WO2013117967A1 - Compositions et procédés de préparation d'échantillons de crachats, et kit - Google Patents

Compositions et procédés de préparation d'échantillons de crachats, et kit Download PDF

Info

Publication number
WO2013117967A1
WO2013117967A1 PCT/IB2012/050628 IB2012050628W WO2013117967A1 WO 2013117967 A1 WO2013117967 A1 WO 2013117967A1 IB 2012050628 W IB2012050628 W IB 2012050628W WO 2013117967 A1 WO2013117967 A1 WO 2013117967A1
Authority
WO
WIPO (PCT)
Prior art keywords
composition
sputum
sputum sample
sample preparation
prepared
Prior art date
Application number
PCT/IB2012/050628
Other languages
English (en)
Inventor
Maloy GHOSH
Prabhakar Kulkarni
Bala S Manian
Sridhar RAMANATHAN
Original Assignee
Reametrix Inc.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Reametrix Inc. filed Critical Reametrix Inc.
Publication of WO2013117967A1 publication Critical patent/WO2013117967A1/fr

Links

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/30—Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/40—Concentrating samples
    • G01N1/4077—Concentrating samples by other techniques involving separation of suspended solids
    • G01N2001/4088—Concentrating samples by other techniques involving separation of suspended solids filtration

Definitions

  • the invention relates generally to compositions and methods for sputum sample preparation and more specifically to compositions and methods for liquefaction of sputum sample and making the Mycobacteria available for further processing while inactivating other contaminant bacteria.
  • Sputum from humans is used to diagnose tuberculosis and detection of infection in cystic fibrosis patients.
  • Sputum is a highly viscous liquid in its native state, which poses several challenges towards sample processing and treatment.
  • a patient is asked to produce a certain volume of sputum, which is then treated appropriately to enable viewing under a microscope for Mycobacteria.
  • the sputum sample may be stained using a suitable reporter such as a dye having specificity towards bacterial cell membrane to facilitate viewing of the Mycobacteria.
  • This technique is also sometimes referred to as acid fast staining.
  • the technique is an inexpensive method, requiring little infrastructure and materials.
  • this method of diagnosing tuberculosis has been found to have only about 50% sensitivity, in other words, there is an equal possibility of detecting presence of tuberculosis as not detecting it when the patient actually has tuberculosis.
  • the lack of sensitivity is due to the inability of the existing sample preparation protocols to make available with equal probability all the Mycobacteria in the sample an equal opportunity to be stained.
  • sensitivity of the detection technology could be improved many fold by concentrating Mycobacterium from total sputum sample.
  • the sensitivity of the testing protocols is affected by the statistics of number of stained bacteria that is actually being viewed under a microscope, for example. This lack of sensitivity results in many patients suffering from tuberculosis not getting treated for it.
  • the traditional sputum liquefaction process typically involves freshly prepared reagents stable for few hours.
  • the sputum samples need to be decontaminated with alkali solution followed by treatment with liquefaction reagent.
  • successful downstream applications require extensive washing steps through centrifugation to remove traces of alkali and liquefaction reagents.
  • the current process thus inherits multiple sources of variability that include, for example, duration of decontamination with alkali, quality of liquefaction reagent prepared each day, washing conditions using centrifugation, and the like. These steps introduce a number of uncertainties and variability in the results.
  • samples are collected from multiple locations and brought to centralized facilities for detailed testing. This could particularly be adoptable in cases of MDR-TB diagnosis where one requires high-end complex technology for confirmed diagnosis.
  • the invention provides a composition for sputum sample preparation.
  • the composition of the invention comprises at least one reducing agent; and at least one chao trope.
  • the invention provides a method for preparing a sputum sample using the composition of the invention.
  • the invention provides a method for decontaminating sputum bacteria from contaminating bacteria using the composition of the invention.
  • the invention provides a sample preparation kit for sputum sample preparation comprising the composition of the invention.
  • the invention provides a method of diagnosis using a prepared sputum sample, wherein the prepared sputum sample is prepared using the method of the invention that includes use of the composition of the invention.
  • Staphylococcus aureus that have been prepared using reagents described herein;
  • Fig. 2 is a photograph of the sample culture of Mycobacterium smegmatis that has been treated with Reagent C to prepare the sample;
  • Fig 4 shows microscopy analysis of sputum sample prepared and stained with
  • Important species within the family of include avium, M. intracellularae, M. gordonae, M. tuberculosis, M. kansasii, M. fortuitum, M. chelonae, M. boyis, M. scrofulaceum, M. paratuberculosis, M. marinum, M. simiae, M. szulgai, M. intracellulare, M. xenopj., M. ulcerans, M. leprae, M.
