CN105087751B - Optochin sensitization test culture medium and its preparation method and application - Google Patents
Optochin sensitization test culture medium and its preparation method and application Download PDFInfo
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Abstract
The present invention relates to a kind of optochin sensitization test culture medium and its preparation method and application, the culture medium includes MH meat soups, glucose, distilled water, optochin, blood meal and acid-base indicator；Correspondingly, the drug selected in preparation method includes acid-base indicator；The culture medium can be used for identifying streptococcus pneumonia, can save and measure the work of scraps of paper antibacterial circle diameter, result is directly read according to color change；Optochin sensitization test can be added in bacterial biochemical assay plate, increase the ability of biochemical identification plate identification bacterium, it is convenient to read the offer of optochin reaction result to semi-automatic, robot.
Description
Technical field
The present invention relates to a kind of optochin sensitization test culture mediums and its preparation method and application.
Background technology
Optochin (optochin) is the peaceful trade name of hydrochloric acid Optochin, there is special suppression to streptococcus pneumonia
It makes and uses, nearly all streptococcus pneumonia then acts on other streptococcus Optochin sensitivities without this.Optochin is sensitive
Experiment is to differentiate a kind of common experiment of streptococcus pneumonia, is had great importance in clinical examination.Traditional optochin
The broth culture of bacterium to be checked is spread evenly across on blood agar plate by sensitization test with cotton swab, and one is sticked after slightly dry containing 5 μ
The g/ piece optochin scraps of paper, 35 DEG C of incubation 18-24h, observe result.It is sensitive when inhibition zone diameter is more than 14mm, is inferred as lung
Scorching streptococcus.
The Chinese invention patent application of Publication No. CN101698876A discloses a kind of streptococcus pneumonia sensitization test paper
Piece, main component optochin, distilled water, filter paper（Diameter 6mm）It mixes in proportion and forms a kind of pneumonia through certain technique
Sensitization test (STT) paper for streptococcus.In use, the broth culture of tested bacterium is spread evenly across blood agar plate with sterile swab stick
On, the optochin scraps of paper is taken to be affixed on tablet, in 35 DEG C~37 DEG C cultivate 18~for 24 hours, observation as a result, inhibition zone in 18mm
It is above sensitivity, no inhibition zone or its inhibition zone are less than 10mm for drug resistance.And provide the minimum standard of its judgement：Sensitivity, it is antibacterial
Ring diameter is generally 15-16mm or bigger, if less than 15mm, should add and do bile solubility test to determine.
Traditional optochin sensitization test is to infer whether tested bacterium colony is streptococcus pneumonia by antibacterial circle diameter, this
Method is not easy to using automatic, semi-automatic Bacteria Identification system.
Invention content
For technological deficiency of the prior art more than overcoming, the present invention provides a kind of optochin sensitization test culture
Base, including MH meat soups, glucose, optochin, blood meal and acid-base indicator.
Preferably, the optochin sensitization test culture medium is made of the following component of mass percent content：
MH broth bouillons 25-35%；
Glucose 63-73%；
Optochin 0.25-0.4%；
Blood meal 1.6-2.5%；
Acid-base indicator 0.14%-0.67%.
In any of the above-described scheme, it is preferred that the optochin sensitization test culture medium by mass percent content such as
Under component be made：
MH meat soups 28.64%；
Glucose 68.56%；
Optochin 0.33%；
Blood meal 2.06%；
Acid-base indicator 0.21%.
Acid-base indicator is added in culture medium, can cause the color of culture medium difference can be presented with the variation of pH
Color, consequently facilitating identification.
In any of the above-described scheme, it is preferred that the pH of the culture medium is 7.2-7.5.Since bacterium is in appropriate pH models
It is rapid to enclose interior growth, in reproductive process, glucose can generate bacterium after utilizing, decomposing along with acid product；Culture medium
PH using above range, bacterial growth, breeding are rapid, convenient for identifying the variation of color；If the pH of culture medium uses alkalinity,
The growth of bacterium may be then influenced, utilizing for glucose is influenced and decomposes, and culture medium cannot be made to change colour, None- identified color
Variation.
In any of the above-described scheme, it is preferred that the acid-base indicator is phenol red and bromocresol purple mixed indicator.It adopts
By the use of phenol red and bromocresol purple mixed indicator as acid-base indicator, the color change interval of indicator can be expanded.
