CN104450860B - A kind of mycoplasma pneumoniae culture medium - Google Patents
A kind of mycoplasma pneumoniae culture medium Download PDFInfo
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- CN104450860B CN104450860B CN201310425112.7A CN201310425112A CN104450860B CN 104450860 B CN104450860 B CN 104450860B CN 201310425112 A CN201310425112 A CN 201310425112A CN 104450860 B CN104450860 B CN 104450860B
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Abstract
The invention discloses a kind of mycoplasma pneumoniae culture medium, it is a kind of mycoplasma pneumoniae biphasic culture, it is to be loaded with a solid medium and a fluid nutrient medium in a vessel, described solid medium shows an inclined-plane inside described container, while the 1/4~1/2 of the inclined-plane is immersed in described fluid nutrient medium；The container also includes the seal member of the inner space of a salable container；For the component of described solid medium in addition to including 15~25g/L agar, remaining component is identical with the component of described fluid nutrient medium.The culture medium of the present invention is used to detect mycoplasma pneumoniae, improves the specificity of detection, and can obtain monoclonal bacterium colony, the convenient further research work for making downstream based on testing result.
Description
Technical field
The present invention relates to microorganism detection field, and in particular to a kind of mycoplasma pneumoniae culture medium.
Background technology
Mycoplasma pneumoniae（Mycoplasma pneumoniae）Category urea substance, acellular wall and precursor, organelle are few.
Mycoplasma pneumoniae mainly causes Eaton agent pneumonia, although Eaton agent pneumonia patient's self-induction symptom is heavier, the general nothing of chest physical examination
Obvious abnormal sign.In addition to the performance of respiratory system, Eaton agent pneumonia can occur together multisystem, multiple organ injury.Due to mycoplasma
The clinical manifestation of pneumonia and chest X-ray do not have characteristic, can not be paid a home visit only according to clinical manifestation and chest X-ray
It is disconnected.To clarify a diagnosis, it is necessary to carry out the detection of pathogen.
At present, the detection of domestic Eaton agent pneumonia relies primarily on Serologic detection, but the specificity of this method and sensitivity
It is poor.Another detection method of mycoplasma pneumoniae is cultivation.The principle of liquid culture method is that mycoplasma pneumoniae decomposes Portugal
Make the pH value of culture medium reduce after grape sugar, make phenolic red indicator yellow by red change.Because some penicillin resistant microorganisms may also divide
Solve glucose, although therefore the method it is more quick, higher false positive be present.And utilize solid medium detection pneumonia branch former
Body although can obtain single bacterium colony, specificity it is very strong, but report detection time it is oversize, be unfavorable for clinic quick diagnosis.Therefore, originally
Need the mycoplasma pneumoniae detection method that the high and can simultaneously of detection specificity enough reaches quick detection purpose badly at present in field.
The content of the invention
The technical problems to be solved by the invention are at present in the detection of mycoplasma pneumoniae, liquid culture method is present
False positive, and use solid medium to detect too time-consuming defect, there is provided a kind of new mycoplasma pneumoniae culture medium, its
It is a kind of mycoplasma pneumoniae biphasic culture.The mycoplasma pneumoniae biphasic culture of the present invention improves the specificity of detection, and
Monoclonal bacterium colony is can obtain, so as to be advantageous to clinical diagnosis.Also, this culture medium only needs a sample inoculation once, so as to
It is easier to laboratory operation, convenient popularization and application in practice.
One of technical scheme provided by the invention is：A kind of mycoplasma pneumoniae culture medium, it is that a kind of mycoplasma pneumoniae is double
Phase culture medium, to be loaded with a solid medium and a fluid nutrient medium in a vessel, described solid medium exists for it
An inclined-plane is showed inside described container, while the 1/4~1/2 of the inclined-plane is immersed in described fluid nutrient medium；
The container also includes the seal member of the inner space of a salable container；
The component of described fluid nutrient medium includes 40~60g/L beef extracts, 7~12g/L peptones, 4~6g/L chlorinations
Sodium, 70~130mL/L cow's serums, 10~20mL/L DMEM Liquid Cultures matrix, 8~15mL/L mass percentage by volume are 10
~40% yeast extract, 3~7g/L glucose, 0.3~0.7mL/L mass percentage by volume be 0.2~0.6% phenolic red indicator,
0.005~0.015g/L Nysfungins, 600~1500IU/L penicillin and 0.5~1.0g/L nicotinamide adenine dinucleotides；
Described DMEM Liquid Culture matrix is made by the following method：Low sugar DMEM powder is dissolved in water and forms 10g/mL concentration；
The component of described solid medium is in addition to including 15~25g/L agar, remaining component and described Liquid Culture
The component of base is identical.
