JPS6142825B2 - - Google Patents
Info
- Publication number
- JPS6142825B2 JPS6142825B2 JP14272878A JP14272878A JPS6142825B2 JP S6142825 B2 JPS6142825 B2 JP S6142825B2 JP 14272878 A JP14272878 A JP 14272878A JP 14272878 A JP14272878 A JP 14272878A JP S6142825 B2 JPS6142825 B2 JP S6142825B2
- Authority
- JP
- Japan
- Prior art keywords
- yeast
- atcc
- brewing
- factor antibody
- bisporus
- 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.)
- Expired
Links
- 240000004808 Saccharomyces cerevisiae Species 0.000 claims description 41
- 235000014680 Saccharomyces cerevisiae Nutrition 0.000 claims description 41
- 239000000427 antigen Substances 0.000 claims description 12
- 102000036639 antigens Human genes 0.000 claims description 12
- 108091007433 antigens Proteins 0.000 claims description 12
- 238000001514 detection method Methods 0.000 claims description 10
- 244000005700 microbiome Species 0.000 claims description 9
- 241000235048 Meyerozyma guilliermondii Species 0.000 claims description 5
- 241000582914 Saccharomyces uvarum Species 0.000 claims description 4
- 241001123227 Saccharomyces pastorianus Species 0.000 claims description 3
- 244000288561 Torulaspora delbrueckii Species 0.000 claims description 2
- 241000222120 Candida <Saccharomycetales> Species 0.000 claims 1
- 241000235070 Saccharomyces Species 0.000 claims 1
- 238000000034 method Methods 0.000 description 17
- 238000012360 testing method Methods 0.000 description 7
- 230000004520 agglutination Effects 0.000 description 5
- 238000006243 chemical reaction Methods 0.000 description 5
- 235000013405 beer Nutrition 0.000 description 4
- 241000894006 Bacteria Species 0.000 description 2
- 241000283973 Oryctolagus cuniculus Species 0.000 description 2
- 241000877401 Saccharomyces ellipsoideus Species 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 2
- 238000013124 brewing process Methods 0.000 description 2
- 238000012136 culture method Methods 0.000 description 2
- 230000003053 immunization Effects 0.000 description 2
- 241000894007 species Species 0.000 description 2
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- KDXKERNSBIXSRK-UHFFFAOYSA-N Lysine Natural products NCCCCC(N)C(O)=O KDXKERNSBIXSRK-UHFFFAOYSA-N 0.000 description 1
- 239000004472 Lysine Substances 0.000 description 1
- 230000000890 antigenic effect Effects 0.000 description 1
- 230000001580 bacterial effect Effects 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- ZXJXZNDDNMQXFV-UHFFFAOYSA-M crystal violet Chemical compound [Cl-].C1=CC(N(C)C)=CC=C1[C+](C=1C=CC(=CC=1)N(C)C)C1=CC=C(N(C)C)C=C1 ZXJXZNDDNMQXFV-UHFFFAOYSA-M 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 238000000691 measurement method Methods 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 230000001717 pathogenic effect Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 230000000405 serological effect Effects 0.000 description 1