In any of the above-described scheme, it is preferred that the quality of phenol red and bromocresol purple in the acid-base indicator indicator
Percentage 4:3-2:1, preferably 2:1.The mass percent of phenol red and bromocresol purple can also be 3:2 and 4:3.Two kinds of instructions
Expand after agent mixing as acid-base indicator, color change interval, effect is better than individual phenol red or bromocresol purple effect.
In any of the above-described scheme, it is preferred that it is anhydrous that phenol red 0.8g and bromocresol purple 0.4g is simultaneously or successively dissolved in 6ml
Ethyl alcohol, then add 4ml pure water, obtain acid-base indicator solution.
Second aspect, the present invention a kind of preparation method of optochin sensitization test culture medium is correspondingly provided, including according to
The weighing drug of secondary progress, dissolving drug, packing, degerming, the drug include acid-base indicator.
Preferably, after further including the dissolving drug, the pH value for adjusting solution is neutrality（7.2-7.5）.If by pH
Value is adjusted to 7.2 hereinafter, the culture medium meeting yellowing being prepared, is not easy to subsequently identify the variation of color.
In any of the above-described scheme, it is preferred that after adjusting pH value, be filtered degerming, obtain degerming solution.Optochin
High pressure degerming cannot be used, filtration sterilization can only be used, the bacterium in drug, solution is removed, so as to ensure the training being prepared
Base is supported when for subsequent applications, as a result accurately.
In any of the above-described scheme, it is preferred that after filtration sterilization, above-mentioned 50 μ l of degerming solution is taken to add in biochemical hole, are had
Conducive to subsequent stoving process.
In any of the above-described scheme, it is preferred that the degerming solution in biochemical hole is dried, is specially dried at 45 DEG C
Bake four to six hours.It is dried under such parameter, the solution in biochemical hole can effectively be made to dry, while avoided
Degree is dry and drying is insufficient.
In any of the above-described scheme, it is preferred that obtained product dispenses after the degerming solution is dried, during packing
It is sealed, vacuumizes, be conducive to preserve dried product for a long time.
In any of the above-described scheme, it is preferred that the product dispensed is carried out irradiation sterilization.Radiation sterilization is to utilize electromagnetism
The electromagnetic wave that radiation generates kills a kind of effective ways of the microorganism on most of substances.The purpose of radio sterilization is different, adopts
Dose of radiation is also different, and the irradiation dose sterilized completely is 25 ~ 50kGy, and the purpose is to kill in addition to bacillus
All microorganisms.The dose of radiation of disinfection be 1 ~ 10kGy, the purpose is to kill irradiation object in asporulate pathogen and
Reduce microbial contamination.In short, for different microorganisms, need to control different dose of radiation and electron energy.In order to the greatest extent
The possible contamination phenomenon for eliminating kit by prolonged verification experimental verification, selects the irradiation of disinfection in production technology
Dosage（8kGy）, in this way so that when the culture medium is used for subsequent applications, as a result accurately.
In any of the above-described scheme, it is preferred that further include, before weighing drug, test to drug.It is marked according to CLSI
Standard is checked whether drug meets the requirements, storing mode, shelf-life and quality including examining drug, so as to ensure to train
Support the sensitivity that base is used to differentiate strain.
In any of the above-described scheme, it is preferred that the acid-base indicator is phenol red and bromocresol purple mixed indicator.It adopts
By the use of phenol red and bromocresol purple mixed indicator as acid-base indicator, the color change interval of indicator is expanded.
In any of the above-described scheme, it is preferred that the quality of phenol red and bromocresol purple in the acid-base indicator indicator
Percentage is 2:1.Expand after two kinds of indicator mixing as acid-base indicator, color change interval, effect is than individual phenol red or bromine
Cresol-purple effect is good.
In any of the above-described scheme, it is preferred that the solution concentration of the acid-base indicator is 0.12g/ml, more preferred
It is that phenol red 0.8g and bromocresol purple 0.4g are dissolved in 6ml absolute ethyl alcohols, then add 4ml pure water, obtain acid-base indicator solution.
The third aspect, the invention further relates to the application of the optochin sensitization test culture medium, by the culture medium
For the identification of streptococcus pneumonia.
Preferably, the culture medium is placed in biochemical identification plate, combines identification strain with other experiments.By the culture
Base is placed in biochemical identification plate, its result can be combined with other experimental results, compared, being capable of a variety of bacterium of accurate judgement
Kind.
