CN106282301A - A kind of mycoplasma pneumoniae fast culture, qualification and the test kit of susceptibility - Google Patents
A kind of mycoplasma pneumoniae fast culture, qualification and the test kit of susceptibility Download PDFInfo
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- CN106282301A CN106282301A CN201610671966.7A CN201610671966A CN106282301A CN 106282301 A CN106282301 A CN 106282301A CN 201610671966 A CN201610671966 A CN 201610671966A CN 106282301 A CN106282301 A CN 106282301A
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- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
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Abstract
The present invention discloses a kind of mycoplasma pneumoniae fast culture, identifies the test kit with susceptibility, belong to susceptibility detection technique field, including culture fluid, diluent, mineral oil and detection plate, containing beef heart infusion 60～80%, NaCl0.6～0.8%, tryptone 0.3～2.0%, Na in described culture fluid2HPO40.05～0.4%, magnesium sulfate 0.03～0.08%, ammonium sulfate 0.1～0.2%, standard calf serum 10.0～30.0%, yolk liquid 5.0～15.0%, glucose 0.5～1.5%, vancomycin 0.001%, amphotericin B 0.001%, chloromycetin 0.01～0.08%, phenol red 0.004～0.02%, methylene blue 0.001～0.005%；It is provided with mycoplasma pneumoniae on described detection plate and differentiates hole and antibiotic drug sensitive detection hole.The test kit of the present invention, sensitivity, quickly, can go out result, and high specificity at 6～10 hours.
Description
Technical field
The present invention relates to susceptibility detection technique field, particularly to a kind of mycoplasma pneumoniae fast culture, qualification and susceptibility
Test kit.
Background technology
Mycoplasma pneumoniae (Mycoplasma pneumoniae Mp) is a kind of pathogenic microorganism that the mankind are newly discovered.It
Be a kind of by patient's mouth and nose secretions through air droplet transmission, easily cause sporadic or regional popular, its cardinal symptom is for exhaling
Inhale road and infect the pathogen of companion's pneumonia i.e. mycoplasma pneumoniae pneumonia (Mycoplasma pneumoniae pneumonia MPP),
The mankind are had the most pathogenic.Mp infects the four seasons and all can fall ill, but autumn and winter is in the majority, and infant and teenager are easy infection people
Group, sickness rate is the 20-30% of infantile pneumonia class, and populous and epidemic season is up to 50%, and popular cycle is 3-7, closely
Popular cycle is had to shorten over Nian, the trend that sickness rate raises.It is long that Mp infects incubation period, and PD is slow, and Symptoms is failed to understand
Aobvious, infect caused respiratory symptom to common bacteria, virus similar, clinic is difficult to diagnosis or Differential Diagnosis and medicine is big
Differing, the clinical treatment cycle is long, and often delay treatment causes serious complication, therefore the quick of Mp infection detects, and accurately examines
Disconnected, timely diagnosis and treatment have important clinical meaning.
Culture method is that microbial pathogens detects " goldstandard ", and the technique of MP fluid medium, formula have passed through tens
The clinical verification in year is the most ripe, but existing susceptibility incubation time is long, generally 24～72 hours, causes the MP inspection of routine
Survey can not routine be carried out, and can not meet the problem that clinical MP infects suspected patient rapid screening in early days.
Summary of the invention
The present invention provides a kind of mycoplasma pneumoniae fast culture, identifies the test kit with susceptibility, solves existing pneumonia and props up
The problem of substance susceptibility detection incubation time length.