In the present invention, described container can be container of this area conventionally used for splendid attire culture medium, such as transparent glass
Pipe or plastic tube, preferably test tube.It is preferred that a diameter of 3~7cm of described container, the length of described container for 6~
15cm.More preferably, a diameter of 5~7cm of described container, the length of described container is 8~10cm.
In the present invention, described seal member is that this area is conventional, as long as it can be close by the inner space of the container
Envelope, makes the inner space of the container be hedged off from the outer world, and prevents the miscellaneous bacteria in air from entering the inside of the container, while can also
Prevent the moisture evaporation in the culture medium inside container.It is preferred that described seal member is can be tight with the container
The lid of closely connected conjunction.Described seal member can also be the plastic seal membrana oralis for sealing.
In the present invention, it is preferred that the component of described fluid nutrient medium includes 45~55g/L beef extracts, 8~10g/L eggs
White peptone, 5~6g/L sodium chloride, 80~110mL/L cow's serums, 12~18mL/L DMEM Liquid Cultures matrix, 10~12mL/L matter
Amount percentage by volume is 20~30% yeast extracts, 4~6g/L glucose, 0.4~0.6mL/L mass percentage by volumes be 0.3~
0.6% phenolic red indicator, 0.008~0.012g/L Nysfungins, 800~1200IU/L penicillin and 0.5~1.0g/L niacinamides
Adenine-dinucleotide.
In the present invention, it is preferred that described solid medium shows an inclined-plane, while institute inside described container
The 1/3 of inclined-plane is stated to be immersed in described fluid nutrient medium.
In the present invention, the preparation method of described solid medium is described in the routine of this area.It is preferred that described solid
Culture medium is prepared by following preparation method：The each component of described solid medium is simply mixed, dissolved with water, is adjusted
Filtration sterilization is carried out after pH again, be then heated to 60~80 DEG C dissolve after, go in described container and be cooled into inclined-plane.It is described
Regulation pH be by pH adjust to this area detect mycoplasma pneumoniae culture medium conventional pH value range, preferably 7.5
~8.0, more preferably it is 7.8.Described water is the conventional water for being used to prepare culture medium in this area, preferably distilled water.Institute
The filtration sterilization stated can also replace with other sterilization methods, as long as it does not destroy the nutritional ingredient of the solid medium i.e.
Can.
Similarly, the preparation method of described fluid nutrient medium is described in the routine of this area.It is preferred that described liquid training
Base is supported to be prepared by following preparation method：The each component of described fluid nutrient medium is simply mixed, dissolved with water, adjusts pH
Carry out filtration sterilization again afterwards, be then heated to 60~80 DEG C dissolve after, go in the container, and cover the solid medium
The 1/4~1/2 of the inclined-plane of formation.Described regulation pH be by pH adjust to this area detect mycoplasma pneumoniae culture medium it is normal
The pH value range of rule, preferably 7.5~8.0, more preferably it is 7.8.Described water is that the conventional preparation that is used in this area is cultivated
The water of base, preferably distilled water.Described filtration sterilization can also replace with other sterilization methods, as long as it does not destroy institute
State the nutritional ingredient of fluid nutrient medium.
