JPH09304390A - Method for detecting microorganism in biological processing vessel - Google Patents
Method for detecting microorganism in biological processing vesselInfo
- Publication number
- JPH09304390A JPH09304390A JP8149886A JP14988696A JPH09304390A JP H09304390 A JPH09304390 A JP H09304390A JP 8149886 A JP8149886 A JP 8149886A JP 14988696 A JP14988696 A JP 14988696A JP H09304390 A JPH09304390 A JP H09304390A
- Authority
- JP
- Japan
- Prior art keywords
- microorganism
- microorganisms
- antibody
- antigen
- treatment tank
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Landscapes
- Treatment Of Biological Wastes In General (AREA)
Abstract
Description
【０００１】[0001]
【発明の属する技術分野】本発明は、下水、工場排水あ
るいは家庭排水などの汚水の浄化処理を行う生物処理槽
内の微生物相の変化を検出する生物処理槽の微生物検出
方法に関するものである。BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for detecting microorganisms in a biological treatment tank, which detects changes in the microflora in the biological treatment tank for purifying sewage, industrial wastewater, domestic wastewater and other wastewater.
【０００２】[0002]
【従来の技術】下水、工場排水、家庭排水などの汚水を
浄化処理するためには、嫌気性細菌や好気性細菌を含む
各種微生物を利用した生物化学的処理法が多く用いられ
ているが、処理を効率良く行うためには処理槽内の微生
物相を把握し、それが変化した場合には運転管理条件を
変えるなどして、常に良好な微生物相を保つことが重要
である。従来、生物処理槽内の微生物相を知るために
は、特定の微生物を単離して純粋培養して得られた菌体
を抗原とする抗体を生産し、特異的抗原抗体反応により
特定微生物が処理槽内に存在することを確認する方法が
提案されている（例えば、特公昭５７−２６１１０号公
報）。2. Description of the Related Art In order to purify sewage, factory wastewater, domestic wastewater, and other wastewater, biochemical treatment methods using various microorganisms including anaerobic bacteria and aerobic bacteria are often used. In order to perform the treatment efficiently, it is important to grasp the microbial flora in the treatment tank and, if it changes, to maintain good microbial flora by changing the operation control conditions. Conventionally, in order to know the microflora in a biological treatment tank, an antibody whose antigen is a bacterial body obtained by isolating and culturing a specific microorganism is produced, and the specific microorganism is treated by a specific antigen-antibody reaction. A method of confirming the presence in the tank has been proposed (for example, Japanese Patent Publication No. 57-26110).
【０００３】[0003]
【発明が解決しようとする課題】ところが、従来技術で
は生物処理槽内の微生物相を検出するためには、槽内に
存在する多くの種類の微生物のそれぞれについて、単
離、純粋培養、抗体生産、抗原抗体反応の検出を行わな
ければならず、多くの時間と労力が必要であり、また、
純粋培養が不可能な微生物に対しては抗体が生産できな
いという問題点があった。そこで、処理槽内の微生物群
に対する抗体の生産に時間と労力を浪費する必要のな
い、抗体を用いて１回の免疫反応により微生物相を判定
できる、生物処理槽の微生物検出方法を提供することを
目的とする。However, in the prior art, in order to detect the microbial flora in the biological treatment tank, isolation, pure culture, and antibody production were carried out for each of many kinds of microorganisms existing in the tank. , Detection of antigen-antibody reaction has to be carried out, which requires a lot of time and labor, and
There is a problem that antibodies cannot be produced against microorganisms that cannot be purely cultured. Therefore, it is an object of the present invention to provide a method for detecting a microorganism in a biological treatment tank, which does not need to waste time and labor for producing an antibody against a microbial group in the treatment tank and can determine the microflora by a single immune reaction using the antibody. With the goal.