In any of the above-described scheme, it is preferred that the application includes the following steps：
（1）Bacteria suspension is configured：It is 0.5 maxwell unit with the streptococcus configuration concentration that the previous day is inoculated with（MCF）Bacteria suspension；
（2）Flora is accessed into the culture medium；
（3）The culture medium of flora aerobic culture 18-24 hours at 35 DEG C will be connected to；
（4）Flora is judged according to culture medium color.
In any of the above-described scheme, it is preferred that the step（2）In, add in 150 microlitres of bacteria suspensions in each biochemistry hole.
The culture medium obtained after the drying adds in 150 microlitres of bacteria suspensions.
In any of the above-described scheme, it is preferred that the step（4）In, by the color result of the culture medium and other knots
Signal in the combination and computer software of fruit is compared, and judges strain.By optochin result of the test and other experiments
As a result it is combined, it can be with precise Identification bacteria to be tested.
During Bacteria Culture is carried out, if bacterium, to optochin sensitivity, the growth of bacterium can be suppressed
Or do not grow, breed completely, then decomposition to glucose does not exist even using less, then culture medium still maintains original
Red；If during Bacteria Culture, bacterium is to optochin drug resistance, then bacterium meeting amount reproduction, a large amount of to decompose, utilize
Glucose, along with the generation of lactic acid so that the pH value of culture medium drops to 6.0 hereinafter, so that culture medium becomes from red
For yellow, therefore, when culture medium shows yellow, it is judged as bacterium to optochin drug resistance；When culture medium is displayed in red, judge
Bacterium is to optochin sensitivity.
Optochin sensitization test culture medium using the present invention can judge tested bacterium colony by culture medium color change
Whether to optochin sensitivity, more than 90% rate of accuracy reached to.This method, which can be saved, measures the work of scraps of paper antibacterial circle diameter, according to
Color change directly reads result；Optochin sensitization test can be added in bacterial biochemical assay plate, increase biochemical identification plate
Identify the ability of bacterium, reading optochin reaction result to semi-automatic, robot provides conveniently.
The optochin sensitization test culture medium of the present invention can singly be not limited by the optochin sensitivity examination of the present invention
The preparation method for testing culture medium obtains.
Specific embodiment
In order to which the technology contents of the present invention are more clearly understood, the present invention is done further with reference to specific embodiment
Explanation, explain.
Used drug, reagent are bought from Chinese medicines group in following embodiment, and distilled water is using two-pass reverse osmosis
Pure water equipment is self-produced.
MH meat soup biochemical reagents；Glucose analysis is pure；Blood meal biochemical reagents；Optochin：Bulk pharmaceutical chemicals；It is phenol red：Instruction
Agent；Bromocresol purple：Indicator.
Embodiment 1
The preparation of acid-base indicator solution：0.8g phenol reds and 0.4g bromocresol purples are removed, is dissolved in 6ml absolute ethyl alcohols, so
4ml distilled water is added in afterwards, is uniformly mixed, is obtained acid-base indicator solution.
Optochin sensitization test culture medium in the present embodiment is to prepare in accordance with the following methods：
Step 1：MH meat soups 1.5000g, glucose 4.3680g is taken to be dissolved in 30mL distilled water, treats both the above ingredient
After being completely dissolved, basal liquid is obtained；
Step 2：10mL basal liquids are taken, add in 0.0240g optochins, 0.0960 blood meal and 0.1ml phenol reds and bromine first
The mixture solution of phenol violet；
Step 3：The pH of the mixed solution obtained in regulating step two is 7.5 or so；
Step 4：The solution that pH is 7.5 is filtered degerming；
Step 5：Solution after filtration sterilization is added in biochemical cup, wherein being added in each biochemical cup molten after degerming
50 microlitres of liquid；
Step 6：All biochemical cups are dried at 45 DEG C, or so about five hours of time obtain dry production
Object；
Step 7：Dried product is vacuum-packed, irradiation sterilization is to get sensitive to the optochin of the present invention
Test medium, wherein, irradiation sterilization irradiation metering used is 8kGy.