The present invention a preferred embodiments be：
A kind of mycoplasma pneumoniae culture medium, it is a kind of mycoplasma pneumoniae biphasic culture, and it is to be in a diameter
5cm, length are that some solid medium and a part of fluid nutrient medium are contained in 8cm lucite pipe, and described consolidates
Body culture medium forms an inclined-plane in described lucite pipe, and the 1/3 of described inclined-plane is immersed in described Liquid Culture
In base, described lucite pipe also includes a lid that can be brought into close contact the lucite pipe；
The component of described fluid nutrient medium includes 45g/L beef extracts, 8g/L peptones, 5g/L sodium chloride, 80mL/L oxen
Serum, 12mL/LDMEM Liquid Cultures matrix, 10mL/L mass percentage by volume be 30% yeast extract, 4g/L glucose,
0.4mL mass percentage by volume is 0.6% phenolic red indicator, 0.008g/L Nysfungins, 800IU/L penicillin and 0.5g/L Buddhist nun gram
Acid amides adenine-dinucleotide；
The component of described solid medium is in addition to including 15.0g/L agar, remaining component and described fluid nutrient medium
Component it is identical.
The present invention another preferred embodiments be：
A kind of mycoplasma pneumoniae culture medium, it is a kind of mycoplasma pneumoniae biphasic culture, and it is to be in a diameter
7cm, length are that some solid medium and a part of fluid nutrient medium are contained in 10cm transparent glass tube, and described consolidates
Body culture medium forms an inclined-plane in described transparent glass tube, and the 1/3 of described inclined-plane is immersed in described Liquid Culture
In base, described transparent glass tube also includes a plastic closures film that can seal the transparent glass tube inner space；
The component of described fluid nutrient medium includes 55g/L beef extracts, 10g/L peptones, 6g/L sodium chloride, 110mL/L
Cow's serum, 18ml/L DMEM Liquid Cultures matrix, 12mL/L mass percentage by volume be 20% yeast extract, 6g/L glucose,
0.6mL mass percentage by volume is 0.3% phenolic red indicator, 0.012g/L Nysfungins, 1200IU/L penicillin and 1.0g/L Buddhist nun gram
Acid amides adenine-dinucleotide；
The component of described solid medium is in addition to including 25g/L agar, remaining component and described fluid nutrient medium
Component is identical.
The two of technical scheme provided by the invention are：It is a kind of that mycoplasma pneumoniae is detected with foregoing mycoplasma pneumoniae culture medium
Method, it comprises the following steps：
（1）Sample to be tested is inoculated in described fluid nutrient medium, jog mixes rear-inclined container, is mixed with a part
The fluid nutrient medium of sample to be tested is laid on described solid medium, that is, completes inoculation；
（2）By step（1）The culture vessel for completing to be inoculated with, which is placed in incubator, to be cultivated, and cultivation temperature is 36~38 DEG C, training
Support the CO in case2Concentration is 5%~10%, and described percentage is percent by volume；
（3）Culture judges Preliminary detection result in 24~48 hours, continues culture to positive sample 3~7 days, obtains and finally detect
As a result；
The standard of described judgement Preliminary detection result is：If fluid nutrient medium reddens and Clear ＆ Transparent, can tentatively report
Accuse positive；The criterion of described final detection result is：If there is fried egg sample colony growth on solid slope, and meet lung
Scorching mycoplasma biochemical character, Final Report mycoplasma pneumoniae are positive.
In the present invention, step（2）Described cultivation temperature is preferably 37 DEG C, the CO in the incubator2Concentration is preferable
Ground is 8%.
In the present invention, for step（3）Described in final detection result be positive sample, can be in solid medium shape
Into inclined-plane on obtain monoclonal bacterium colony, to facilitate the further research work in downstream.
It on the basis of common sense in the field is met, above-mentioned each optimum condition, can be combined, it is each preferably real to produce the present invention
Example.
Agents useful for same and raw material of the present invention are commercially available.
The positive effect of the present invention is：
Compared with traditional mycoplasma pneumoniae fluid nutrient medium, mycoplasma pneumoniae biphasic culture of the invention improves inspection
The specificity of survey, and monoclonal bacterium colony is can obtain, the convenient further research work for making downstream based on testing result.In general liquid
Body culture medium detection method false positive is up to 5~15%, and very big inconvenience is brought to actual diagnosis.It is and former with traditional pneumonia branch
Body solid medium is compared, and the culture medium can obtain Preliminary detection result in advance.General traditional solid medium is examined
Surveying result needs 7~10 days, present invention only requires 1~5 day, so as to be advantageous to accelerate diagnosis speed in practical application.This hair
Bright mycoplasma pneumoniae biphasic culture only needs a sample inoculation once, convenient in reality so as to be easier to laboratory operation
Trample middle popularization and application.