【０００４】[0004]
【課題を解決するための手段】上記問題を解決するため
に、本発明は汚水の浄化処理を行う生物処理槽中の微生
物を抗原とし、前記抗原を脊椎動物の血液中に注入する
ことによりポリクローナル抗体を生産し、前記ポリクロ
ーナル抗体を前記生物処理槽内の微生物と特異的抗原抗
体反応を行わせることにより前記生物処理槽内の微生物
の有無、存在量を検出する生物処理槽の微生物検出方法
において、前記微生物を複数種の微生物の細胞体の混合
物からなるものとし、前記微生物の細胞体の混合物に殺
菌処理、界面活性剤による可溶化処理または熱処理のい
ずれかの処理を施したものを前記抗原として前記抗体を
生産するようにしたものである。なお、前記殺菌処理は
紫外線あるいは有機溶媒により行うようにすると良い。
あるいは前記微生物の細胞体の混合物に施した前記処理
に替えて、前記微生物の細胞体の混合物から抽出した膜
画分を前記抗原として前記抗体を生産するようにしても
良い。In order to solve the above problems, the present invention uses a microorganism in a biological treatment tank for purifying sewage as an antigen, and injects the antigen into the blood of a vertebrate to produce a polyclonal antibody. In the method for detecting a microorganism in a biological treatment tank, the presence of a microorganism in the biological treatment tank is detected by producing an antibody, and the polyclonal antibody is allowed to undergo a specific antigen-antibody reaction with the microorganism in the biological treatment tank. The microorganism is composed of a mixture of cell bodies of a plurality of kinds of microorganisms, and the mixture of microbial cell bodies is subjected to sterilization treatment, solubilization treatment with a surfactant, or heat treatment. As the above-mentioned antibody is produced. The sterilization treatment may be performed with ultraviolet rays or an organic solvent.
Alternatively, instead of the treatment performed on the mixture of the cell bodies of the microorganism, the membrane fraction extracted from the mixture of the cell bodies of the microorganism may be used as the antigen to produce the antibody.
【０００５】[0005]
【発明の実施の形態】以下、本発明の実施例を図に基づ
いて説明する。図１は生物処理槽内の微生物群を検出す
るためのポリクローナル抗体を生産する手順を示すフロ
ーチャートである。図２はポリクローナル抗体と微生物
の細胞体との反応の検出手順を示すフローチャートであ
る。ここで、生物処理槽内の微生物群は原生動物や細菌
からなる多数種類の微生物を含むものである。最初にポ
リクローナル抗体の生産手順について述べる。図１にお
いて、まずステップＳＴ１にて良好な運転状態もしくは
処理能力が低下した状態、負荷の高い状態、負荷の低い
状態など様々な既知の運転状態にある生物処理槽から少
量の処理水を採取する。次にステップＳＴ２にて砂泥や
ゴミを除いた後、遠心分離操作などにより処理水中の微
生物群の細胞体すべてを回収する。このすべての回収さ
れた微生物群の細胞体にステップＳＴ３にて殺菌処理、
界面活性剤による可溶化処理または熱処理（例えば、６
０〜１００°Ｃ）のいずれかの処理を施したものを抗原
として使用して、抗体の生産を行う。殺菌処理は紫外線
あるいは有機溶媒によるもので行うと良い。なお、有機
溶媒による殺菌処理はフェノ−ル処理、ホルマリン処理
あるいはアルコ−ル処理などの処理を用いてもよい。あ
るいは、ステップＳＴ３に替えてステップＳＴ４にて超
音波処理や遠心処理などにより複数種の微生物の細胞体
の混合物から抽出した膜画分を使用して抗体の生産を行
うようにしても良い。さらに続いて、ステップＳＴ５に
て、上記ステップＳＴ３あるいはステップＳＴ４の処理
により得られた細胞体を適当な期間をおいて脊椎動物の
血液中に数回注入し、ステップＳＴ６にて生体の抗原抗
体反応を利用して、その動物にポリクローナル抗体を生
産させる。ステップＳＴ７にて数週間後に、動物の全血
を採取し、生産されたポリクローナル抗体（γグロブリ
ン分画）を生産する。ステップＳＴ８にて生産した抗体
はステップＳＴ９にて、そのまま、または、蛍光色素に
より標識した後、サンプルチューブなどに分注して保存
しておく。一般に、生物処理槽内で優占的な微生物種
は、処理水の状態（有機物濃度、溶存酸素濃度等）によ
り大きく変化する（例えば、溶存酸素量が低い場合に
は、鞭毛虫類や分散状細菌が多くみられるが溶存酸素量
が高くなるにつれ、繊毛虫類や糸状性細菌があらわれ
る）ことが知られている。図１の手法により作製された
ポリクローナル抗体は、抗原とした微生物群を採取した
状態（例えば、良好な運転状態）にある生物処理槽中の
優占する複数種の微生物に特異的に反応する複数種の抗
体分子を多く含み、まったく異なった状態の処理槽内に
