cancer cell line TA-1, and FIG. 3 is a flow cytometer for the same cell line by a flow cytometer. It is the obtained nucleic acid histogram. As is clear from both figures, the histogram obtained by the method proposed this time was bimodal in the G1 and G2 + M phases of the cell cycle, and was almost the same as the nucleic acid histogram obtained by the flow cytometer.
なお、本実施例においては特異および非特異吸光画像と
して緑および青成分の画像を用いたが、核酸染色に応じ
て特異および非特異的に吸光する色成分の２画像であれ
ば同様に処理することができ、本実施例の色成分の画像
に限定されるものではない。Although the images of the green and blue components were used as the specific and non-specific absorption images in this example, two images of color components that absorb specifically and non-specifically according to the nucleic acid staining are similarly processed. However, the image is not limited to the color component image of this embodiment.
以上説明したようにこの発明は、カラーテレビジョンカ
メラと画像処理装置を用いたので、顕微鏡標本上の細胞
のように、吸光物質が不均一に存在する場合でも、フロ
ーサイトメトリーに匹敵する細胞周期の解析が可能な精
度で核酸量の算出が簡単に行なえるようになり次のよう
な効果を有する。As described above, since the present invention uses the color television camera and the image processing device, even when the light-absorbing substance is nonuniformly present like the cells on the microscope specimen, the cell cycle which is comparable to the flow cytometry. It becomes possible to easily calculate the amount of nucleic acid with such an accuracy that the analysis can be performed, and it has the following effects.
イ）テレビジョンカメラから画像解析装置に入力される
データはカラー画像データであり、１画像を微細な画素
点に分割して計測、解析処理するため、計測点を走査す
る必要がない。また、一個の核当たりに換算すると多数
の画素点に分割して計測でき、２波長走査法で計測焦点
面積をさらに微小にした場合と同等の精度が得られる。
即ち計測核領域の正確な区分が可能となり、計測焦点内
の分布誤差により生じる分布誤差がほとんど無視でき
る。B) The data input from the television camera to the image analysis device is color image data, and one image is divided into fine pixel points for measurement and analysis processing, so there is no need to scan the measurement points. Further, when converted into one nucleus, the measurement can be performed by dividing into a large number of pixel points, and the same accuracy as when the measurement focal area is further reduced by the two-wavelength scanning method can be obtained.
That is, the measurement nucleus region can be accurately divided, and the distribution error caused by the distribution error in the measurement focus can be almost ignored.
ロ）各計測焦点（画素）毎の計測値をカラーマルチデー
タとして入力処理するため、同一画素における任意の多
波長の吸光度を同時に計算できる。(B) Since the measurement value for each measurement focus (pixel) is input and processed as color multi-data, it is possible to simultaneously calculate the absorbance at any multi-wavelength in the same pixel.
ハ）計測データを計測点の集合である画像のまま解析処
理（画像間演算）するため、高速処理が可能である。C) Since the analysis data (inter-image calculation) is performed on the measurement data as an image that is a set of measurement points, high-speed processing is possible.
また顕微鏡永久標本上で正確な核酸ヒストグラムの解析
が可能となり、検体摂取量、処理法に制約がある一般の
臨床細胞診標本への応用についても十分に考えられる。Further, it becomes possible to analyze the nucleic acid histogram accurately on the permanent specimen of the microscope, and it is sufficiently possible to apply it to general clinical cytology specimens in which the amount of sample intake and the processing method are limited.
第１図はこの発明を適用した装置の一実施例を示すブロ
ック図、第２図はこの発明の方法によって求めた培養が
ん細胞の核酸ヒストグラム（出現頻度を表わすグラ
フ）、第３図はフローサイトメータによって求めた培養
がん細胞の核酸ヒストグラム（出現頻度を表わすグラ
フ）である。 １……顕微鏡標本、７……光学式顕微鏡、８……カラー
テレビジョンカメラ、９，１０……Ａ／Ｄ変換器、１１
……緑画像メモリ、１２……青画像メモリ、１３……特
異透過率画像抽出処理部、１４……領域区分処理部、１
５……濃淡値計測処理部。FIG. 1 is a block diagram showing an embodiment of an apparatus to which the present invention is applied, FIG. 2 is a nucleic acid histogram (graph showing the frequency of appearance) of cultured cancer cells obtained by the method of the present invention, and FIG. 3 is a flow chart. It is a nucleic acid histogram of a cultured cancer cell (graph showing the frequency of appearance) determined by a cytometer. 1 ... Microscope specimen, 7 ... Optical microscope, 8 ... Color television camera, 9, 10 ... A / D converter, 11
... green image memory, 12 ... blue image memory, 13 ... peculiar transmittance image extraction processing unit, 14 ... area division processing unit, 1
5 ... Gray value measurement processing unit.