- 235000015041 whisky Nutrition 0.000 description 1
Landscapes
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Description
【発明の詳細な説明】
本発明は醸造に有害な微生物の検出方法に関
し、詳しくはビール等の醸造中に出現することが
ある醸造に有害な微生物を検出する方法に関す
る。
現在、ビール醸造工程中に現れる野生酵母の検
出にはLysine培地、Crystal―Violet培地等が広
く使用されているが、これらは培養を伴う手法で
あるため、迅速性に欠けている。
また、別法として螢光抗体法で代表される血清
学的な手法によつて有害微生物を検出することも
提案されている。この方法は迅速性という点では
上記手法よりすぐれているが、数種類の抗血清を
使用し、それぞれの結果を総合して判定しなけれ
ばならないため簡便性に欠けるものである。たと
えば、本発明者らが先に開発した手法においては
現在、我国で市販されている病原性キヤンデイダ
属同定用因子抗体を使用するけれども、５種類も
の因子抗体を用いる必要があるため、操作が煩雑
となる傾向があり効果的な手法とは云えない。
本発明は上記の欠点を解消し、単一の因子抗体
を用いることによつて醸造に有害な微生物を迅速
かつ簡便に検出できる方法を提供するものであ
る。
本発明はキヤンデイダ属もしくはサツカロミセ
ス属に属する酵母を抗原として得た抗血清を醸造
用酵母で吸収処理して因子抗体を調製し、該因子
抗体を被検酵母と接触させ、その際に凝集反応を
生起するか否かによつて該酵母の性質を判定し、
醸造に有害な微生物を検出するものである。
本発明者らは単一の因子抗体によつて醸造工程
中に発生する有害な微生物を迅速かつ正確に検出
する方法を開発すべく研究を重ねた結果、抗血清
として下記のものを用い、かつ該抗血清を醸造用
酵母を吸収菌として吸収処理して調製した因子抗
体が、、本発明の目的に適合することを見出した
のである。
抗血清として用いるものは、キヤンデイダ・ギ
リアモンデイ（Candidaguilliermondi，ATCC―
9058）、サツカロミセス・セレビシエー
（Saccharomyces cerevisiae Ｊ―6001）、サツカ
ロミセス・パストリアヌス（S.pastrorianus，
BSRI―１―12）、サツカロミセス・ロゼイ（S.
rosei，ATCC―0428）、サツカロミセス・ビスポ
ラス（S.bisporus，ATCC―0723）およびサツカ
ロミセス・ウバルム（S.uvarum，BSRI―１―
９）の酵母を抗原としてそれぞれ兎を免疫して得
られるものである。
上記菌株を抗原として兎を免疫して得た抗血清
を醸造用酵母で吸収処理することによつて因子抗
体を調製する。醸造用酵母は使用目的に応じて選
定すればよく、たとえばビール醸造用酵母につい
ては下面ビール酵母あるいは上面ビール酵母があ
り、その他清酒酵母、ワイン酵母、ウイスキー酵
母、アルコール酵母等がある。
因子抗体は単一のものでもよく、あるいは２以
上を組合せて使用することができる。とりわけサ
ツカロミセス・パストリアヌスを抗原として用い
た因子抗体とサツカロミセス・ビスポラスを抗原
として用いた因子抗体とを組合せたものやサツカ
ロミセス・セレビシエーを抗原として用いた因子
抗体とキヤンデイダ・ギリアモンデイを抗原とし
て用いた因子抗体とを組合せたものは非常に高率
で有害微生物を検出することができる。
本発明に係る因子抗体を被検酵母と接触させる
と、被験酵母が吸収菌として用いた醸造用酵母で
ある場合を除いて凝集反応が生起する。このよう
な被験酵母の挙動は該酵母の有する抗原構造の差
異に起因するものであり、本発明によれば単一の
因子抗体を用いるのみで醸造に有害な微生物を迅
速かつ正確に検出することができる。因子抗体と
被検酵母との接触はスライドグラス上で行なわし
めることができ、短時間で凝集反応の有無を確認
することが可能である。しかも必要があれば一時
に多数の被検酵母について試験することができ
る。
本発明の方法に使用する因子抗体は、既知の螢
光抗体法および培養法に対しても有効に使用でき
る。
次に本発明の実施例を示す。
実施例 １
サツカロミセス・ビスポラス（S.bisporus，
ATCC―0723）を抗原として抗血清を得、該抗血
清を下面ビール酵母（S.carlsbergensis，IAM―
4778）で吸収処理して常法により因子抗体を調製
した。
次いで、この因子抗体を用いて野生酵母16株に
ついてスライド凝集反応を試みたところ、検出率
は75％であつた。
実施例 ２
サツカロミセス・ビスポラスの代りにサツカロ
ミセス・ウバルム（S.uvarum，BSRI―１―９）
を用いたこと以外は実施例１と同様の方法により
スライド凝集反応を試みたところ、検出率は45％
であつた。
実施例 ３
サツカロミセス・ビスポラスの代りにキヤンデ
イダ・ギリアモンデイ（C.guilliermondii，
ATCC―9058）を用いたこと以外は実施例１と同
様の方法によりスライド凝集反応を試みたとこ
ろ、検出率は69％であつた。
実施例 ４
因子抗体として、サツカロミセス・パストリア
ヌス（S.pastorianus，BSRI―１―12）およびサ
ツカロミセス・ビスポラス（S.bisporus，ATCC
―0723）をそれぞれ抗原として用いて調製した２
種の因子抗体を組合せて用い、その他は実施例１
と同じ操作を試みたところ、野生酵母の検出率は
94％であつた。
実施例 ５
因子抗体として、サツカロミセス・セレビシエ
ー（S.cerevisiae，Ｊ―6001）およびキヤンデイ
ダ・ギリアモンデイ（C.guilliermondii，ATCC
―9058）をそれぞれ抗原として用いて調製した２
種の因子抗体を組合せて用い、その他は実施例１
と同じ操作を試みたところ、野生酵母の検出率は
88％であつた。
実施例 ６
因子抗体として、サツカロミセス・セレビシエ
（S.cerevisiae，Ｊ―6001）、サツカロミセス・ビ