多く出現する微生物に特異的な抗体分子は含まない。DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS An embodiment of the present invention will be described below with reference to the drawings. FIG. 1 is a flow chart showing a procedure for producing a polyclonal antibody for detecting a microbial group in a biological treatment tank. FIG. 2 is a flowchart showing a procedure for detecting a reaction between a polyclonal antibody and a cell body of a microorganism. Here, the group of microorganisms in the biological treatment tank includes many kinds of microorganisms composed of protozoa and bacteria. First, the procedure for producing a polyclonal antibody will be described. In FIG. 1, first, in step ST1, a small amount of treated water is collected from a biological treatment tank in various known operating states such as a favorable operating state or a state where the treatment capacity is low, a high load state and a low load state. . Next, after removing sand and dirt in step ST2, all the cell bodies of the microbial group in the treated water are collected by centrifugation or the like. Sterilization is performed on the cell bodies of all the collected microorganisms in step ST3.
Solubilization or heat treatment with a surfactant (for example, 6
Antibodies are produced by using, as an antigen, one that has been subjected to any treatment of 0 to 100 ° C. The sterilization treatment may be performed with ultraviolet rays or an organic solvent. The sterilization treatment with an organic solvent may be a treatment such as phenol treatment, formalin treatment or alcohol treatment. Alternatively, instead of step ST3, in step ST4, antibody production may be performed using a membrane fraction extracted from a mixture of cell bodies of a plurality of types of microorganisms by ultrasonication or centrifugation. Then, in step ST5, the cell bodies obtained by the processing in step ST3 or step ST4 are injected into the blood of the vertebrate several times with an appropriate period, and in step ST6, the antigen-antibody reaction of the living body is performed. To produce a polyclonal antibody in the animal. After several weeks in step ST7, whole blood of the animal is collected to produce the produced polyclonal antibody (γ-globulin fraction). The antibody produced in step ST8 is stored in the sample tube or the like as it is or after being labeled with a fluorescent dye in step ST9. In general, the microbial species that predominate in biological treatment tanks vary greatly depending on the state of treated water (organic matter concentration, dissolved oxygen concentration, etc.) (for example, when the dissolved oxygen amount is low, flagellates or dispersed species are used). Many bacteria are seen, but as the amount of dissolved oxygen increases, ciliates and filamentous bacteria appear). The polyclonal antibody produced by the method of FIG. 1 is a multiple antibody that specifically reacts with a plurality of dominant microorganisms in a biological treatment tank in a state (for example, in a good operating state) in which a group of microorganisms used as an antigen is collected. It does not contain the antibody molecules that are rich in the species antibody molecules, but are specific to the microorganisms that often appear in the treatment tanks in completely different states.