  • M. tuberculosis which already infects millions of people each year, is an important Mycobacteria from an epidemiologic and clinical viewpoint.
  • M. averium, M. boyis, M. intracellularae, M. africanum, M. leprae, M. chelonae, M. paratuberculosis, and M. marinum are also significant from an epidemiological and clinical viewpoint.
  • sputum is mucus that is made available from the lower airways of the lungs. Sputum is useful for microbiological investigations of respiratory infections, such as tuberculosis. Without being bound to any theory, it is known that sputum is a complex polymeric structure made up of mucins. N and C terminal of the mucin monomers contain large number of cysteine residues. Disulphide bridges between cysteine molecules of mucin results in long and complex polymeric structure. In addition, heavy glycosylation of mucin renders a hydrophobic structure to it. The Mycobacteria, when present, are entrapped within the structure of mucins.
  • Sputum solubility may also vary from sample to sample depending on the complexity. Usually, when sputum is being extracted from a patient, some quantity of saliva is also extracted out. In existing diagnostic methods, microflora contained within the saliva may interfere with any diagnostic testing conditions.
  • the invention provides a composition for sputum sample preparation.
  • the composition of the invention comprises at least one reducing agent and at least one chao trope.
  • Useful chaotropes in the invention include, but not limited to, guanidinium salts such as guanidinium thiocyanate, guanidinium isothiocyanate, guanidinium hydrochloride.
  • the chaotrope used in the invention is guanidinium hydrochloride.
  • the chaotrope is present in a concentration range in the sample such that in the final liquid sample it is made available at a concentration that ranges from about 2 moles per liter to about 6 moles per liter.
  • the composition of the invention may also include a reporter.
  • Useful reporters in the invention include dyes. Dyes, as used herein, include coloring and staining reagents that provide a response at a wavelength suitable for observation using appropriate techniques. In some instances, dyes provide a response in visible wavelengths and thus can be observed using visual techniques such as microscopy. In other instances, the dyes provide responses in infra-red wavelength, and can be observed using, for example, an infra red spectrometer.
  • dyes may be fluorescent dyes that are capable of fluorescing. Such dyes are known in the art, and are contemplated to be included in this invention.
  • the reporter is a thiazole orange derivative dye.
  • Other exemplary reporters include, for example, nucleic acid binding dyes, viability dyes/reagents, surface markers, acid fast staining, and the like, and combinations thereof.
  • compositions of the invention are made by techniques known in the art, and may include, for example, solid mixing, blending, compounding, solution mixing, and the like. The exact choice of methods depend on various factors, such as, physical nature of components (for example, whether the components are solid or liquid), density, stability, and the like, and combinations thereof. In some instances, the components may be combined in solution in a common solvent, and subsequently the solvent is removed using known techniques, such as boiling, rotary evaporation, lyophilization etc. The exact methods for making the composition of the invention will become obvious to one skilled in the art.
  • the composition of the invention may be made available as a solid formulation or a liquid formulation. In some instances, the composition is a solid formulation. Solid formulations provide advantages such as of ease of transportation, better storage and stability, reduced wastage, ease of handling, and the like.
  • the composition of the invention is capable of liquefying sputum at room temperature within 10-15 minutes. Further, the composition is capable of being unitized and be provided as a solid formulation, wherein the solid formulation is used as such to liquefy sputum samples. This provides the added advantage of stability and storability at room temperature. Also, the composition is capable of preferentially inactivating or lysing contaminant bacteria like Escherichia coli, Staphylococcus aureus etc. while not lysing or inactivating any Mycobacteria present in sputum. The reporter when present then links to the Mycobacteria.
  • composition of the invention facilitates diagnosis of sputum for any afflictions such as tuberculosis using simple methods such as microscopy, infra-red readers, fluorescence readers, and the like.
  • a sputum sample may be liquefied that allows for diagnosis of afflictions such as tuberculosis, infections in cystic fibrosis patients, and the like.
  • the composition of the invention allows for using the whole sputum sample instead of only a portion of it, which now allows for rendering infection detection from a mere possibility to almost a certainty, as long as the sample contains the infection causing pathogen.
