CN100447234C - Method of simultaneously separating and cultivating pathogenicity Neisseria and mycoplasma - Google Patents
Method of simultaneously separating and cultivating pathogenicity Neisseria and mycoplasma Download PDFInfo
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- CN100447234C CN100447234C CNB200510024388XA CN200510024388A CN100447234C CN 100447234 C CN100447234 C CN 100447234C CN B200510024388X A CNB200510024388X A CN B200510024388XA CN 200510024388 A CN200510024388 A CN 200510024388A CN 100447234 C CN100447234 C CN 100447234C
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Abstract
The present invention relates to a method of simultaneously separating and culturing pathogenic neisseria and mycoplasma. The method comprises the following steps: (1) inoculation: firstly, a material is taken from a suspect patient when the suspect patient has physical examination, and then the material is directly inoculated on a culture medium of an inner chip of a sealed biological culturing assembly; (2) culture: a gas generation sheet is put in the assembly, the assembly is covered with a sealing cover, and culture is carried out for 24 to 72 hours at 30 to 40 DEG C; (3) separation and observation: the pathogenic neisseria can generate a needle head-shaped macroscopic transparent bacterial colony, and a brown bacterial colony on the pathogenic mycoplasma can be observed under a microscope of 40 to 100 times. Compared with the prior art, the present invention has the advantages of simple operation, low cost, etc.
Description
Technical field
The present invention relates to the method for a kind of while separation and Culture pathogenicity Neisseria and mycoplasma, this method can be used on the clinical medicine suspicious patient body being looked into sample and carries out separation and Culture pathogenicity Neisseria (Diplococcus gonorrhoeae and meningococcus) and mycoplasma (Ureaplasma urealyticum, mycoplasma hominis and mycoplasma pneumoniae).
Background technology
The mankind are infected and suffer from transmissible disease by many invasive organisms.Need on the clinical medicine suspicious infected patient is carried out clear and definite pathogeny body analysis and test, carry out the Clinics and Practices of disease to help the doctor.Lab investigation to microorganism separation transmissible disease pathogeny body must rely on from suspicious patient's health check-up sample and be separated to pathogenic microorganism, determine the kind of pathogenic micro-organism, select which kind of antiseptic-germicide to treat to help the doctor.The means of conventional or traditional suspicious pathogenic micro-organism of separation and Culture and method comprise that employing flat garden shape plate or three-dimensional oval plate carry out separation and Culture to a kind of suspicious pathogenic micro-organism, generally need 2～3 kinds of substratum, and need at CO
2Carry out in the incubator, the circular plate of above-mentioned routine divides the loam cake two portions of going to the bottom, and does not seal.There are defectives such as complicated operation, cost height in the method for this traditional suspicious pathogenic micro-organism of separation and Culture, need improve and improve, thereby guarantee specificity, susceptibility and repetition stability to clinical diagnosis.
Summary of the invention
Purpose of the present invention is exactly that a kind of method simple to operate, separation and Culture pathogenicity Neisseria and mycoplasma when cost is low is provided in order to overcome the defective that above-mentioned prior art exists.
Purpose of the present invention can be achieved through the following technical solutions: the method for a kind of while separation and Culture pathogenicity Neisseria and mycoplasma is characterized in that this method may further comprise the steps:
(1) inoculation is drawn materials during at first from suspicious patient's corpse or other object for laboratory examination and chemical testing, and direct inoculation is on the substratum of the interior chip of stopping property biological culture assembly then;
(2) cultivate, in this assembly, place gas generant tablet and cover sealing cover, cultivated 24～72 hours down at 30～40 ℃;
(3) observe separation, pathogenicity Neisseria can grow the transparent bacterium colony of macroscopic syringe needle shape, and pathogenic mycoplasma can be observed brown bacterium colony under 40～100 power microscopes.
Described stopping property biological culture assembly is by transparent outer tube, and with the supporting sealing cover of this outer tube, and interior chip forms, and described outer tube is flats.
The formulation by weight and the compound method thereof of described substratum are as follows: 4% agar, add 10% histocyte nutrient solution, 2% animal serum, 10% yeast extract, 0.1% phenol red, 0.1% urea arginine, 0.1% manganous sulfate, 0.1% fungistat, surplus is a distilled water, transfer pH to 6 ± 0.5, aseptic watering on interior chip waited to solidify and got final product.
Described fungistat comprises polymyxin 7.5 μ g/ml, amphotericin B 6 μ g/ml and Viola crystallinas.
Described culture temperature is preferably 36～38 ℃, and described incubation time is preferably 24～48 hours.
Described gas generant tablet comprises carbonic acid gas generation sheet, micro amount of oxygen generation sheet or anaerobism generation sheet.