For solving above-mentioned technical problem, the technical scheme is that
A kind of mycoplasma pneumoniae fast culture, identify and the test kit of susceptibility, including culture fluid, diluent, mineral oil and
Detection plate, in described culture fluid containing beef heart infusion 60～80%, NaCl0.6～0.8%, tryptone 0.3～2.0%,
Na2HPO40.05～0.4%, magnesium sulfate 0.03～0.08%, ammonium sulfate 0.1～0.2%, standard calf serum 10.0～
30.0%, yolk liquid 5.0～15.0%, glucose 0.5～1.5%, vancomycin 0.001%, amphotericin B 0.001%, chlorine
Mycin 0.01～0.08%, phenol red 0.004～0.02%, methylene blue 0.001～0.005%；It is provided with lung on described detection plate
Scorching mycoplasma differentiates hole and antibiotic drug sensitive detection hole.
Wherein it is preferred to, described diluent be concentration be the NaCl aqueous solution of 0.5～1.0%.
Wherein it is preferred to, described mycoplasma pneumoniae differentiates to be loaded with respectively in hole dry arginine, chlorinated triphenyl base four
Nitrogen azoles indigo plant, methylene blue and Beta-lactam medicine.
Wherein it is preferred to, described antibiotic drug sensitive detection group includes organizing susceptibility detector unit, often group susceptibility detector unit more
Including two susceptibility micropores.
Wherein it is preferred to, it is loaded with the same medicine of variable concentrations in two susceptibility micropores of same group of susceptibility detector unit
Thing.
Wherein it is preferred to, the medicine that described susceptibility micropore contains is erythromycin, acetylspiramycin, josamycin, strongly
Mycin, Roxithromycin, ofloxacin, azithromycin, clarithromycin or ciprofloxacin.
Wherein it is preferred to, described mineral oil is saxol.
The present invention is to combine with biochemical reaction cultivating, and mycoplasma pneumoniae has the ability decomposing Fructus Vitis viniferae product acid, makes training
The pH of nutrient solution reduces, according to the color change judged result of acid-base indicator instruction.If detection in antibiotic drug sensitive detection hole
Mycoplasma pneumoniae is to coated antibiotic sensitive, and its enzymatic activity is suppressed, and color will not be caused to change.
Beneficial effects of the present invention:
1. the test kit of the present invention, sensitivity, quickly, can go out result at 6～10 hours, and similar mycoplasma pneumoniae reagent
Box needs just to go out result in 24～48 hours.
2. the test kit high specificity of the present invention, is not suitable for the growth of other mycoplasma and other antibacterial, passes through the present invention
Middle culture fluid, is simultaneously introduced vancomycin, amphotericin B and the chloromycetin not affecting mycoplasma growth, is suppressed by other miscellaneous bacteria
Or kill.
Test kit the most of the present invention can carry out drug sensitive test to Multiple Classes of Antibiotics simultaneously, contains wide, clinical strong adaptability.
Test kit the most of the present invention is cultivated and is synchronized to carry out with susceptibility, convenient and swift, more effective treats Clinical detection and treatment.
Detailed description of the invention
Below the technical scheme in the embodiment of the present invention is clearly and completely described, it is clear that described embodiment
It is only a part of embodiment of the present invention rather than whole embodiments.Based on the embodiment in the present invention, the common skill in this area
The every other embodiment that art personnel are obtained under not making creative work premise, broadly falls into the model of present invention protection
Enclose.
Embodiment 1
The present embodiment puies forward a kind of mycoplasma pneumoniae fast culture, identifies and the test kit of susceptibility, including culture fluid, 0.8%
NaCl aqueous solution, saxol and detection plate, containing beef heart infusion 70%, NaCl0.6%, Trypsin in described culture fluid
Peptone 1.0%, Na2HPO40.2%, magnesium sulfate 0.05%, ammonium sulfate 0.14%, standard calf serum 20%, yolk liquid 7%, Portugal
Grape sugar 0.92%, vancomycin 0.001%, amphotericin B 0.001%, chloromycetin 0.05%, phenol red 0.008%, methylene blue
0.003%；It is provided with mycoplasma pneumoniae on described detection plate and differentiates hole and antibiotic drug sensitive detection hole.
Wherein, described mycoplasma pneumoniae differentiate to be loaded with respectively in hole dry arginine, triphenyltetrazolium chloride blue,
Methylene blue and Beta-lactam medicine.