スポラス（S.bisporus，ATCC―0723）およびサ
ツカロミセス・ウバルム（S.uvarum，BSRI―１
―９）をそれぞれ抗原として用いて調製した３種
の因子抗体を組合せて用い、その他は実施例１と
同じ操作を試みたところ、野生酵母の検出率は63
％であつた。
実施例 ７
因子抗体として実施例４で使用した２種の因子
抗体の組合せＡあるいは実施例５で使用した２種
類の因子抗体の組合せＢを用いて、野生酵母72
株、下面ビール酵母63株について試験したとこ
ろ、検出率は野生酵母に対してはA91％，B83％
であり、下面ビール酵母に対してはＡ，Ｂともに
100％であつた。なお、野生酵母のうちサツカロ
ミセス属の菌株32株について同様に試験したとこ
ろ、検出率はA100％，B86％であつた。
参考例 １
殺菌ビール１にサツカロミセス・エリプソイ
デウス（S.ellipsoideus）および下面ビール酵母
の細胞を各々100個づつ添加した試料を膜過し
た後、実施例４で使用した２種の因子抗体の組合
せを用い、螢光抗体法により螢光細胞数を求め、
かつ麦汁培地を使用する培養法により細胞数を求
めたところ、次のような結果を得、両測定法とも
一致していた。
【表】DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a method for detecting microorganisms harmful to brewing, and more particularly to a method for detecting microorganisms harmful to brewing that may appear during the brewing of beer or the like. Currently, Lysine medium and Crystal-Violet medium are widely used to detect wild yeasts that appear during the beer brewing process, but these methods involve culturing and therefore lack speed. Furthermore, as an alternative method, it has been proposed to detect harmful microorganisms by a serological method such as a fluorescent antibody method. Although this method is superior to the above-mentioned methods in terms of speed, it is less convenient because it requires the use of several types of antisera and the combined results of each. For example, the method previously developed by the present inventors uses factor antibodies for identifying pathogenic Candeida spp., which are currently commercially available in Japan, but it requires the use of five types of factor antibodies, making the operation complicated. Therefore, it cannot be said to be an effective method. The present invention overcomes the above-mentioned drawbacks and provides a method for quickly and conveniently detecting microorganisms harmful to brewing by using a single factor antibody. The present invention involves preparing a factor antibody by absorbing an antiserum obtained using yeast belonging to the genus Candeida or Satucharomyces with brewer's yeast, and bringing the factor antibody into contact with a test yeast. Determining the properties of the yeast based on whether or not it occurs,
This detects microorganisms that are harmful to brewing. The present inventors have conducted extensive research to develop a method for quickly and accurately detecting harmful microorganisms generated during the brewing process using a single factor antibody. It has been found that a factor antibody prepared by absorbing the antiserum using brewer's yeast as an absorption bacterium is suitable for the purpose of the present invention. The antiserum used is Candida guilliermondi (ATCC-
9058), Saccharomyces cerevisiae J-6001, S.pastrorianus,
BSRI-1-12), Satucharomyces rosei (S.