Brief description of the drawings
Fig. 1 is the schematic diagram of mycoplasma pneumoniae biphasic culture of the present invention.
Embodiment
The present invention is further illustrated below by the mode of embodiment, but does not therefore limit the present invention to described reality
Apply among a scope.The experimental method of unreceipted actual conditions in the following example, conventionally and condition, or according to business
Product specification selects.
In following embodiments, low sugar DMEM powder is purchased from Sigma companies.
Cow's serum is purchased from Sigma companies.
Beef extract is purchased from Sigma companies.
Peptone is purchased from Sigma companies.
Other reagents or raw material are conventional commercial commodity unless otherwise instructed.
The preparation of the mycoplasma pneumoniae biphasic culture of embodiment 1
It is standby that DMEM powder is dissolved in water formation 10g/L concentration.Weigh or 50g beef extracts, 10g albumen are measured with measurer
Peptone, 5g sodium chloride, 100mL cow's serums, 10mL DMEM Liquid Cultures matrix, 10mL25% yeast extracts, 5g glucose,
0.5mL0.4% phenolic red indicators, 0.01g Nysfungins, 1000IU penicillin and 0.5g/L nicotinamide adenine dinucleotides, will
Each component is added in 1000mL distilled water after being simply mixed, and adjusts pH to 7.8, then degerming with 0.22 μm of bacteriological filtration membrane filtration, so
After be heated to 60 DEG C of dissolve fully each components, then the culture medium is divided into two parts of account for total amount respectively 3/4 and 1/4, wherein accounting for
3/4 a addition 11g agar powders, after heat of solution to be added, it is 5cm to be transferred to a sterile diameter, and length is the transparent of 10cm
In plastic tube, plastic tube is tilted to be cooled into an inclined-plane, i.e. solid culture base section.Again by remaining without agar powder
Culture medium is poured into shape plastic tube in the slope, and covers the 1/3 of this inclined-plane, i.e. Liquid Culture base section.Plastic tube is with one
Individual cap covers its oral area that can be brought into close contact with it.It is specific as shown in Figure 1.In Fig. 1, described mycoplasma pneumoniae two-phase
Culture medium includes the inclined-plane and lid 1 that container 2, fluid nutrient medium 3, solid medium 4 are formed.
The preparation of the mycoplasma pneumoniae biphasic culture of embodiment 2
It is standby that DMEM powder is dissolved in water formation 10g/L concentration.Weigh or with measurer measure 45g beef extracts, 8g peptones,
5g sodium chloride, 80mL cow's serums, 12mL DMEM Liquid Cultures matrix, 12mL20% yeast extracts, 4g glucose, 0.4mL0.3%
Phenolic red indicator, 0.008g Nysfungins, 800IU penicillin and 0.5g/L nicotinamide adenine dinucleotides, by each component letter
Added after single mixing in 1000mL distilled water, adjust pH to 7.5, it is then degerming with 0.22 μm of bacteriological filtration membrane filtration, it is then heated to
60 DEG C of dissolve fully each components, then the culture medium is divided into two parts of account for total amount respectively 3/4 and 1/4, wherein accounting for 3/4 portion
15g agar powders are added, after heat of solution to be added, it is 3cm to be transferred to a sterile diameter, and length is in 7cm transparent glass tube, is inclined
Oblique glass tube is to be cooled into an inclined-plane, i.e. solid culture base section.The remaining culture medium without agar powder is poured into again
In shape glass tube in the slope, and the 1/3 of this inclined-plane is covered, i.e. Liquid Culture base section.Glass tube can be with it with one
Its oral area of the cap covers being brought into close contact.