Wherein it is preferred to, described antibiotic drug sensitive detection group includes nine groups of susceptibility detector units, often group susceptibility detector unit
Including two susceptibility micropores.
Wherein, the same medicine of variable concentrations it is loaded with in two susceptibility micropores of same group of susceptibility detector unit.Described
Susceptibility micropore contain medicine be erythromycin, acetylspiramycin, josamycin, doxycycline, Roxithromycin, ofloxacin,
Azithromycin, clarithromycin and ciprofloxacin.
Embodiment 2
The present embodiment puies forward a kind of mycoplasma pneumoniae fast culture, identifies and the test kit of susceptibility, including culture fluid, 0.9%
NaCl aqueous solution, saxol and detection plate, containing beef heart infusion 60%, NaCl0.8%, Trypsin in described culture fluid
Peptone 2.0%, Na2HPO40.05%, magnesium sulfate 0.08%, ammonium sulfate 0.2%, standard calf serum 30.0%, yolk liquid
5.308%, glucose 1.5%, vancomycin 0.001%, amphotericin B 0.001%, chloromycetin 0.08%, phenol red 0.02%,
Methylene blue 0.005%；It is provided with mycoplasma pneumoniae on described detection plate and differentiates hole and antibiotic drug sensitive detection hole.
Wherein, described mycoplasma pneumoniae differentiate to be loaded with respectively in hole dry arginine, triphenyltetrazolium chloride blue,
Methylene blue and Beta-lactam medicine.
Wherein it is preferred to, described antibiotic drug sensitive detection group includes seven groups of susceptibility detector units, often group susceptibility detector unit
Including two susceptibility micropores.
Wherein, the same medicine of variable concentrations it is loaded with in two susceptibility micropores of same group of susceptibility detector unit.Described
The medicine that susceptibility micropore contains is erythromycin, josamycin, doxycycline, ofloxacin, azithromycin, clarithromycin and ring third
Sha Xing.
Embodiment 3
The present embodiment puies forward a kind of mycoplasma pneumoniae fast culture, identifies and the test kit of susceptibility, including culture fluid, 0.5%
NaCl aqueous solution, saxol and detection plate, containing beef heart infusion 72%, NaCl0.7%, Trypsin in described culture fluid
Peptone 0.37%, Na2HPO470.4%, magnesium sulfate 0.037%, ammonium sulfate 0.1～0.2%, standard calf serum 10.949%, ovum
Yellow liquor 15.0%, glucose 0.5%, vancomycin 0.001%, amphotericin B 0.001%, 0.04% chloromycetin, phenol red
0.004%, methylene blue 0.005%；It is provided with mycoplasma pneumoniae on described detection plate and differentiates hole and antibiotic drug sensitive detection hole.
Wherein, described mycoplasma pneumoniae differentiate to be loaded with respectively in hole dry arginine, triphenyltetrazolium chloride blue,
Methylene blue and Beta-lactam medicine.
Wherein it is preferred to, described antibiotic drug sensitive detection group includes eight groups of susceptibility detector units, often group susceptibility detector unit
Including two susceptibility micropores.
Wherein, the same medicine of variable concentrations it is loaded with in two susceptibility micropores of same group of susceptibility detector unit.Described
Susceptibility micropore contain medicine be erythromycin, acetylspiramycin, josamycin, doxycycline, Roxithromycin, ofloxacin,
Clarithromycin and ciprofloxacin.
Embodiment 4
The present embodiment puies forward a kind of mycoplasma pneumoniae fast culture, identifies and the test kit of susceptibility, including culture fluid, 1.0%
NaCl aqueous solution, saxol and detection plate, containing beef heart infusion 80%, NaCl0.7%, Trypsin in described culture fluid
Peptone 1.2%, Na2HPO40.3%, magnesium sulfate 0.05%, ammonium sulfate 0.15%, standard calf serum 11.334%, yolk liquid
5%, glucose 1.2%, vancomycin 0.001%, amphotericin B 0.001%, chloromycetin 0.05%, phenol red 0.01%, methylene
Blue 0.004%；It is provided with mycoplasma pneumoniae on described detection plate and differentiates hole and antibiotic drug sensitive detection hole.