  • the method of the invention may include a step of shaking to facilitate intimate contact between components to provide the prepared sputum sample. While the method does not require other steps such as mixing, vortexing and the like to facilitate intimate contact between components, it may still be performed to provide the prepared sputum sample. It is known that shaking, mixing vortexing, and other steps done to a certain extent causes aerosol build-up, this is considered to be a health risk for the operators as Mycobacteria spread easily through aerosol. Hence, it is generally desirable to develop compositions and methods that do not involve such steps to any extent.
  • the present invention provides compositions and methods that address issues related to containment of aerosolized Mycobacteria to circumvent this important health issue.
  • the sample may be subjected to a centrifuging step to concentrate the sample to improve Mycobacteria availability in a suitable manner.
  • the method of the invention may further comprise filtering the prepared sputum sample. Filtration is performed through a suitable filtering medium.
  • Useful filtering medium in the invention include, but not limited to, paper-based filters, nylon filters, Teflon filters, silica-based filters, glass wool, and the like, and combinations thereof. Several such variations and combinations of filters are known to one of ordinary skill in the art, and are contemplated to be within the scope of the invention.
  • an annular filter is used.
  • glass wool is used as the filtration medium.
  • Other suitable filters are known and can be used appropriately. This filtration ensures that no extraneous precipitated material interferes with the diagnostic testing, thus increasing sensitivity of the testing procedure. Filtration may also be effected by the application of suction or vacuum.
  • the method of the invention further comprises concentrating the prepared sputum sample.
  • Methods of concentrating the prepared sputum sample include, for example, filtering, centrifuging, induced sedimentation technique, electrical field, magnetic field, affinity based separation, charge based adhesion, and the like, and combinations thereof. Filtration may be achieved using methods and apparatus as described herein.
  • Affinity based separation include the use of polymeric materials that show affinity towards Mycobacteria, which are made available commercially from a variety of sources, such as TB BeadsTM available from Microsens, London, UK, and have also been described in for example US2010/0143883. Such polymeric materials are known in the art, and are contemplated to be within the scope of the invention.
  • the prepared sample is rendered conducive for any one or more testing methods already known in the art.
  • the prepared sample may be subjected to direct detection methods such as conventional AFB smear test (ZN staining) using light microscopy.
  • Other forms of microscopy are also known to one skilled in the art, and include, for example, light microscopy, electron microscopy such as TEM and SEM, and others, and combinations thereof, and are contemplated to be within the scope of the invention.
  • Fluorescence based microscopy or by using reporters that include, for example, but not limited to, nucleic acid binding dyes, viability dyes/reagents, surface markers, acid fast staining, or detection of the Mycobacteria using intrinsic autofluorescence.
  • a particular fluorescence device useful for detection includes the commercially available fluorescent readers from ReaMetrix Inc., USA. Such techniques are well-known in the art.
  • the invention provides a method for diagnosing using the prepared sputum sample, and/or the filtered prepared sputum sample, as the situation demands, for the presence or absence of Mycobacteria.
  • the method for testing may include any of the qualitative or quantitative tests known in the art. For example, a semi quantitative test using the AFB smear test may be performed using the prepared sputum sample to provide results that fall under the categories of 1+, 2+, 3+ and the like using the well developed techniques in the art.
  • compositions and methods described herein allows for short turnaround times (-30 minutes) from sample collection to diagnostic data interpretation.
  • the rapidity and high sensitivity of the process allows same day diagnosis and treatment a reality for the patients.
  • the quick turnaround time prevents further dissemination of disease within the community thereby reducing the epidemic burden.
  • the prepared sputum sample is further viable for culture based diagnostic protocols, and could be used for nucleic acid isolation and amplification based diagnostics.
  • the compositions and methods described herein can be adopted in resource poor settings nearer to patients.
  • the prepared sputum sample may be stored for future processing for diagnosis.
  • the prepared sample may be further processed to facilitate transportation of the prepared sample to a suitable location for testing, diagnosis or detection.
  • Techniques for storage of nucleic acid are known in the art, and include, for example, FTA® card or FTA® elute commercially made available from, for example, General Electric, USA and the like.
  • the sample may be processed to inactivate the bacteria while maintaining the cell intact.
  • the nucleic acid may be isolated and optionally amplified.