Compared with prior art, the present invention has following characteristics: 1, can separate two kinds of different microorganisms (pathogenicity Neisseria and mycoplasma) simultaneously, especially in the clinical experiment diagnosis, promising diagnosis and differential diagnosis gonorrhoea and non-pouring mycoplasma provide scientific basis; 2, when observations, but not only the naked eyes direct viewing cultivate and biochemical reaction (mycoplasma product alkali) can also examine under a microscope the mycoplasma bacterium colony; 3, use CO
2Take place to save CO when sheet replaces the out-of-date methods cultivation
2Incubator is operated very easyly, helps not having CO
2The hospitals at different levels of incubator equipment use.4, saved the usage quantity of substratum greatly, the usage quantity of substratum only is 1/3rd of a traditional method; 5, the present invention can directly finish the method comparison of pathogenicity Neisseria and mycoplasma in the assembly of stopping property.
Embodiment
The invention will be further described below in conjunction with embodiment.
Embodiment 1
The method of a kind of while separation and Culture pathogenicity Neisseria and mycoplasma, this method may further comprise the steps:
(1) inoculation is drawn materials during at first from suspicious patient's corpse or other object for laboratory examination and chemical testing, and direct inoculation is on the substratum of the interior chip of stopping property biological culture assembly then;
(2) cultivate, in this assembly, place CO
2Sheet, little burst size oxygen generation sheet taking place and cover sealing cover, cultivated 72 hours down at 30 ℃;
(3) observe separation, pathogenicity Neisseria can grow the transparent bacterium colony of macroscopic syringe needle shape, and pathogenic mycoplasma can be observed brown bacterium colony under 40～100 power microscopes.
Described stopping property biological culture assembly is by transparent outer tube, and with the supporting sealing cover of this outer tube, and interior chip forms, and described outer tube is flats.
The formulation by weight and the compound method thereof of described substratum are as follows: 4% agar, add 10% histocyte nutrient solution, 2% animal serum, 10% yeast extract, 0.1% phenol red, 0.1% urea arginine, 0.1% manganous sulfate, 0.1% fungistat, surplus is a distilled water, transfer pH to 6 ± 0.5, aseptic watering on interior chip waited to solidify and got final product.Described fungistat comprises polymyxin 7.5 μ g/ml, amphotericin B 6 μ g/ml and Viola crystallinas.
Embodiment 2
The method of a kind of while separation and Culture pathogenicity Neisseria and mycoplasma, this method may further comprise the steps:
(1) inoculation is drawn materials during at first from suspicious patient's corpse or other object for laboratory examination and chemical testing, and direct inoculation is on the substratum of the interior chip of stopping property biological culture assembly then;
(2) cultivate, in this assembly, place CO
2Sheet, anaerobism generation sheet taking place and cover sealing cover, cultivated 24 hours down at 40 ℃;
(3) observe separation, pathogenicity Neisseria can grow the transparent bacterium colony of macroscopic syringe needle shape, and pathogenic mycoplasma can be observed brown bacterium colony under 40～100 power microscopes.
Described stopping property biological culture assembly is by transparent outer tube, and with the supporting sealing cover of this outer tube, and interior chip forms, and described outer tube is flats.
The formulation by weight and the compound method thereof of described substratum are as follows: 4% agar, add 10% histocyte nutrient solution, 2% animal serum, 10% yeast extract, 0.1% phenol red, 0.1% urea arginine, 0.1% manganous sulfate, 0.1% fungistat, surplus is a distilled water, transfer pH to 6 ± 0.5, aseptic watering on interior chip waited to solidify and got final product.Described fungistat comprises polymyxin 7.5 μ g/ml, amphotericin B 6 μ g/ml and Viola crystallinas.
Application examples 1
The venereal disease outpatient service prescription on individual diagnosis person of hospital, for being arranged, gonorrhoea or non-gonococcal urea road inflammation used this novel method by suspicious, draw materials during from suspicious patient's corpse or other object for laboratory examination and chemical testing, direct inoculation will be put as CO in this assembly on the substratum of the interior chip of cultivating assembly more then
2The generation sheet closes the lid and plugs 36～38 ℃ of cultivations 24～72 hours, the result of inoculation culture is observed in taking-up, getting property Neisseria direct inoculation is to transparent interior chip, mycoplasma needs to amplify 40～100 times at microscopically and sees feature, can judge tentatively whether suspicious patient is separated to pathogenic bacterium in experiment, Neisseria, mycoplasma or two kinds of blended pathogenic agent according to turning out feature.
Application examples 2
Obstetrics and gynecology hospital outpatient service prescription on individual diagnosis person, by suspicious be infertility, use this novel method, draw materials during from suspicious patient's corpse or other object for laboratory examination and chemical testing, direct inoculation will be put as CO in this assembly on the substratum of the interior chip of cultivating assembly more then
2The generation sheet closes the lid and plugs 36～38 ℃ of cultivations 24～72 hours, the result of inoculation culture is observed in taking-up, getting property Neisseria direct inoculation is to transparent interior chip, mycoplasma needs to amplify 40～100 times at microscopically and sees feature, can judge tentatively whether suspicious patient is separated to pathogenic bacterium in experiment, Neisseria, mycoplasma or two kinds of blended pathogenic agent according to turning out feature.
