EP0335866A1 - Test device and method of characterizing biological material - Google Patents

Test device and method of characterizing biological material

Info

Publication number
EP0335866A1
EP0335866A1 EP87906634A EP87906634A EP0335866A1 EP 0335866 A1 EP0335866 A1 EP 0335866A1 EP 87906634 A EP87906634 A EP 87906634A EP 87906634 A EP87906634 A EP 87906634A EP 0335866 A1 EP0335866 A1 EP 0335866A1
Authority
EP
European Patent Office
Prior art keywords
reagent
zone
zones
test device
well
Prior art date
Legal status (The legal status 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 status listed.)
Withdrawn
Application number
EP87906634A
Other languages
German (de)
English (en)
French (fr)
Inventor
Anne Bolmström
Magnus Ericsson
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Biodisk AB
Original Assignee
Biodisk AB
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 Biodisk AB filed Critical Biodisk AB
Publication of EP0335866A1 publication Critical patent/EP0335866A1/en
Withdrawn legal-status Critical Current

Links

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/5302—Apparatus specially adapted for immunological test procedures
    • G01N33/5304—Reaction vessels, e.g. agglutination plates
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • 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
    • C12Q1/18—Testing for antimicrobial activity of a material

Definitions

  • the present invention relates to a test device for characterization of biological materials by means of microbiological, immunological and/or biochemical analyses.
  • the invention further comprises a method of characterizing biologic materials.
  • test device for characterization of biological materials by means of microbiological, immunological and/or biochemical analyses, in which test device there is a plurality of. wells, each comprising two distinct zones communicating with one another, one of which is a reagent zone and the other a reading zone.
  • the shape of the test device can be rectangular, round or oval, and the wells in the test device can be of different designs. This invention is not restricted to any special shape of the test device or any special designs according to which the wells are made in the test device. Some embodiments of the test device according to the invention are shown in Fig. 1 consisting of the test plate A, a reagent zone B and a reading zone C of each well.
  • the shape of the wells can vary within the scope of the invention and according to one embodiment the wells have the shape of a key-hole, the circular portion being the reagent zone and the elongated portion the reading zone.
  • each well consists of an elongated portion comprising the two zones where constricting projections are arranged between the zones.
  • the ends of the wells can be rounded off or straight.
  • each well consists of a square reagent zone and a squar reading zone which are of different sizes.
  • Fig. 2 shows some different embodiments of wells according to the invention, B designating the reagent zone, C the reading zone and D the constricting projections.
  • other shapes of the wells can also be used and of importance is that the carrier of the active substance is retained in the reagent zone. Tapering zones for example are also possible.
  • the wells can also have such a shape that the reagent zone and reading zone are lying above each other, i. e. in the vertical plane of the test device or consecutively, i. e. in the horizontal plane of the test device.
  • the designations in the figure have the meanings indicated above.
  • the reading zone can be covered from above. In this figure the area where communication between the two zones takes place has been designated E.
  • the test device is preferably of some inert, material such as plastic or glass and it can be in one piece or made with wells after which possible constricting projections are applied.
  • a perspective view of a well in the shape of a key-hole comprising a reagent zone B and a reading zone C.
  • a carrier F is applied which is impregnated with the antimicrobial substance, the minimum inhibitory concentration of which is to be determined.
  • a suspension G of microorganisms in a medium is uniformly distributed across the two zones B and C and covering the top of the carrier F.
  • A, B and C are as defined above.
  • the number indiciated in the figure are markings of the carriers. In this case the amount of antibiotic is indicated on the respective carrier.
  • the carriers marked by plus and minus signs are positive and negative controls, respectively.
  • This invention also relates to a method of characterizing biological materials by means of microbiological, immunological and/or biochemical analyses, according to which the characterization is carried out in two zones communicating with one another, one of which is a reagent zone and the other a reading zone, a reagent on a carrier is applied in the reagent zone and the biologic material to be characterized is distributed across the two zones, after which the whole is mixed, incubated, if required, and a possible reaction is read in the reading zone.