【０００６】次に図２において、測定対象となる処理槽
内の微生物群と図１に示した手順により作製したポリク
ローナル抗体の抗原反応を測定する手順を述べる。ま
ず、ステップＳＴ１０にて処理水の一部を採取し、ステ
ップＳＴ１１にて微生物群の細胞体を回収する。次に、
ステップＳＴ１２にて上記抗体と混合、反応させた後、
ステップＳＴ１３にて洗浄して過剰の未反応の抗体を除
去する。このプロセスは、溶液中に懸濁した状態で行っ
ても、微生物をメンブレン上に固着させた状態で行って
もよい。蛍光色素標識した抗体を使用した場合はステッ
プＳＴ１４にて、励起光を照射して得られる蛍光の強度
から微生物群と反応した抗体の量を定量的に検出する。
蛍光色素標識していない抗体を用いる場合には、ステッ
プＳＴ１５にて細胞体との反応後に市販の蛍光標識した
２次抗体等を利用して反応量を検出できる。上記ポリク
ローナル抗体は、ステップＳＴ９にて、抗原を作製する
元となった状態（例えば、良好な運転状態）の処理水中
の微生物群とは反応性が高いが、処理槽の状態が変化す
るにつれ反応性が低下し、まったく異なった状態（例え
ば、処理能力が著しく低下した場合）に出現する微生物
群にはほとんど反応しない。したがって、図１の手法に
より作製したポリクローナル抗体を利用すれば、ステッ
プＳＴ１２にて１回の図２に示した抗原抗体反応測定に
より、生物処理槽内の微生物相およびその変化を定量的
に判定することが可能になる。また、ステップＳＴ１６
にてこの測定値を指標として生物処理槽の運転条件を調
整し、管理することが可能になる。また、様々な既知の
運転状態にある処理槽の微生物群を抗原とした複数種類
のポリクローナル抗体を生産し、それぞれについて上記
の測定を定期的に行えば、その時点の処理槽の状態、さ
らには、どの状態に向かって変化しつつあるかを知るこ
とができ、その判定を指標として処理槽の運転管理を容
易に行うこともできる。上記手段により、短時間、少労
力で生物処理槽内の微生物相の判定を行うことができ
る。Next, referring to FIG. 2, a procedure for measuring the antigen reaction of the microorganism group in the treatment tank to be measured and the polyclonal antibody prepared by the procedure shown in FIG. 1 will be described. First, in step ST10, a part of the treated water is collected, and in step ST11, the cell bodies of the microorganism group are collected. next,
After mixing and reacting with the above antibody in step ST12,
Excess unreacted antibody is removed by washing in step ST13. This process may be performed in a state of being suspended in a solution or in a state in which microorganisms are fixed on a membrane. When a fluorescent dye-labeled antibody is used, in step ST14, the amount of the antibody that has reacted with the microorganism group is quantitatively detected from the intensity of fluorescence obtained by irradiating the excitation light.
When an antibody not labeled with a fluorescent dye is used, the reaction amount can be detected by using a commercially available fluorescently labeled secondary antibody or the like after the reaction with the cell body in step ST15. The above-mentioned polyclonal antibody is highly reactive with the microorganism group in the treated water in the state (for example, good operating state) from which the antigen was prepared in step ST9, but reacts as the state of the treatment tank changes. It is less reactive and reacts poorly to microbial populations that appear in completely different states (eg, significantly reduced throughput). Therefore, if the polyclonal antibody produced by the method of FIG. 1 is used, the microbial flora in the biological treatment tank and its change are quantitatively determined by the antigen-antibody reaction measurement shown in FIG. 2 once in step ST12. It will be possible. In addition, step ST16
It becomes possible to adjust and manage the operating conditions of the biological treatment tank by using this measured value as an index. In addition, if a plurality of types of polyclonal antibodies are produced using the microorganisms in the processing tank in various known operating states as antigens, and the above measurement is periodically performed for each, the state of the processing tank at that time, It is possible to know which state is changing, and the operation management of the processing tank can be easily performed using the determination as an index. By the above means, the determination of the microflora in the biological treatment tank can be performed in a short time and with little labor.
【０００７】[0007]
【発明の効果】以上に述べたように、本発明によれば、
処理槽内の微生物群に対する抗体の生産に時間と労力を
浪費する必要のない、抗体を用いて１回の免疫反応によ
り微生物相を判定できる、生物処理槽の微生物検出方法
を得る効果がある。As described above, according to the present invention,
There is an effect of obtaining a method for detecting a microorganism in a biological treatment tank, which does not need to waste time and labor for producing an antibody against a group of microorganisms in the treatment tank and can determine the microflora by a single immune reaction using the antibody.
【図１】複数種の微生物を検出するポリクローナル抗体
の生産手順を示すフローチャートである。FIG. 1 is a flowchart showing a procedure for producing a polyclonal antibody that detects a plurality of types of microorganisms.