  • composition and method of the invention greatly improves time to result and sensitivity - the time to diagnosis is less than 30 minutes and more than 90% sensitivity of detection of mycobacterium species in sputum may be achieved. This will also facilitate improved detection of "smear negative, culture positive" TB samples as described in current literature, which is a major challenge in TB diagnosis arena. Further, the composition and method of the invention are designed for room temperature processing without any specialized instrumentation, thus enabling the resource poor settings to diagnose sputum samples with high degree of sensitivity for any appropriate afflictions, such as tuberculosis.
  • the sample preparation kit is a cylinder with a thin solid layer located near the bottom portion of the cylinder, wherein the solid layer comprises the composition of the invention.
  • the filtration unit such as a membrane filter.
  • a pre-filter may also be disposed ahead of the membrane filter.
  • the sample preparation kit may be made up of disposable plastics easy for disposal.
  • the sample preparation kit may further include components to enable inactivation of Mycobacteria with intact cell, which thus facilitates direct detection of the Mycobacteria through appropriate methods as described herein.
  • the sample preparation kit may be used for isolation and storage of bacterial nucleic acid as described herein, and the nucleic acid may be used for future detection using methods described herein.
  • Techniques for storage of nucleic acid are known in the art, and include, for example, FTA® card or FTA® elute and the like.
  • the stored nucleic acid samples could be transported to suitable locations (such as a centralized testing laboratory) for further testing or for repository collections.
  • sample preparation kit Other suitable components for the sample preparation kit will become obvious to one skilled in the art.
  • Some exemplary components include, for example, a cartridge or holder or cassette or similar arrangement for membrane to enable better capture of concentrated prepared sample.
  • the membrane may be covered with a slit like arrangement or similar arrangement to ensure uniform distribution of the filtrate on the cartridge.
  • the sample preparation kit may comprise a flat base to keep the membrane flat.
  • the sample preparation may further comprise a separate connector for collecting the waste suitably located for secure collection and disposal.
  • the connector for waste collection may comprise an option for connecting multiple sample preparation kits to a vacuum manifold or mechanical hand pump or similar device, which when operated will create pressure differential between the protected chambers and rest of the assembly to pass the sample through the cartridge or holder or cassette or similar arrangement containing the membrane.
  • the waste collection container is configured to be permanently closed for easy disposal.
  • the sample preparation kit is configured to be leak proof and, once sputum sample has been collected, it is configured to be permanently sealed to ensure no contamination occurs. While one particular configuration of the sample preparation kit has been described here, other variations will become obvious to one skilled in the art, and are contemplated to be within the scope of the invention. For example, the components of the composition may be made available individually, and brought together in a liquid form through secure ports upon a trigger event, such as opening of one or more valves.
  • Range of concentrations of critical components of the sputum preparation reagent compositions were tested. Fixed volume of the reagent was added to sputum sample and mixed by inverting the tube for 10 times. The sputum sample preparation process was carried out either at 37°C for 15 minutes or at room temperature for 10 minutes. The effect of sputum preparation reagent was recorded after stipulated time. Modified sputum compaction assay was carried out on prepared sputum to estimate the effect of different compositions of the sample preparation reagent on the nature of the final prepared sputum sample. The prepared sputum sample was characterized for their pellet content in the following qualitative manner: Minute size pellet; Medium Size pellet; and Large Size size pellet. Following reagent concentrations were tested
  • TCEP concentration was varied from lOmM to lOOmM
  • the three reducing agents are compatible with sputum preparation processing at room temperature for 10 minutes.
  • EXAMPLE 3 The following reagents were used to prepare sputum sample in the manner described herein. The final prepared sputum samples were characterized quantitatively and the results are provided in Table 2.
  • Reagent A Reducing agent: Beta-Mercaptoethanol
  • Reducing agent Tris(2-carboxyethyl)phosphine hydrochloride
  • Reducing agent Mixture of N- Acetyl Cysteine and Lysine
  • composition described in Reagent C is found to be stable at room temperature. Further, the prepared sputum sample shows higher volume of liquid sample and smaller pellet sizes, which is indicative better solubility of the sputum sample thereby release of entrapped Mycobacteria.
  • compositions of the selected buffers are designed to include high concentration of chaotrope to disrupt cell wall of commonly occurring Gram positive and Gram negative bacteria.