Optochin culture medium in the present embodiment can be used for the identification of streptococcus pneumonia, include the following steps：
（1）Bacteria suspension is configured：With the bacteria suspension that the streptococcus configuration concentration that the previous day is inoculated with is 0.5 maxwell unit；
（2）Flora is accessed into the culture medium：150 microlitres of bacteria suspensions is taken to add in each biochemical hole；
（3）The culture medium for being connected to flora is cultivated 18 hours at 35 DEG C；
（4）Flora is judged according to culture medium color：If culture medium color is yellow, illustrate the flora in the bacteria suspension added in
To optochin drug resistance；If culture medium is red, illustrate the flora added in optochin sensitivity, you can be judged as the bacterium added in
Group contains streptococcus pneumonia.
In the present embodiment, rate of accuracy reached to 97.3%.
Embodiment 2.1
As different from Example 1, the optochin sensitization test culture medium in the present embodiment is to make in accordance with the following methods
It is standby：
Step 1：MH meat soups 3.1257g, glucose 5.6700g is taken to be dissolved in 70mL distilled water, treats both the above ingredient
After being completely dissolved, basal liquid is obtained；
Step 2：10mL basal liquids are taken, add in 0.0225g optochins, 0.1575g blood meals and 0.2ml phenol reds and bromine
The mixture of cresol-purple；
Step 3：The pH of the mixed solution obtained in regulating step two is 7.5 or so；
Step 4：The solution that pH is 7.5 is filtered degerming；
Step 5：Solution after filtration sterilization is added in biochemical cup, wherein being added in each biochemical cup molten after degerming
50 microlitres of liquid；
Step 6：All biochemical cups are dried at 45 DEG C, or so about five hours of time obtain dry production
Object；
Step 7：Dried product is vacuum-packed, irradiation sterilization is to get sensitive to the optochin of the present invention
Test medium.
In the present embodiment, rate of accuracy reached to 98.4%.
Embodiment 2.2
As different from Example 1, the optochin sensitization test culture medium in the present embodiment is to make in accordance with the following methods
It is standby：
Step 1：MH meat soups 2.3792g, glucose 5.3720g is taken to be dissolved in 60mL distilled water, treats both the above ingredient
After being completely dissolved, basal liquid is obtained；
Step 2：10mL basal liquids are taken, add in 0.0280g optochins, 0.2000g blood meals and 0.17ml phenol reds and bromine
The mixture solution of cresol-purple；
Step 3：The pH of the mixed solution obtained in regulating step two is 7.5 or so；
Step 4：The solution that pH is 7.5 is filtered degerming；
Step 5：Solution after filtration sterilization is added in biochemical cup, wherein being added in each biochemical cup molten after degerming
50 microlitres of liquid；
Step 6：All biochemical cups are dried at 45 DEG C, or so about five hours of time obtain dry production
Object；
Step 7：Dried product is vacuum-packed, irradiation sterilization is to get sensitive to the optochin of the present invention
Test medium.
In the present embodiment, rate of accuracy reached to 95.1%.
Embodiment 2.3
As different from Example 1, the optochin sensitization test culture medium in the present embodiment is to make in accordance with the following methods
It is standby：
Step 1：MH meat soups 2.4693g, glucose 5.8876g is taken to be dissolved in 50mL distilled water, treats both the above ingredient
After being completely dissolved, basal liquid is obtained；
Step 2：10mL basal liquids are taken, add in 0.0283g optochins, 0.1767g blood meals and 0.15ml phenol reds and bromine
The mixture solution of cresol-purple；
Step 3：The pH of the mixed solution obtained in regulating step two is 7.5 or so；
Step 4：The solution that pH is 7.5 is filtered degerming；
Step 5：Solution after filtration sterilization is added in biochemical cup, wherein being added in each biochemical cup molten after degerming
50 microlitres of liquid；
Step 6：All biochemical cups are dried at 45 DEG C, or so about five hours of time obtain dry production
Object；
Step 7：Dried product is vacuum-packed, irradiation sterilization is to get sensitive to the optochin of the present invention
Test medium.
In the present embodiment, rate of accuracy reached to 99%.