【図２】ポリクローナル抗体と微生物の細胞体との反応
の検出手順を示すフローチャートである。FIG. 2 is a flowchart showing a procedure for detecting a reaction between a polyclonal antibody and a cell body of a microorganism.
Claims (3)
生物を抗原とし、前記抗原を脊椎動物の血液中に注入す
ることによりポリクローナル抗体を生産し、前記ポリク
ローナル抗体を前記生物処理槽内の微生物と特異的抗原
抗体反応を行わせることにより前記生物処理槽内の微生
物の有無、存在量を検出する生物処理槽の微生物検出方
法において、 前記微生物を複数種の微生物の細胞体の混合物からなる
ものとし、 前記微生物の細胞体の混合物に殺菌処理、界面活性剤に
よる可溶化処理または熱処理のいずれかの処理を施した
ものを前記抗原として前記抗体を生産することを特徴と
する生物処理槽の微生物検出方法。1. A polyclonal antibody is produced by injecting the antigen into the blood of a vertebrate using the microorganism in the biological treatment tank for purifying wastewater as an antigen, and the polyclonal antibody is produced in the biological treatment tank. In the method of detecting a microorganism in a biological treatment tank for detecting the presence or absence of the microorganism in the biological treatment tank by causing a specific antigen-antibody reaction with the microorganism, the microorganism comprises a mixture of cell bodies of a plurality of types of microorganisms. In the biological treatment tank, the mixture of cell bodies of the microorganism is subjected to sterilization treatment, solubilization treatment with a surfactant, or heat treatment to produce the antibody as the antigen. Microbial detection method.
により行うことを特徴とする請求項１記載の生物処理槽
の微生物検出方法。2. The method for detecting microorganisms in a biological treatment tank according to claim 1, wherein the sterilization treatment is performed with ultraviolet rays or an organic solvent.
生物を抗原とし、前記抗原を脊椎動物の血液中に注入す
ることによりポリクローナル抗体を生産し、前記ポリク
ローナル抗体を前記生物処理槽内の微生物と特異的抗原
抗体反応を行わせることにより前記生物処理槽内の微生
物の有無、存在量を検出する生物処理槽の微生物検出方
法において、 前記微生物を複数種の微生物の細胞体の混合物からなる
ものとし、 前記微生物の細胞体の混合物から抽出した膜画分を前記
抗原として前記抗体を生産することを特徴とする生物処
理槽の微生物検出方法。3. A polyclonal antibody is produced by injecting the antigen into the blood of a vertebrate using the microorganism in the biological treatment tank for purification of wastewater as an antigen, and the polyclonal antibody is produced in the biological treatment tank. In the method of detecting a microorganism in a biological treatment tank for detecting the presence or absence of the microorganism in the biological treatment tank by causing a specific antigen-antibody reaction with the microorganism, the microorganism comprises a mixture of cell bodies of a plurality of types of microorganisms. A method for detecting a microorganism in a biological treatment tank, which comprises producing the antibody using a membrane fraction extracted from a mixture of cell bodies of the microorganism as the antigen.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP8149886A JPH09304390A (en) | 1996-05-20 | 1996-05-20 | Method for detecting microorganism in biological processing vessel |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP8149886A JPH09304390A (en) | 1996-05-20 | 1996-05-20 | Method for detecting microorganism in biological processing vessel |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH09304390A true JPH09304390A (en) | 1997-11-28 |
Family
ID=15484793
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP8149886A Pending JPH09304390A (en) | 1996-05-20 | 1996-05-20 | Method for detecting microorganism in biological processing vessel |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH09304390A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2008018486A1 (en) * | 2006-08-09 | 2008-02-14 | Kurita Water Industries Ltd. | Method and apparatus for biological treatment of organic wastewater |
-
1996
- 1996-05-20 JP JP8149886A patent/JPH09304390A/en active Pending
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2008018486A1 (en) * | 2006-08-09 | 2008-02-14 | Kurita Water Industries Ltd. | Method and apparatus for biological treatment of organic wastewater |
| JP2008036580A (en) * | 2006-08-09 | 2008-02-21 | Kurita Water Ind Ltd | Biological treatment method of organic wastewater |
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