  • sputum sample preparation reagent To test the effectiveness of the sputum sample preparation reagent the following experiment was carried out. Logarithmically growing Escherichia coli (representing Gram -ve bacteria), Staphylococcus aureus (representing Gram +ve bacteria) and Mycobacterium smegmatis (noninfectious Mycobacteria) was treated with the selected sputum sample preparation reagents as described in Example 3. Treated bacterial cells were washed and plated on suitable solid culture medium. The plates were incubated at 37°C incubator for appropriate period for bacterial growth. Growth of E_. coli and S_.
  • FIG. 1 shows photographs of the cultures of S_. aureus and E_. coli, generally represented by numeral 10.
  • Numeral 12 represents the photograph of sputum sample containing S_. aureus that was treated with Reagent A
  • 14 represents the photograph of the sputum sample containing S_, aureus that was treated with Reagent C
  • 16 shows the photograph of the sample containing S_. aureus that was treated with Reagent E.
  • numeral 18 represents the photograph of sample containing E, coli that was treated with Reagent A
  • 20 represents the photograph of the sample containing coli that was treated with Reagent C
  • 22 shows the photograph of the sputum sample containing E_. coli that was treated with Reagent E
  • Numerals 24 and 26 show the photographs of the samples containing S_. aureus and E_. coli respectively that were not treated with any of the reagents.
  • the culture plates treated with all three sputum sample preparation reagents did not show any bacterial growth in case of E coli and Staphylococcus aureus suggesting the reagent is suitable for decontamination of sputum samples.
  • FIG. 2 shows photographs represented generally by numeral 28, of a sample containing Mycobacterium smegmatis that was treated with Reagent C as represented by numeral 30, as compared to the negative control sample that was not treated with any of the reagents as represented by numeral 32.
  • Treatment of Mycobacterium smegmatis with sputum sample preparation Reagent C has no effect on growth of bacteria after 3 days of culture.
  • the data suggest suitability of the sputum sample preparation reagents in downstream processing like, direct detection of Mycobacteria, nucleic acid staining and culture based detection of Mycobacterium in TB diagnosis.
  • compositions of the invention can suitably liquefy the sputum and can further be designed to differentially inactivate and/or lyse any contaminant bacteria present in the sputum while excluding the Mycobacteria.
  • the sputum preparation compositions and methods described herein neutralize the effect of any contaminating bacteria without compromising the detectability of the Mycobacteria.
  • Numeral 36 shows a photograph of the sputum sample as received.
  • Numeral 38 represents the sputum sample to which the liquefaction reagent was added.
  • Numeral 40 shows a photograph of the tube containing the sputum sample with the liquefaction reagent after 2 minutes of addition of the reagent.
  • Numeral 42 is a photograph of the tube containing the sputum sample with the liquefaction reagent after 10 minutes of addition of the reagent.
  • Numeral 44 shows a photograph of the pellet containing the Mycobacterium after centrifugation.
  • the resuspended pellet was used for making a smear on glass slide as suggested by WHO protocol for smear preparation.
  • the smear was air-dried and heat fixed and stained with acid fast staining following WHO recommended Ziehl-Neelsen staining.
  • Mycobacterial cells were clearly visible in light microscopy as distinct violet rod shaped cells in sputum samples positive for Tuberculosis infection.
  • the sputum samples of confirmed negative for Tuberculosis did not produce any staining pattern characteristic of Mycobacteria staining.
  • Fig 4 shows microscopy analysis of sputum samples prepared and stained with ZN staining protocol as described in Example 5 depicted by numeral 46.
  • the left panel represented by numeral 48, shows rod shaped Mycobacteria cells in TB positive sputum sample stained with ZN staining protocol as depicted by numeral 52; the right panel, depicted by numeral 50 shows result for identically processed TB negative sputum sample.
  • Tuberculosis Control Programme (RNTCP) center St John's Hospital, Bangalore, India were processed inside "Biosafety Level 3 facility" following established protocols.
  • Reagent C described in Example 3 was used for sputum sample preparation following the method described as follows:
  • Comparative Example 2 The method described in Comparative Example 2 is the reference method based on which the calculations have been performed and expressed as percentages in Table 3.
  • the method of the invention using Reagent C identified additional 50% TB positive samples as compared to the WHO protocol followed at the RNTCP center.
  • the distribution of positive samples has a distinctive pattern; the method of Example 6 identified significantly more positives in the "scanty" and "AFB 1+” categories as shown in Table 4.
  • TABLE 4 Distribution of TB patient sample based on the methods described herein, wherein the data is represented as percentage of total samples analyzed