rosei, ATCC-0428), S.bisporus (ATCC-0723) and S.uvarum (BSRI-1-)
9) are obtained by immunizing rabbits using yeast as an antigen. A factor antibody is prepared by immunizing a rabbit with the above bacterial strain as an antigen and absorbing the antiserum obtained with brewer's yeast. Brewing yeast may be selected depending on the purpose of use; for example, yeast for beer brewing includes bottom beer yeast and top beer yeast, and other types include sake yeast, wine yeast, whiskey yeast, alcohol yeast, and the like. A single factor antibody may be used, or two or more may be used in combination. In particular, a combination of a factor antibody using Satucharomyces pastorianus as an antigen and a factor antibody using Satucharomyces bisporus as an antigen, a factor antibody using Satucharomyces cerevisiae as an antigen, and a factor antibody using Candeida guilliamondei as an antigen. A combination of these can detect harmful microorganisms at a very high rate. When the factor antibody according to the present invention is brought into contact with a test yeast, an agglutination reaction occurs except when the test yeast is a brewing yeast used as an absorption bacterium. Such behavior of the test yeast is due to differences in the antigenic structure of the yeast, and according to the present invention, microorganisms harmful to brewing can be quickly and accurately detected using only a single factor antibody. I can do it. Contact between the factor antibody and the test yeast can be carried out on a slide glass, making it possible to confirm the presence or absence of an agglutination reaction in a short time. Furthermore, if necessary, a large number of test yeasts can be tested at the same time. The factor antibodies used in the method of the present invention can also be effectively used in known fluorescent antibody methods and culture methods. Next, examples of the present invention will be shown. Example 1 S.bisporus (S.bisporus,
Antiserum was obtained using S. carlsbergensis (ATCC-0723) as an antigen, and the antiserum was used as an antigen for S.
4778) and prepared the factor antibody by a conventional method. Next, when slide agglutination reaction was attempted on 16 wild yeast strains using this factor antibody, the detection rate was 75%. Example 2 S. uvarum (BSRI-1-9) instead of S. bisporus
Slide agglutination reaction was attempted using the same method as in Example 1, except that the detection rate was 45%.
It was hot. Example 3 C. guilliermondii (C. guilliermondii,
When a slide agglutination reaction was attempted in the same manner as in Example 1 except that ATCC-9058) was used, the detection rate was 69%. Example 4 As factor antibodies, S. pastorianus (BSRI-1-12) and S. bisporus (ATCC
-0723) respectively as antigens.
A combination of species factor antibodies was used, and the rest was as in Example 1.
When I tried the same operation as , the detection rate of wild yeast was
It was 94%. Example 5 S. cerevisiae (J-6001) and C. guilliermondii (ATCC) were used as factor antibodies.
-9058) were prepared using each as an antigen.
A combination of species factor antibodies was used, and the rest was as in Example 1.
When I tried the same operation as above, the detection rate of wild yeast was
It was 88%. Example 6 As factor antibodies, S. cerevisiae (J-6001), S. bisporus (S. bisporus, ATCC-0723), and S. uvarum (BSRI-1)
-9) respectively as antigens, and using a combination of three types of factor antibodies prepared, and performing the same procedure as in Example 1, the wild yeast detection rate was 63.
It was %. Example 7 Wild yeast 72
When testing 63 strains of bottom brewer's yeast, the detection rate was 91% for wild yeast and 83% for B.