  • the reagent is introduced into the reagent zone on a suitable carrier such as a porous or nonporous disc, a bead coated with the required amount of reagent, or it can be added in the form of a compressed tablet containing the required amount of reagent.
  • the biological material to be characterized is thereafter applied into each well.
  • the biological material is present in or tranferred into liquid form and is preferably applied by means of a pipette, in exact volumes to each well.
  • the test device is thereafter rotated or shaken cautionsly to obtain a uniform mixture of the reagent and biological material in the whole well. De pending on the type of analyses the test device is rotated, shaken, centrifuged and/or incubated for part of or during the whole reaction after which the results in the different wells are read.
  • the whole procedure can preferably be automated.
  • An apparatus can e.g. dispense the active carriers, and a system can automatically pipette the test sample.
  • the reading can also be conducted using a suitable spectophotometric apparatus illuminating the reading zone.
  • test devices can be used when characterizing biological materials such as in microbiological, immunological and biochemical analyses, for example determination of concentrations of substances which are active against microorganisms or other biologic cells.
  • biologic materials can for example be other biologic cells, antibodies, antigens, enzymes, blood, serum or other biologic mateial in the form of liquids.
  • the reagent may consist of biologic or chemically active substances such as antimicrobial substances, microbiologically active substances, substances with an inhibiting, lethal, toxic, mutagenic or growth-promoting effect on cells.
  • Test devices with unlabelled wells can be used for different kinds of reagents and in different concentrations. The latter are applied by the aid of suitable carriers on which the reagent is applied in a desired concentration. Carriers with reagents are packed in separate units and the type of reagent as well as amount is preferably indicated by specific labelling on the carriers itself. Altgruatively, the wells on the test device can be individually labelled for different types of analyses.
  • microorganism includes bacteria such as enterobacteriaceae, staphyloccoci, streptococci, hemophilus, neisseriaceae, bacteriodes and clostridia, mycobacteria, actinomyces, mycoplasma, nocardia, virues and fungi such as mould-fungus, yeast-fungus and Candida.
  • biological cells comprises cancer cells, normal human cells, animal cells, insect cells and plant cells.
  • Antimicrobial substances are antibiotics, for example aminoglycosides, ⁇ -lactam antibiotics, macrolide antibiotics, polymyxins, polypeptides and other chemotherapeut ics such as sulf onamides, antimycot ics, for example 5-fluorocytosine, amphotericin, antiviral agents such as adenine arabinoside (Ara-A), trif luorothymidine, antituberculous agents such as i ⁇ oniazide and cycloserine, as well as disinfectants, antiseptics and preservative such as chlorohexidine, ethanol and bensalkonium chloride.
  • antibiotics for example aminoglycosides, ⁇ -lactam antibiotics, macrolide antibiotics, polymyxins, polypeptides and other chemotherapeut ics such as sulf onamides, antimycot
  • Substances that can have an inhibitory, lethal, toxic or mutagenic effect on cells are anticancer agents, e.g. antracyclines, mitomycins, cancerognic/mutagenic substances such as dimethyl nitrosamine, herbicides such as paraquat and pesticides as well as insecticides such as dicophane (DDT) and chloroquine.
  • anticancer agents e.g. antracyclines, mitomycins, cancerognic/mutagenic substances such as dimethyl nitrosamine, herbicides such as paraquat and pesticides as well as insecticides such as dicophane (DDT) and chloroquine.
  • Microbiologically active substances besides the indicated antimicrobial substances are bacteriocin, for example colicin, enzymes, for example peroxidases, various growth factors such as amino acids, vitamins, carbohydrates and other nutrients, various body fluids and components thereof such as serum, blood and specific components in serum, for example ⁇ -lysine.
  • Biologically active substances are interferons such as human interferon, HuIFN- ⁇ , IF4N- ⁇ , enzymes such as ⁇ -lactamases, proteases and ureases, enzyme specific substrates such as nitrates, carbohydrates and proteins, ⁇ -lactam antibiotics, aminoglycosides and chloramphenicol, antigens such as immuneglobulins, polysaccharides and toxins from microorganisms. It is for instance possible to determine the minimum inhibitorys concentration of an antimicrobial substance by means of the method of the invention. The determination is carried out in two zones communicating with each other, one being a reagent zone and the other a reading zone.