Claims (5)
1. method of separation and Culture pathogenicity Neisseria and mycoplasma simultaneously is characterized in that this method may further comprise the steps:
(1) inoculation is drawn materials during at first from suspicious patient's corpse or other object for laboratory examination and chemical testing, and direct inoculation is on the substratum of the interior chip of stopping property biological culture assembly then;
(2) cultivate, in this assembly, place gas generant tablet and cover sealing cover, cultivated 24～72 hours down at 30～40 ℃;
(3) observe separation, pathogenicity Neisseria can grow the transparent bacterium colony of macroscopic syringe needle shape, and pathogenic mycoplasma can be observed brown bacterium colony under 40～100 power microscopes;
Described stopping property biological culture assembly is by transparent outer tube, and with the supporting sealing cover of this outer tube, and interior chip forms, and described outer tube is flats.
2. the method for a kind of while separation and Culture pathogenicity Neisseria according to claim 1 and mycoplasma, it is characterized in that, the formulation by weight and the compound method thereof of described substratum are as follows: 4% agar, add 10% histocyte nutrient solution, 2% animal serum, 10% yeast extract, 0.1% phenol red, 0.1% urea arginine, 0.1% manganous sulfate, 0.1% fungistat, surplus is a distilled water, transfer pH to 6 ± 0.5, aseptic watering on interior chip waited to solidify and got final product.
3. the method for a kind of while separation and Culture pathogenicity Neisseria according to claim 2 and mycoplasma is characterized in that, described fungistat comprises polymyxin 7.5 μ g/ml, amphotericin B 6 μ g/ml and Viola crystallinas.
4. the method for a kind of while separation and Culture pathogenicity Neisseria according to claim 1 and mycoplasma is characterized in that described culture temperature is preferably 36～38 ℃, and described incubation time is preferably 24～48 hours.
5. the method for a kind of while separation and Culture pathogenicity Neisseria according to claim 1 and mycoplasma is characterized in that described gas generant tablet comprises carbonic acid gas generation sheet, micro amount of oxygen generation sheet or anaerobism generation sheet.
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| Application Number | Priority Date | Filing Date | Title |
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| CNB200510024388XA CN100447234C (en) | 2005-03-15 | 2005-03-15 | Method of simultaneously separating and cultivating pathogenicity Neisseria and mycoplasma |
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| Application Number | Priority Date | Filing Date | Title |
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| CNB200510024388XA CN100447234C (en) | 2005-03-15 | 2005-03-15 | Method of simultaneously separating and cultivating pathogenicity Neisseria and mycoplasma |
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| CN1834225A CN1834225A (en) | 2006-09-20 |
| CN100447234C true CN100447234C (en) | 2008-12-31 |
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Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101177668B (en) * | 2007-12-05 | 2010-05-19 | 四川省医学科学院（四川省人民医院） | Novel neisseria gonorrhoeae culture medium and method for making same |
| CN109385381B (en) * | 2018-11-16 | 2019-10-08 | 山东恒辰生物科技有限公司 | A kind of Urogenital Mycoplasma biphasic culture |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1086739C (en) * | 1996-07-05 | 2002-06-26 | 中国科学院动物研究所 | Single tube synchronous three-step PCR checkout method for chlamydia trachomatis and Neisseria gonorrhoeae |
| WO2004026336A1 (en) * | 2002-09-23 | 2004-04-01 | Vital Biotech (Hong Kong) Limited | Improvements in or relating to vaccines |
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2005
- 2005-03-15 CN CNB200510024388XA patent/CN100447234C/en not_active Expired - Fee Related
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1086739C (en) * | 1996-07-05 | 2002-06-26 | 中国科学院动物研究所 | Single tube synchronous three-step PCR checkout method for chlamydia trachomatis and Neisseria gonorrhoeae |
| WO2004026336A1 (en) * | 2002-09-23 | 2004-04-01 | Vital Biotech (Hong Kong) Limited | Improvements in or relating to vaccines |
Non-Patent Citations (2)
| Title |
|---|
| 淋病奈瑟菌、衣原体、支原体检测695例分析. 吴祥林，陈继中，潘晓龙.蚌埠医学院学报，第26卷第3期. 2001 |
| 淋病奈瑟菌、衣原体、支原体检测695例分析. 吴祥林，陈继中，潘晓龙.蚌埠医学院学报，第26卷第3期. 2001 * |
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| CN1834225A (en) | 2006-09-20 |
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Application publication date: 20060920 Assignee: Ningbo four Ming en Biotechnology Co., Ltd. Assignor: Shanghai Enkang Biotechnology Co., Ltd. Contract record no.: 2013330000003 Denomination of invention: Method of simultaneously separating and cultivating pathogenicity Neisseria and mycoplasma Granted publication date: 20081231 License type: Exclusive License Record date: 20130115 |
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Granted publication date: 20081231 Termination date: 20130315 |