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Biomedical Technology (AREA)
  • Molecular Biology (AREA)
  • Physics & Mathematics (AREA)
  • Chemical & Material Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Pathology (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
  • Investigating Or Analysing Biological Materials (AREA)

Abstract

Dans un aspect, l'invention concerne une composition pour la préparation d'échantillons de crachats. La composition comprend au moins un agent réducteur et au moins un agent chaotropique. Dans un autre aspect, l'invention concerne un procédé de préparation d'un échantillon de crachat liquéfié en utilisant la composition de l'invention. Dans encore un autre aspect, l'invention propose un kit de préparation d'échantillons pour la préparation d'échantillons de crachats comprenant la composition de l'invention. Dans d'autres aspects, l'invention fournit des techniques de diagnostic utilisant les compositions et les procédés de préparation d'échantillons décrits ici.
PCT/IB2012/050628 2012-02-10 2012-02-13 Compositions et procédés de préparation d'échantillons de crachats, et kit WO2013117967A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
IN3410/CHE/2011 2012-02-10
IN3410CH2011 2012-02-10

Publications (1)

Publication Number Publication Date
WO2013117967A1 true WO2013117967A1 (fr) 2013-08-15

Family

ID=45819245

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/IB2012/050628 WO2013117967A1 (fr) 2012-02-10 2012-02-13 Compositions et procédés de préparation d'échantillons de crachats, et kit

Country Status (1)

Country Link
WO (1) WO2013117967A1 (fr)

Cited By (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN104189473A (zh) * 2014-09-19 2014-12-10 强红枫 一种治疗结核病的药物组合物及其制备方法
WO2017055494A1 (fr) * 2015-10-02 2017-04-06 Roche Diagnostics Gmbh Réactif préanalytique de mycobactéries
CN108414313A (zh) * 2018-02-27 2018-08-17 中国科学院北京基因组研究所 一种痰液液化试剂、含有其的试剂盒及其应用
JP2020503071A (ja) * 2017-01-16 2020-01-30 スペクトラム・ソリューションズ・エルエルシーＳｐｅｃｔｒｕｍ Ｓｏｌｕｔｉｏｎｓ Ｌ．Ｌ．Ｃ． 核酸保存溶液ならびに製造方法および使用方法
CN111218444A (zh) * 2020-04-24 2020-06-02 广州安必平医药科技股份有限公司 一种痰液保存液
WO2020160307A1 (fr) * 2019-01-30 2020-08-06 Cedars-Sinai Medical Center Traitement et analyse d'échantillons biologiques liquides visqueux
WO2022195531A1 (fr) * 2021-03-18 2022-09-22 Vimal Soomnath Pumposh Dispositif à usage unique permettant d'identifier des expectorations

Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1996009308A1 (fr) * 1994-09-19 1996-03-28 Promega Corporation Procede particulierement efficace pour isoler des substances cibles a l'aide d'un dispositif de separation a echantillons multiples
WO2000000813A1 (fr) * 1998-06-30 2000-01-06 Lamina, Inc. Composition fixative cytologique et histologique et procedes d'utilisation
WO2000006780A1 (fr) * 1998-07-31 2000-02-10 Ambion, Inc. Procedes et reactifs permettant de preserver l'arn dans des prelevements cellulaires et tissulaires
EP1804045A1 (fr) * 2005-12-30 2007-07-04 Qiagen GmbH Procédé pour traitement d'un échantillon biologique
EP1965190A1 (fr) * 2007-02-27 2008-09-03 Qiagen GmbH Fixation d'un échantillon biologique
US20100143883A1 (en) 2006-11-29 2010-06-10 Microsens Medtech Limited Capture of mycobacteria like micro-organisms

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1996009308A1 (fr) * 1994-09-19 1996-03-28 Promega Corporation Procede particulierement efficace pour isoler des substances cibles a l'aide d'un dispositif de separation a echantillons multiples
WO2000000813A1 (fr) * 1998-06-30 2000-01-06 Lamina, Inc. Composition fixative cytologique et histologique et procedes d'utilisation
WO2000006780A1 (fr) * 1998-07-31 2000-02-10 Ambion, Inc. Procedes et reactifs permettant de preserver l'arn dans des prelevements cellulaires et tissulaires
EP1804045A1 (fr) * 2005-12-30 2007-07-04 Qiagen GmbH Procédé pour traitement d'un échantillon biologique
US20100143883A1 (en) 2006-11-29 2010-06-10 Microsens Medtech Limited Capture of mycobacteria like micro-organisms
EP1965190A1 (fr) * 2007-02-27 2008-09-03 Qiagen GmbH Fixation d'un échantillon biologique