Wherein, described mycoplasma pneumoniae differentiate to be loaded with respectively in hole dry arginine, triphenyltetrazolium chloride blue,
Methylene blue and Beta-lactam medicine.
Wherein it is preferred to, described antibiotic drug sensitive detection group includes seven groups of susceptibility detector units, often group susceptibility detector unit
Including two susceptibility micropores.
Wherein, the same medicine of variable concentrations it is loaded with in two susceptibility micropores of same group of susceptibility detector unit.Described
Susceptibility micropore contain medicine be erythromycin, acetylspiramycin, doxycycline, Roxithromycin, ofloxacin, azithromycin and
Ciprofloxacin.
The mycoplasma pneumoniae fast culture of above-described embodiment, identify as follows with the test kit preparation method of susceptibility:
(1) preparation of culture fluid: weigh beef heart infusion, NaCl, tryptone, Na by proportioning2HPO4, magnesium sulfate, sulphuric acid
Ammonium, standard calf serum, yolk liquid, glucose, vancomycin, amphotericin B 0.001%, chloromycetin, phenol red and methylene blue,
Vibration makes dissolving, regulates pH value 7.4 ± 0.2,0.22 μm filtering with microporous membrane, to obtain final product.
(2) preparation of drug sensitive plate is differentiated: with methanol as diluent, prepare arginine, chlorinated triphenyl base four respectively by concentration
Nitrogen azoles indigo plant, methylene blue and Beta-lactam medicine solution, calculated by 100 μ l/ holes, arginine final concentration of 0.05%, chlorination three
Final concentration 0.05%, methylene blue 0.2% and the beta-lactam 100mg/L that phenyl tetrazole is blue.
(3) susceptibility micropore liquid: the judgement medicaments insensitive announced with state-owned Clinical Laboratory Standard committee (NCCLS)
MIC value be low concentration standard (L), tolerance MIC value be high standard (H), with methanol as diluent, 9 kinds of medicines of preparation respectively
18 kinds of concentration；50 μ l/ holes add in specific micropore, are dried, encapsulate, to obtain final product.
Use the detection method of this test kit detection mycoplasma pneumoniae, comprise the following steps:
Sample swab is put into abundant eluting in the culture fluid of culture tube extract；
Again culture fluid is added with the amount of 100 IlL per well (ul/ hole) in the micropore differentiating plate to susceptibility
Mineral oil is added to each micropore with the amount in 1 every hole；
Cultivate 24～72 hours in the environment of test kit is placed on 35 DEG C～37 DEG C, judge knot according to the change of micropore color
Really.
Result judges as follows:
Differentiate district: it is Mp that Arg-TTC-MB-β-Lac shows as yellow-red-Huang green-yellow, is otherwise other mycoplasma；
Susceptibility district: same drug low concentration (L) hole, high concentration (H) Kong Junhong, sentences sensitivity；Low concentration (L) hole turns yellow, height
Concentration (H) Kong Hong, quick in sentencing；Low concentration (L) hole, high concentration (H) Kong Junhuang, sentence drug resistance.
Mentioned reagent box has carried out quality evaluation, and wherein, the total coincidence rate of replica test is more than 96%, and stability is tried
Test this reagent of table to preserve at-20 DEG C and have good stability for 6 months.
The foregoing is only presently preferred embodiments of the present invention, not in order to limit the present invention, all essences in the present invention
Within god and principle, any modification, equivalent substitution and improvement etc. made, should be included within the scope of the present invention.