Embodiment 2.4
As different from Example 1, the optochin sensitization test culture medium in the present embodiment is to make in accordance with the following methods
It is standby：
Step 1：MH meat soups 2.0111g, glucose 4.7691g is taken to be dissolved in 40mL distilled water, treats both the above ingredient
After being completely dissolved, basal liquid is obtained；
Step 2：10mL basal liquids are taken, add in 0.0217g optochins, 0.1827g blood meals and 0.13ml phenol reds and bromine
The mixture solution of cresol-purple；
Step 3：The pH of the mixed solution obtained in regulating step two is 7.5 or so；
Step 4：The solution that pH is 7.5 is filtered degerming；
Step 5：Solution after filtration sterilization is added in biochemical cup, wherein being added in each biochemical cup molten after degerming
50 microlitres of liquid；
Step 6：All biochemical cups are dried at 45 DEG C, or so about five hours of time obtain dry production
Object；
Step 7：Dried product is vacuum-packed, irradiation sterilization is to get sensitive to the optochin of the present invention
Test medium.
In the present embodiment, rate of accuracy reached to 96.3%.
Embodiment 2.5
As different from Example 1, the optochin sensitization test culture medium in the present embodiment is to make in accordance with the following methods
It is standby：
Step 1：MH meat soups 2.2176g, glucose 4.5759g is taken to be dissolved in 70mL distilled water, treats both the above ingredient
After being completely dissolved, basal liquid is obtained；
Step 2：Take 10mL basal liquids, add in 0.0231g optochins, 0.1610g blood meals and 0.185ml phenol reds and
The mixture solution of bromocresol purple；
Step 3：The pH of the mixed solution obtained in regulating step two is 7.5 or so；
Step 4：The solution that pH is 7.5 is filtered degerming；
Step 5：Solution after filtration sterilization is added in biochemical cup, wherein being added in each biochemical cup molten after degerming
50 microlitres of liquid；
Step 6：All biochemical cups are dried at 45 DEG C, or so about five hours of time obtain dry production
Object；
Step 7：Dried product is vacuum-packed, irradiation sterilization is to get sensitive to the optochin of the present invention
Test medium.
In the present embodiment, rate of accuracy reached to 93.8%.
Embodiment 2.6
As different from Example 1, the optochin sensitization test culture medium in the present embodiment is to make in accordance with the following methods
It is standby：
Step 1：MH meat soups 2.1511g, glucose 4.6900g is taken to be dissolved in 55mL distilled water, treats both the above ingredient
After being completely dissolved, basal liquid is obtained；
Step 2：10mL basal liquids are taken, add in 0.0259g optochins, 0.1141g blood meals and 0.16ml phenol reds and bromine
The mixture solution of cresol-purple；
Step 3：The pH of the mixed solution obtained in regulating step two is 7.5 or so；
Step 4：The solution that pH is 7.5 is filtered degerming；
Step 5：Solution after filtration sterilization is added in biochemical cup, wherein being added in each biochemical cup molten after degerming
50 microlitres of liquid；
Step 6：All biochemical cups are dried at 45 DEG C, or so about five hours of time obtain dry production
Object；
Step 7：Dried product is vacuum-packed, irradiation sterilization is to get sensitive to the optochin of the present invention
Test medium.
In the present embodiment, rate of accuracy reached to 92.7%.
Embodiment 2.7
As different from Example 1, the optochin sensitization test culture medium in the present embodiment is to make in accordance with the following methods
It is standby：
Step 1：MH meat soups 2.0433g, glucose 4.7446g is taken to be dissolved in 65mL distilled water, treats both the above ingredient
After being completely dissolved, basal liquid is obtained；
Step 2：10mL basal liquids are taken, add in 0.0182g optochins, 0.1722g blood meals and 0.18ml phenol reds and bromine
The mixture solution of cresol-purple；
Step 3：The pH of the mixed solution obtained in regulating step two is 7.5 or so；
Step 4：The solution that pH is 7.5 is filtered degerming；
Step 5：Solution after filtration sterilization is added in biochemical cup, wherein being added in each biochemical cup molten after degerming
50 microlitres of liquid；
Step 6：All biochemical cups are dried at 45 DEG C, or so about five hours of time obtain dry production
Object；
Step 7：Dried product is vacuum-packed, irradiation sterilization is to get sensitive to the optochin of the present invention
Test medium.
In the present embodiment, rate of accuracy reached to 91.7%.