Cited By (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN104189473A (zh) * 2014-09-19 2014-12-10 强红枫 一种治疗结核病的药物组合物及其制备方法
WO2017055494A1 (fr) * 2015-10-02 2017-04-06 Roche Diagnostics Gmbh Réactif préanalytique de mycobactéries
JP2020503071A (ja) * 2017-01-16 2020-01-30 スペクトラム・ソリューションズ・エルエルシーＳｐｅｃｔｒｕｍ Ｓｏｌｕｔｉｏｎｓ Ｌ．Ｌ．Ｃ． 核酸保存溶液ならびに製造方法および使用方法
CN108414313A (zh) * 2018-02-27 2018-08-17 中国科学院北京基因组研究所 一种痰液液化试剂、含有其的试剂盒及其应用
WO2020160307A1 (fr) * 2019-01-30 2020-08-06 Cedars-Sinai Medical Center Traitement et analyse d'échantillons biologiques liquides visqueux
US20220186281A1 (en) * 2019-01-30 2022-06-16 Cedars-Sinai Medical Center Method for processing and analysis of viscous liquid biological samples
CN111218444A (zh) * 2020-04-24 2020-06-02 广州安必平医药科技股份有限公司 一种痰液保存液
CN111218444B (zh) * 2020-04-24 2020-09-01 广州安必平医药科技股份有限公司 一种痰液保存液
WO2022195531A1 (fr) * 2021-03-18 2022-09-22 Vimal Soomnath Pumposh Dispositif à usage unique permettant d'identifier des expectorations

Similar Documents

Publication Publication Date Title
WO2013117967A1 (fr) Compositions et procédés de préparation d'échantillons de crachats, et kit
Emerson et al. Schrödinger’s microbes: tools for distinguishing the living from the dead in microbial ecosystems
US9944974B2 (en) Method for the specific isolation of nucleic acids of interest
US7858363B2 (en) Plasmid DNA isolation
Marei et al. Evaluation of a rapid bacteriophage-based method for the detection of Mycobacterium tuberculosis in clinical samples
US9719128B2 (en) Selective ultrasonic lysis of blood and other biological fluids and tissues
An et al. Development and calibration of real-time PCR for quantification of airborne microorganisms in air samples
US11753616B2 (en) Composition and method for stabilizing and maintaining the viability of hardy microorganisms
EP1774041A2 (fr) Compositions et méthodes permettant de préparer des acides nucléiques à partir d'échantillons microbiens
US9677981B2 (en) Sample concentrator and method of use
EP3348634B1 (fr) Procédé de séparation d'acide nucléique contenu dans un échantillon comprenant un acide nucléique, et dispositif associé
US20220195490A1 (en) Method for recovering microbial cells
KR20200062321A (ko) 샘플에서 온전한 미생물의 농도를 결정하는 방법
Mangayarkarasi et al. Jayaprakash Ergonomic diagnostic tool based on chip mini RT-PCR for diagnosis of pulmonary and extra pulmonary tuberculosis
JP2013042750A (ja) 核酸抽出方法、核酸抽出試薬キットおよび核酸抽出用試薬
Shenai et al. Evaluation of a new phage amplification technology for rapid diagnosis of tuberculosis
US8758990B2 (en) Methods for detecting and differentiating Mycobacterium genus and Mycobacterium avium complex in a sample or culture
Iqbal et al. Importance of polymerase chain reaction in diagnosis of pulmonary and extra-pulmonary tuberculosis
JP2004180551A (ja) 微生物の収集方法およびそれを用いた遺伝子の増幅方法若しくは検出方法
Ok et al. Comparison of swabbing solution volume and gDNA extraction kits on DNA recovery from rigid surface
WO2013117968A1 (fr) Procédés de lyse de bactéries provenant d'un échantillon et d'isolement des composants cellulaires de celles-ci, et kits correspondants
US20120264119A1 (en) New method for decontamination and processing of clinical specimens from a patient
Kretzer et al. Sample preparation–an essential prerequisite for high-quality bacteria detection
KR20180136908A (ko) 병원체 검출 방법
Betters et al. Technical Evaluation of Sample-Processing, Collection, and Preservation Methods

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 12708588

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 12708588

Country of ref document: EP

Kind code of ref document: A1