The preparation of the mycoplasma pneumoniae biphasic culture of embodiment 3
It is standby that DMEM powder is dissolved in water formation 10g/L concentration.Weigh or 55g beef extracts, 10g albumen are measured with measurer
Peptone, 6g sodium chloride, 110mL cow's serums, 18mL DMEM Liquid Cultures matrix, 10mL30% yeast extracts, 6g glucose,
0.6mL0.6% phenolic red indicators, 0.012g Nysfungins, 1200IU penicillin and 1.0g/L nicotinamide adenine dinucleotides,
Added after each component is simply mixed in 1000mL distilled water, adjust pH to 8.0, it is then degerming with 0.22 μm of bacteriological filtration membrane filtration,
60 DEG C of dissolve fully each components are then heated to, then the culture medium is divided into two parts of account for total amount respectively 3/4 and 1/4, wherein
3/4 a addition 18g agar powders are accounted for, after heat of solution to be added, it is 7cm to be transferred to a sterile diameter, and length is the saturating of 15cm
In bright plastic tube, plastic tube is tilted to be cooled into an inclined-plane, i.e. solid culture base section.Again agar powder is free of by remaining
Culture medium pour into shape plastic tube in the slope, and cover the 1/3 of this inclined-plane, i.e. Liquid Culture base section.Plastic tube with
One cap covers its oral area that can be brought into close contact with it.
The preparation of the mycoplasma pneumoniae biphasic culture of embodiment 4
It is standby that DMEM powder is dissolved in water formation 10g/L concentration.Weigh or with measurer measure 50g beef extracts, 9g peptones,
6g sodium chloride, 100mL cow's serums, 18mL DMEM Liquid Cultures matrix, 10mL30% yeast extracts, 6g glucose, 0.6mL0.6%
Phenolic red indicator, 0.012g Nysfungins, 1000IU penicillin and 1.0g/L nicotinamide adenine dinucleotides, by each component letter
Added after single mixing in 1000mL distilled water, adjust pH to 7.8, it is then degerming with 0.22 μm of bacteriological filtration membrane filtration, it is then heated to
60 DEG C of dissolve fully each components, then the culture medium is divided into two parts of account for total amount respectively 3/4 and 1/4, wherein accounting for 3/4 portion
18g agar powders are added, after heat of solution to be added, it is 5cm to be transferred to a sterile diameter, and length is in 8cm test tube, tilts test tube
To be cooled into an inclined-plane, i.e. solid culture base section.It is oblique that the remaining culture medium without agar powder is poured into the formation again
In the test tube in face, and the 1/3 of this inclined-plane is covered, i.e. Liquid Culture base section.Test tube is with a lid that can be brought into close contact with it
Son covers its oral area.
Application of the biphasic culture of embodiment 5 in mycoplasma pneumoniae is detected
1st, the process of detection specifically comprises the following steps：
（1）With sterile disposable swab collect specimen（Subject's nasopharynx secretions or sputum specimen）Afterwards, it is seeded to
In the fluid nutrient medium of biphasic culture（Swab is corresponded in liquid phase part and ground for several times on chamber wall）, then mix, and incline
Oblique liquid phase for several times, to flood culture medium slant, completes inoculation.The culture medium for completing inoculation is placed into incubator and cultivated.
Condition of culture is：37℃、8%CO2。
（2）As a result observation：
（a）Criterion：, can be positive with preliminary report if fluid nutrient medium reddens and Clear ＆ Transparent in 24~48h；Report
Accuse positive sample and continue culture 3~7 days, if there is fried egg sample colony growth on solid slope, and meet mycoplasma pneumoniae life
Change feature, Final Report mycoplasma pneumoniae is positive；If it exceeds 48h fluid nutrient mediums are still non-discolouring, mycoplasma pneumoniae can be reported
It is negative.
2nd, sample to be tested is detected with the culture medium obtained by embodiment 1, and result is analyzed：
50 subjects are chosen to be tested, it is as a result positive for 15 reports, reddened respectively at 16~48h fluid nutrient mediums
And it is Clear ＆ Transparent, and Final Report is positive when cultivating 4~5 days, and monoclonal bacterium colony is obtained, separately there are 35 reports negative.Tool
Body testing result is as shown in table 1.
The testing result of table 1
It can be seen from the data of table 1 present invention biphasic culture it is very convenient in actual applications, culture 16~
During 48h can preliminary report testing result, and cultivate 4~5 days when can report final detection result.