Embodiment 2.8
As different from Example 1, the optochin sensitization test culture medium in the present embodiment is to make in accordance with the following methods
It is standby：
Step 1：MH meat soups 1.7969g, glucose 5.0386g is taken to be dissolved in 45mL distilled water, treats both the above ingredient
After being completely dissolved, basal liquid is obtained；
Step 2：10mL basal liquids are taken, add in 0.0189g optochins, 0.1225g blood meals and 0.19ml phenol reds and bromine
The mixture solution of cresol-purple；
Step 3：The pH of the mixed solution obtained in regulating step two is 7.5 or so；
Step 4：The solution that pH is 7.5 is filtered degerming；
Step 5：Solution after filtration sterilization is added in biochemical cup, wherein being added in each biochemical cup molten after degerming
50 microlitres of liquid；
Step 6：All biochemical cups are dried at 45 DEG C, or so about five hours of time obtain dry production
Object；
Step 7：Dried product is vacuum-packed, irradiation sterilization is to get sensitive to the optochin of the present invention
Test medium.
In the present embodiment, rate of accuracy reached to 94.9%.
Embodiment 2.9
As different from Example 1, the optochin sensitization test culture medium in the present embodiment is to make in accordance with the following methods
It is standby：
Step 1：MH meat soups 1.8662g, glucose 4.9350g is taken to be dissolved in 35mL distilled water, treats both the above ingredient
After being completely dissolved, basal liquid is obtained；
Step 2：10mL basal liquids are taken, add in 0.0203g optochins, 0.1547g blood meals and 0.20ml phenol reds and bromine
The mixture solution of cresol-purple；
Step 3：The pH of the mixed solution obtained in regulating step two is 7.5 or so；
Step 4：The solution that pH is 7.5 is filtered degerming；
Step 5：Solution after filtration sterilization is added in biochemical cup, wherein being added in each biochemical cup molten after degerming
50 microlitres of liquid；
Step 6：All biochemical cups are dried at 45 DEG C, or so about five hours of time obtain dry production
Object；
Step 7：Dried product is vacuum-packed, irradiation sterilization is to get sensitive to the optochin of the present invention
Test medium.
In the present embodiment, rate of accuracy reached to 96.1%.
Embodiment 2.10
As different from Example 1, the acid-base indicator solution is prepared as：Take phenol red 1.6g and bromocresol purple
0.8g is dissolved in 10ml absolute ethyl alcohols, is then added in 6.77ml distilled water, is obtained acid-base indicator solution.
And when preparing the optochin sensitization test culture medium, the optochin sensitization test culture medium in the present embodiment
It is to prepare in accordance with the following methods：
Step 1：MH meat soups 1.8662g, glucose 4.9119g is taken to be dissolved in 35mL distilled water, treats both the above ingredient
After being completely dissolved, basal liquid is obtained；
Step 2：10mL basal liquids are taken, add in 0.0203g optochins, 0.1547g blood meals and 0.20ml phenol reds and bromine
The mixture solution of cresol-purple；
Step 3：The pH of the mixed solution obtained in regulating step two is 7.5 or so；
Step 4：The solution that pH is 7.5 is filtered degerming；
Step 5：Solution after filtration sterilization is added in biochemical cup, wherein being added in each biochemical cup molten after degerming
50 microlitres of liquid；
Step 6：All biochemical cups are dried at 45 DEG C, or so about five hours of time obtain dry production
Object；
Step 7：Dried product is vacuum-packed, irradiation sterilization is to get sensitive to the optochin of the present invention
Test medium.
In the present embodiment, rate of accuracy reached to 95.9%.
Embodiment 2.11
As different from Example 1, the acid-base indicator solution is prepared as：Take phenol red 0.6g and bromocresol purple
0.3g is dissolved in 6ml absolute ethyl alcohols, is then added in 4.5ml distilled water, is obtained acid-base indicator solution.
And when preparing the optochin sensitization test culture medium, the optochin sensitization test culture medium in the present embodiment
It is to prepare in accordance with the following methods：
Step 1：MH meat soups 1.5036g, glucose 4.3680g is taken to be dissolved in 30mL distilled water, treats both the above ingredient
After being completely dissolved, basal liquid is obtained；
Step 2：Take 10mL basal liquids, add in 0.0240g optochins, 0.00960g blood meals and 0.10ml phenol reds and
The mixture solution of bromocresol purple；
Step 3：The pH of the mixed solution obtained in regulating step two is 7.5 or so；
Step 4：The solution that pH is 7.5 is filtered degerming；
Step 5：Solution after filtration sterilization is added in biochemical cup, wherein being added in each biochemical cup molten after degerming
50 microlitres of liquid；
Step 6：All biochemical cups are dried at 45 DEG C, or so about five hours of time obtain dry production
Object；
Step 7：Dried product is vacuum-packed, irradiation sterilization is to get sensitive to the optochin of the present invention
Test medium.