For bottom brewer's yeast, both A and B
It was 100%. When 32 strains of wild yeast belonging to the genus Satucharomyces were similarly tested, the detection rate was 100% for A and 86% for B. Reference Example 1 A sample obtained by adding 100 cells each of S. ellipsoideus and S. ellipsoideus to sterilized beer 1 was filtered through a membrane, and then the combination of the two factor antibodies used in Example 4 was obtained. Determine the number of fluorescent cells by fluorescent antibody method using
When the number of cells was determined by a culture method using a wort medium, the following results were obtained, and both measurement methods were in agreement. 【table】
Claims (1)
属する酵母を抗原として得た抗血清を醸造用酵母
で吸収処理して因子抗体を調製し、該因子抗体を
酵母と接触させることを特徴とする醸造に有害な
微生物の検出方法。 ２ キヤンデイダ属もしくはサツカロミセス属に
属する酵母がキヤンデイダ・ギリアモンデイ
（Candida guilliermondii，ATCC―9058）、サツ
カロミセス・セレビシエ―（Saccharomyces
cerevisiae Ｊ―6001）、サツカロミセス・パスト
リアヌス（S.Pastorianus，BSRI―１―12）、サ
ツカロミセス・ロゼイ（S.rosei，ATCC―
0428）、サツカロミセス・ビスポラス（S.
bisporus，ATCC―0723）およびサツカロミセ
ス・ウバルム（S.uvarum，BSRI―１―９）の中
から選ばれた１種もしくは２種以上の酵母である
特許請求の範囲第１項記載の検出方法。 ３ 醸造用酵母がビール酵母である特許請求の範
囲第１項記載の検出方法。[Claims] 1. Antiserum obtained using yeast belonging to the genus Candeida or Satucharomyces as an antigen is absorbed and treated with brewer's yeast to prepare a factor antibody, and the factor antibody is brought into contact with the yeast. How to detect microorganisms harmful to brewing. 2 Yeasts belonging to the genus Candida or Saccharomyces include Candida guilliermondii (ATCC-9058) and Saccharomyces cerevisiae.
cerevisiae J-6001), S.Pastorianus (BSRI-1-12), S.rosei (ATCC-)
0428), Satucharomyces bisporus (S.
The detection method according to claim 1, which is one or more yeasts selected from S. bisporus, ATCC-0723) and S. uvarum, BSRI-1-9. 3. The detection method according to claim 1, wherein the brewing yeast is beer yeast.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP14272878A JPS5568299A (en) | 1978-11-18 | 1978-11-18 | Method of detecting microorganism harmful to brewage |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP14272878A JPS5568299A (en) | 1978-11-18 | 1978-11-18 | Method of detecting microorganism harmful to brewage |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPS5568299A JPS5568299A (en) | 1980-05-22 |
| JPS6142825B2 true JPS6142825B2 (en) | 1986-09-24 |
Family
ID=15322197
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP14272878A Granted JPS5568299A (en) | 1978-11-18 | 1978-11-18 | Method of detecting microorganism harmful to brewage |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPS5568299A (en) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| AU547928B2 (en) * | 1981-11-03 | 1985-11-14 | Sutka, K. | Antigenic preparation from candida guilliermondii |
| FR2598513B1 (en) * | 1986-02-06 | 1991-12-13 | Chemunex | NOVEL ANTIBODY ANTIBODIES CAPABLE OF RECOGNIZING MULTIPLE YEASTS, HYBRID CELL LINES PRODUCING SUCH ANTIBODIES, THEIR PREPARATION AND THEIR USES AND APPLICATIONS IN DETECTION AND POSSIBLY IN YEAST NUMBERING |
-
1978
- 1978-11-18 JP JP14272878A patent/JPS5568299A/en active Granted
Also Published As
| Publication number | Publication date |
|---|---|
| JPS5568299A (en) | 1980-05-22 |
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