Claims (10)
1. a kind of mycoplasma pneumoniae culture medium, it is characterised in that it is a kind of mycoplasma pneumoniae biphasic culture, and it is at one
A solid medium and a fluid nutrient medium are loaded with container, described solid medium shows inside described container
One inclined-plane, while the 1/4~1/2 of the inclined-plane is immersed in described fluid nutrient medium；The container can also including one
Seal the seal member of the inner space of the container；
The component of described fluid nutrient medium includes 40~60g/L beef extracts, 7~12g/L peptones, 4~6g/L sodium chloride, 70
~130mL/L cow's serums, 10~20mL/L DMEM Liquid Cultures matrix, 8~15mL/L mass percentage by volume are 10~40%
Yeast extract, 3~7g/L glucose, 0.3~0.7mL/L mass percentage by volume are 0.2~0.6% phenolic red indicator, 0.005
~0.015g/L Nysfungins, 600~1500IU/L penicillin and 0.5~1.0g/L nicotinamide adenine dinucleotides；It is described
DMEM Liquid Culture matrix be made by the following method：Low sugar DMEM powder is dissolved in water and forms 10g/mL concentration；
The component of described solid medium is in addition to including 15~25g/L agar, remaining component and described fluid nutrient medium
Component is identical.
2. mycoplasma pneumoniae culture medium as claimed in claim 1, it is characterised in that a diameter of 3~7cm of described container,
The length of described container is 6~15cm.
3. mycoplasma pneumoniae culture medium as claimed in claim 1, it is characterised in that described seal member be and the container
The lid or the plastic seal membrana oralis for sealing that can be brought into close contact.
4. mycoplasma pneumoniae culture medium as claimed in claim 1, it is characterised in that the component of described fluid nutrient medium includes
45~55g/L beef extracts, 8~10g/L peptones, 5~6g/L sodium chloride, 80~110mL/L cow's serums, 12~18mL/L
DMEM Liquid Cultures matrix, 10~12mL/L mass percentage by volume are 20~30% yeast extracts, 4~6g/L glucose, 0.4
~0.6mL/L mass percentage by volume be 0.3~0.6% phenolic red indicator, 0.008~0.012g/L Nysfungins, 800~
1200IU/L penicillin and 0.5~1.0g/L nicotinamide adenine dinucleotides.
5. mycoplasma pneumoniae culture medium as claimed in claim 1, it is characterised in that described solid medium by preparing as follows
Method is prepared：The each component of described solid medium is simply mixed, dissolved with water, was filtered out again after adjusting pH
Bacterium, be then heated to 60~80 DEG C dissolve after, go in described container and be cooled into inclined-plane；Described fluid nutrient medium is by such as
Lower preparation method is prepared：The each component of described fluid nutrient medium is simply mixed, dissolved with water, is carried out again after adjusting pH
Filtration sterilization, be then heated to 60~80 DEG C dissolve after, go in the container, and cover the solid medium formed it is oblique
The 1/4~1/2 of face.
6. mycoplasma pneumoniae culture medium as claimed in claim 5, it is characterised in that described regulation pH be by pH adjust to
7.5~8.0.
7. mycoplasma pneumoniae culture medium as claimed in claim 1, it is characterised in that it is a kind of mycoplasma pneumoniae biphasic culture
Base, it in a diameter is that 5cm, length are to contain some solid medium and a part in 8cm lucite pipe that it, which is,
Fluid nutrient medium, described solid medium form an inclined-plane, 1/3 leaching on described inclined-plane in described lucite pipe
Not in described fluid nutrient medium, described lucite pipe also includes a lid that can be brought into close contact the lucite pipe
Son；
The component of described fluid nutrient medium include 45g/L beef extracts, 8g/L peptones, 5g/L sodium chloride, 80mL/L cow's serums,
12mL/LDMEM Liquid Cultures matrix, 10mL/L mass percentage by volume are 30% yeast extract, 4g/L glucose, 0.4mL matter
Amount percentage by volume is 0.6% phenolic red indicator, 0.008g/L Nysfungins, 800IU/L penicillin and 0.5g/L niacinamide glands
Purine dinucleotides；
The component of described solid medium is in addition to including 15.0g/L agar, remaining component and the group of described fluid nutrient medium
Split-phase is same.