In the present embodiment, rate of accuracy reached to 97.9%.
Embodiment 3.1
It is unlike the embodiments above, in the present embodiment, in the method for preparing optochin sensitization test culture medium,
Step 3 is adjusted in the pH for adjusting mixed solution to 7.7 or so.In the present embodiment, rate of accuracy reached to 91%.
Embodiment 3.2
It is unlike the embodiments above, in the present embodiment, in the method for preparing optochin sensitization test culture medium,
Step 3 is adjusted in the pH for adjusting mixed solution to 7.6 or so.In the present embodiment, rate of accuracy reached to 90.8%.
Embodiment 3.3
It is unlike the embodiments above, in the present embodiment, in the method for preparing optochin sensitization test culture medium,
Step 3 is adjusted in the pH for adjusting mixed solution to 7.4 or so.In the present embodiment, rate of accuracy reached to 95%.
Embodiment 3.4
It is unlike the embodiments above, in the present embodiment, in the method for preparing optochin sensitization test culture medium,
Step 3 is adjusted in the pH for adjusting mixed solution to 7.3 or so.In the present embodiment, rate of accuracy reached to 92.1%.
Embodiment 4
It is unlike the embodiments above, in the present embodiment, in the method for preparing optochin sensitization test culture medium,
It before drug is weighed, tests to drug, whether drug used in examination meets CLSI standards, and the project of inspection includes drug
Whether condition of storage meets regulation, and whether drug is within the shelf-life, and whether the properties such as character of drug change, the mark of drug
Know content etc., it is only standard compliant just to use, in principle, as possible using the drug of the same nature of identical producer production.
Embodiment 5
It is unlike the embodiments above, in the present embodiment, optochin sensitization test culture medium is being used for strain
During identification, the culture medium is added in biochemical identification hole, by the color result of the culture medium and the group of other 23 kinds of results
Conjunction is compared with the signal in computer software, judges strain.
Embodiment 6.1
It is unlike the embodiments above, in the present embodiment, the quality hundred of phenol red and bromocresol purple in the indicator
Divide than being 3:2.In the present embodiment, rate of accuracy reached to 92%.
Embodiment 6.2
It is unlike the embodiments above, in the present embodiment, the quality hundred of phenol red and bromocresol purple in the indicator
Divide than being 4:3.In the present embodiment, rate of accuracy reached to 90%.
Embodiment 7.1
Unlike the embodiments above, in the present embodiment, the optochin culture medium is used for the mirror of streptococcus pneumonia
Periodically, the culture medium for being connected to flora is cultivated 24 hours at 35 DEG C.
Embodiment 7.2
Unlike the embodiments above, in the present embodiment, the optochin sensitization test culture medium is used for pneumonia chain
During the identification of coccus, the culture medium for being connected to flora is cultivated 20 hours at 35 DEG C.
Embodiment 7.3
It is unlike the embodiments above to be, in the present embodiment, it is connected to the culture medium C O of flora2Under the concentration of 5-10%, 35
It is cultivated 20 hours at DEG C.
Comparative example
By streptococcus pneumonia sensitization test a kind of disclosed in the Chinese invention patent application of Publication No. CN101698876A
The scraps of paper are compared with the optochin sensitization test culture medium in the present invention, and Publication No. very low in pneumonia streptococcus bacteria concentration
A kind of antibacterial circle diameter of sensitization test (STT) paper for streptococcus pneumoniae is small disclosed in the Chinese invention patent application of CN101698876A
When 10 mm, each formula of the optochin sensitization test culture medium in the present invention remains to the variation significantly to distinguish between colors.
It is worth noting that, above example is only the preferred embodiment of the invention content of the present invention, for explaining,
Illustrate invention content, it is impossible to for limiting the content invented, and, to those skilled in the art, do not departing from this hair
Under the premise of bright total design, several changes for being not required to creative work can also be made, and this change should also belong to this hair
Bright protection domain.
Claims (19)
1. a kind of optochin sensitization test culture medium, including MH meat soups, glucose, distilled water, optochin, blood meal and soda acid
Indicator, the acid-base indicator are phenol red and bromocresol purple mixed indicator.