───────────────────────────────────────────────────── フロントページの続き (72)発明者 古市 恒晃 東京都港区西新橋１丁目15番１号 日本ア ビオニクス株式会社内 (56)参考文献 特開 昭58−153144（ＪＰ，Ａ) 特開 昭61−226638（ＪＰ，Ａ) ─────────────────────────────────────────────────── ─── Continuation of the front page (72) Inventor Tsuneaki Furuichi 1-15-1 Nishi-Shimbashi, Minato-ku, Tokyo Within Japan Avionics Co., Ltd. (56) Reference JP-A-58-153144 (JP, A) JP Sho 61-226638 (JP, A)
Claims (1)
をカラー画像装置を用いることによって算出するカラー
画像解析装置を用いた細胞の核酸量の顕微測光方法にお
いて、 核酸染色を施した細胞の顕微鏡像をカラーテレビジョン
カメラで撮影して画像解析装置に入力しカラーマルチ画
像を作り出し、 その画像中で核酸染色の吸光域のある色成分と吸光域の
ない色成分とのそれぞれの画像を背景画像で画素毎に画
像間除算することによって輝度の減衰割合である透過率
の画像を作り出し、 この透過率画像間の演算処理を行ない核酸染色の特異透
過率画像を作り出し、 これを対数濃度変換して核酸染色特異吸光度画像を作り
出し、 核領域内の画素と周囲の画素とを２値区分して計測領域
を核に限定し、 核酸染色特異吸光画像の計測する核領域毎に含まれる画
素の濃淡値を計測し、 この計測値を集計して一つ一つの細胞の核酸相当量を算
出することを特徴とするカラー画像解析装置を用いた細
胞の核酸量の顕微測光方法。1. A method for microscopically measuring the amount of nucleic acid in a cell using a color image analyzer for calculating the amount of nucleic acid in a microscopic specimen stained with nucleic acid by using a color image device A microscopic image of is captured by a color television camera and input to an image analysis device to create a color multi-image. In the image, images of color components with absorptive region and those with absorptive region of nucleic acid stain are obtained. An image of the transmittance, which is the attenuation rate of the brightness, is created by dividing the image between pixels in the background image, and the calculation processing between the transmittance images is performed to create the specific transmittance image of the nucleic acid stain. To create a nucleic acid-staining specific absorbance image, and binarize the pixels in the nuclear region and surrounding pixels to limit the measurement region to the nucleus. Microscopic photometry of the amount of nucleic acid in cells using a color image analyzer characterized by measuring the gray value of pixels contained in each cell and totaling the measured values to calculate the amount of nucleic acid equivalent in each cell. Method.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63047115A JPH0641909B2 (en) | 1988-02-29 | 1988-02-29 | Microphotometric method for measuring nucleic acid content of cells using color image analyzer |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63047115A JPH0641909B2 (en) | 1988-02-29 | 1988-02-29 | Microphotometric method for measuring nucleic acid content of cells using color image analyzer |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPH01219654A JPH01219654A (en) | 1989-09-01 |
| JPH0641909B2 true JPH0641909B2 (en) | 1994-06-01 |
Family
ID=12766174
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP63047115A Expired - Lifetime JPH0641909B2 (en) | 1988-02-29 | 1988-02-29 | Microphotometric method for measuring nucleic acid content of cells using color image analyzer |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH0641909B2 (en) |
Families Citing this family (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6519355B2 (en) * | 2001-03-28 | 2003-02-11 | Alan C. Nelson | Optical projection imaging system and method for automatically detecting cells having nuclear and cytoplasmic densitometric features associated with disease |
| JP5245424B2 (en) | 2008-01-25 | 2013-07-24 | 日本電気株式会社 | Pathological tissue imaging system, pathological tissue imaging method, and pathological tissue imaging program |
| JP5344073B2 (en) * | 2012-09-14 | 2013-11-20 | 日本電気株式会社 | Pathological tissue imaging system, pathological tissue imaging method, and pathological tissue imaging program |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS58153144A (en) * | 1982-03-09 | 1983-09-12 | Nippon Kokan Kk <Nkk> | Apparatus for measuring organization composition ratio in massive material having a plurality of organizations with different reflectivities |
| JPS61226638A (en) * | 1985-03-30 | 1986-10-08 | Shimadzu Corp | Microscopic spectral image forming device |
-
1988
- 1988-02-29 JP JP63047115A patent/JPH0641909B2/en not_active Expired - Lifetime
Also Published As
| Publication number | Publication date |
|---|---|
| JPH01219654A (en) | 1989-09-01 |
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