8. mycoplasma pneumoniae culture medium as claimed in claim 1, it is characterised in that it is a kind of mycoplasma pneumoniae biphasic culture
Base, it in a diameter is that 7cm, length are to contain some solid medium and a part in 10cm transparent glass tube that it, which is,
Fluid nutrient medium, described solid medium form an inclined-plane, 1/3 leaching on described inclined-plane in described transparent glass tube
Not in described fluid nutrient medium, described transparent glass tube, which also includes one, can seal the transparent glass tube inner space
Plastic closures film；
The component of described fluid nutrient medium includes 55g/L beef extracts, 10g/L peptones, 6g/L sodium chloride, 110mL/L ox bloods
Clearly, 18ml/L DMEM Liquid Cultures matrix, 12mL/L mass percentage by volume be 20% yeast extract, 6g/L glucose,
0.6mL mass percentage by volume is 0.3% phenolic red indicator, 0.012g/L Nysfungins, 1200IU/L penicillin and 1.0g/L Buddhist nun
Gram acid amides adenine-dinucleotide；
The component of described solid medium is in addition to including 25g/L agar, remaining component and the component of described fluid nutrient medium
It is identical.
9. detect pneumonia branch to a kind of non-diagnostic purposes of mycoplasma pneumoniae culture medium with as described in any one of claim 1~8
The method of substance, it comprises the following steps：
(1) sample to be tested is inoculated in described fluid nutrient medium, jog mixes rear-inclined container, is mixed with a part to be measured
The fluid nutrient medium of sample is laid on described solid medium, that is, completes inoculation；
(2) culture vessel that step (1) is completed to be inoculated with is placed in incubator and cultivated, cultivation temperature is 36~38 DEG C, incubator
Interior CO2Concentration is 5%~10%, and described percentage is percent by volume；
(3) cultivate 24~48 hours and judge Preliminary detection result, continue culture to positive sample 3~7 days, obtain finally detection knot
Fruit；
The standard of described judgement Preliminary detection result is：, can be with preliminary report sun if fluid nutrient medium reddens and Clear ＆ Transparent
Property；The criterion of described final detection result is：If there is fried egg sample colony growth on solid slope, and meet pneumonia branch
Substance biochemical character, Final Report mycoplasma pneumoniae are positive.
10. method as claimed in claim 9, it is characterised in that in step (2), described cultivation temperature is 37 DEG C, the training
Support the CO in case2Concentration is 8%.
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| CN105145350B (en) * | 2015-07-22 | 2017-09-15 | 青岛农业大学 | A kind of preparation method and application of solid-liquid two-phase PEG culture mediums |
| CN106350460A (en) * | 2016-11-21 | 2017-01-25 | 四川农业大学 | Mould solid culture medium based on DMEM (Dulbecco Modified Eagle Medium) and preparation method of mould solid culture medium |
| CN106591177B (en) * | 2016-11-29 | 2020-04-21 | 中国农业科学院兰州兽医研究所 | Low-serum efficient culture medium for mycoplasma capricolum goat pneumonia subspecies and preparation method thereof |
| 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 |
| CN111154696A (en) * | 2020-02-12 | 2020-05-15 | 石河子大学 | Mycoplasma hyopneumoniae culture medium and preparation method thereof |
| CN111304279A (en) * | 2020-03-09 | 2020-06-19 | 珠海市银科医学工程股份有限公司 | Mycoplasma identification agar culture medium and preparation method thereof |
| CN115873780A (en) * | 2022-12-10 | 2023-03-31 | 刘鹤然 | Cell culture medium and culture method thereof |
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| CN101838626A (en) * | 2010-05-12 | 2010-09-22 | 上海交通大学 | Bartonia body solid-liquid double-phase slant culture medium as well as preparation method and application method thereof |
| CN103757112A (en) * | 2014-01-15 | 2014-04-30 | 珠海市银科医学工程有限公司 | Mycobacterium separation and culture kit and testing method thereof |
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| 双相培养基检测泌尿生殖道支原体方法及评价;张银旺,等;《现代检验医学杂志》;20081130;第23卷(第6期);第77页左栏第7-13行，第1.2.1.1节-1.2.2.1节，第79页左栏倒数第14行至右栏第10行 * |
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