2. culture medium as described in claim 1, it is characterised in that：The optochin sensitization test culture medium is by quality percentage
The following component of number content is made：
MH broth bouillons 25-35%；
Glucose 63-73%；
Optochin 0.25-0.4%；
Blood meal 1.6-2.5%；
Acid-base indicator 0.14-0.67%.
3. culture medium as claimed in claim 2, it is characterised in that：The pH of the culture medium is 7.2-7.5.
4. culture medium as claimed in claim 3, it is characterised in that：The quality hundred of phenol red and bromocresol purple in the indicator
Divide than being 4:3-2:1.
5. the preparation method of the optochin sensitization test culture medium as described in any one of claim 1-4, including successively into
Capable weighing drug, dissolving drug, packing, degerming.
6. preparation method as claimed in claim 5, it is characterised in that：After further including the dissolving drug, the pH of solution is adjusted
It is worth for 7.3-7.7.
7. preparation method as claimed in claim 6, it is characterised in that：After adjusting pH value, degerming is filtered, it is molten to obtain degerming
Liquid.
8. preparation method as claimed in claim 7, it is characterised in that：After filtration sterilization, above-mentioned 50 μ l of degerming solution is taken to add in
In biochemical hole.
9. preparation method as claimed in claim 8, it is characterised in that：Degerming solution in biochemical hole is dried, specifically
To toast four to six hours at 45 DEG C.
10. preparation method as claimed in claim 9, it is characterised in that：After the degerming solution is dried obtained product into
Row packing, when packing, are sealed, vacuumize.
11. the preparation method as described in claim 5 or 10, it is characterised in that：The product dispensed is subjected to irradiation sterilization.
12. preparation method as claimed in claim 11, it is characterised in that：It further includes, before weighing drug, drug is examined
It tests.
13. preparation method as claimed in claim 12, it is characterised in that：The solution concentration of the acid-base indicator is 0.12g/
ml。
14. preparation method as claimed in claim 13, it is characterised in that：Phenol red 0.8g and bromocresol purple 0.4g are dissolved in 6ml
Absolute ethyl alcohol, then add 4ml pure water, obtain acid-base indicator solution.
15. the application of the optochin sensitization test culture medium as described in any one of claim 1-4, it is characterised in that：It will
The culture medium is used for the identification of the streptococcus pneumonia of non-diagnostic purpose.
16. application as claimed in claim 15, it is characterised in that：The culture medium is placed in biochemical identification plate, with other objects
Matter joint identification strain.
17. the application as described in claim 15 or 16, it is characterised in that：Include the following steps：
（1）Bacteria suspension is configured：With the bacteria suspension that the streptococcus configuration concentration that the previous day is inoculated with is 0.5 maxwell unit；
（2）Flora is accessed into the culture medium；
（3）The culture medium for being connected to flora is cultivated 18-24 hours at 35 DEG C；
（4）Flora is judged according to culture medium color.
18. application as claimed in claim 17, it is characterised in that：The step（2）In, it is micro- to add in 150 in each biochemistry hole
Rise bacteria suspension.
19. application as claimed in claim 18, it is characterised in that：The step（4）In, by the color result of the culture medium
It is compared with the signal in the combination and computer software of other results, judges strain.
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Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101698876A (en) * | 2007-12-18 | 2010-04-28 | 杨宇 | Sensitization test (STT) paper for streptococcus pneumoniae |
-
2015
- 2015-07-06 CN CN201510389463.6A patent/CN105087751B/en active Active
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101698876A (en) * | 2007-12-18 | 2010-04-28 | 杨宇 | Sensitization test (STT) paper for streptococcus pneumoniae |
Non-Patent Citations (4)
| Title |
|---|
| 241株肺炎链球菌血清分型及耐药性研究;李马超等;《中国疫苗和免疫》;20130831;第19卷(第4期);336-340页 * |
| 微量法在致病性大肠杆菌药敏测定中的应用研究;陈翠珍等;《微量法在致病性大肠杆菌药敏测定中的应用研究》;19871231;4-5页 * |
| 肺炎链球菌分布及耐药分析;张艳等;《昆明医科大学学报》;20141231;第35卷(第7期);135-137页 * |
| 肺炎链球菌的感染分布与耐药性分析;沈丽珍等;《中国卫生检验杂志》;20130930;第23卷(第12期);2691-2692页 * |
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