Claims (7)
1. a mycoplasma pneumoniae fast culture, identify the test kit with susceptibility, it is characterised in that: include culture fluid, diluent,
Mineral oil and detection plate, in described culture fluid containing beef heart infusion 60～80%, NaCl0.6～0.8%, tryptone 0.3～
2.0%, Na2HPO40.05～0.4%, magnesium sulfate 0.03～0.08%, ammonium sulfate 0.1～0.2%, standard calf serum 10.0
～30.0%, yolk liquid 5.0～15.0%, glucose 0.5～1.5%, vancomycin 0.001%, amphotericin B 0.001%,
Chloromycetin 0.01～0.08%, phenol red 0.004～0.02%, methylene blue 0.001～0.005%；It is provided with on described detection plate
Mycoplasma pneumoniae differentiates hole and antibiotic drug sensitive detection hole.
2. seek a kind of mycoplasma pneumoniae fast culture described in 1 according to right, identify the test kit with susceptibility, it is characterised in that:
Described diluent be concentration be the NaCl aqueous solution of 0.5～1.0%.
A kind of mycoplasma pneumoniae fast culture the most according to claim 1, qualification and the test kit of susceptibility, its feature exists
In: described mycoplasma pneumoniae differentiates hole, be loaded with that dry arginine, triphenyltetrazolium chloride be blue respectively, methylene blue and β-
Lactam drugs.
A kind of mycoplasma pneumoniae fast culture the most according to claim 1, qualification and the test kit of susceptibility, its feature exists
In: described antibiotic drug sensitive detection group includes organizing susceptibility detector unit more, and often group susceptibility detector unit includes two susceptibility micropores.
A kind of mycoplasma pneumoniae fast culture the most according to claim 4, qualification and the test kit of susceptibility, its feature exists
In: it is loaded with the same medicine of variable concentrations in two susceptibility micropores of same group of susceptibility detector unit.
6. to mark a kind of mycoplasma pneumoniae fast culture described in 5 according to right, identify the test kit with susceptibility, its feature exists
In: the medicine that described susceptibility micropore contains is erythromycin, acetylspiramycin, josamycin, doxycycline, Roxithromycin, oxygen
Flucloxacillin, azithromycin, clarithromycin or ciprofloxacin.
A kind of mycoplasma pneumoniae fast culture the most according to claim 1, qualification and the test kit of susceptibility, its feature exists
In: described mineral oil is saxol.
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| CN201610671966.7A CN106282301A (en) | 2016-08-15 | 2016-08-15 | A kind of mycoplasma pneumoniae fast culture, qualification and the test kit of susceptibility |
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| CN201610671966.7A CN106282301A (en) | 2016-08-15 | 2016-08-15 | A kind of mycoplasma pneumoniae fast culture, qualification and the test kit of susceptibility |
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101555517A (en) * | 2009-05-15 | 2009-10-14 | 钱叶林 | Mycoplasma pneumoniae rapid identification and cultivation susceptibility test kit and test method |
| CN201545845U (en) * | 2009-07-01 | 2010-08-11 | 郭秋丽 | Mycoplasma and candida joint identification drug susceptibility reagent kit |
| CN102888441A (en) * | 2011-07-20 | 2013-01-23 | 农高惠 | Kit for fast culture, identification and drug sensitivity test of myeoplasmapneumoniae (Mp) and detection method therefor |
-
2016
- 2016-08-15 CN CN201610671966.7A patent/CN106282301A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101555517A (en) * | 2009-05-15 | 2009-10-14 | 钱叶林 | Mycoplasma pneumoniae rapid identification and cultivation susceptibility test kit and test method |
| CN201545845U (en) * | 2009-07-01 | 2010-08-11 | 郭秋丽 | Mycoplasma and candida joint identification drug susceptibility reagent kit |
| CN102888441A (en) * | 2011-07-20 | 2013-01-23 | 农高惠 | Kit for fast culture, identification and drug sensitivity test of myeoplasmapneumoniae (Mp) and detection method therefor |
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Application publication date: 20170104 |