  • the antimicrobial substance is introduced into the reagent zone on a suitable carrier such as one of those indicated, for example a porous disc.
  • a suspension of the microorganism, the sensitivity of which is to be determined, is thereafter introduced into each well.
  • the suspension is in a suitable culture medium. This is preferably applied by means of a pipette, in exact volumes of microorganism in substrate to each well.
  • the test device is thereafter tilted carefully to achieve a uniform mixture of the substance and the suspension of microorganisms in the whole well. After incubation e.g. at 37°C over night, the results are read, i.e. inhibition of growth of the microorganism in the different wells.
  • the antimicrobial substance is present in different concentrations on labelled carriers in the different wells it is easy to establish directly the minimum inhibitory concentration of the substance.
  • MBC minimum bactericidal concentration
  • the inactivating substance is preferably applied to the reagent zone on a carrier, such as any of those described earlier. After incubation, the presence growth is read and the MCB is determined.
  • microorganisms By means of the method of the invention it is also possible to identify microorganisms by determining their biochemical patterns based on fermentation, oxidation or assimilation of different substances.
  • a test substance is introduced into the reagent zone with or without the addition of other reagents for the particular reaction, for example a pH-colour indicator on a suitable carrier, such as a bead coated with the appropriate amount thereof.
  • a suspension of the microorganism, the biochemical pattern of which is to be determined is thereafter applied to each well.
  • the supension is made in a suitable culture medium. This is applied for instance with a pipette in exact amounts of microorganisms suspended in culture medium to each well.
  • test device To detain a uniform mixture of the substances and the suspension of microorganisms in the whole well, the test device is carefully rotated or shaken. After incubation the result is read in different ways depending on the biochemical reaction used.
  • the assimilation pattern is established for instance by detecting growth/no growth, while fermentation or oxidation patterns can be indicated by colour change of a pH-indicator.
  • substances used in the determination of assimilation patterns are carbohydrates such as glucose, mannose, lactose, raffinose, melibiose, cellobiose, glycerol, xylitol and gluconate. These substances can be combined with suitable pH-indicators such as phenol red, bromothymol blue, gromocresol purple for fermentation and oxidation studies.
  • the method of the invention can also be used for characterizing enzymatic patterns of biologic materials (so-called zymograms) by determining enzyme/substrate reactions, in which the determination is carried out in the two zones communicating with one another.
  • a chromogenic substrate for example a naphthalene derivative on a suitable carrier, for example in the form of a tablet containing the desired amount of the chromogenic substrate, is introduced into the reagent zone.
  • a suspension of the biological material, the zymogram of which is to be determined, is thereafter added to each well. This is preferably applied by means of a pipette, in exact volumes of biological material to each well.
  • test device is thereafter rotated carefully so that a uniform mixture of chromogenic substrate and the suspension of the biological material is obtained through out the whole well.
  • the test device is then incubated and the result is thereafter read, i.e. colour changes of the enzyme/substrate reactions in the different wells.
  • Chromogenic substrates that can be used in zymogram studies are for instance chromogenic naphthalene derivatives such as naphthyl phosphate to detect phosphatases, naphthyl butyrate to detect esterases, naphthyl myristate to detect upases, naphthylamides to detect araylamidases and trypsin, naphtyl galactopyranosides to detect galactosidases, naphthyl glucopyranosides to detect glucosidases, nitrocephin to detect ⁇ -lactamases etc.
  • chromogenic naphthalene derivatives such as naphthyl phosphate to detect phosphatases, naphthyl butyrate to detect esterases, naphthyl myristate to detect upases, naphthylamides to detect araylamidases and trypsin, naphtyl galactopyranosides to detect
  • the method of the invention can be used for characterizing and quantifying different biologically active substances based on their immunologic properties.
  • a specific antibody-latex conjugate is applied in the reagent zone and suspension of the biological material, the serologic pattern of which is to be determined, is uniformly distributed across the two zones.
  • the device is thereafter rotated to obtain a uniform mixture of the reagent and the sample.
  • An antigen-antibody reaction resulting in agglutination, i.e. clumping, will take place if the sample contains antigens which are specific to antibody-conjugate present in the reagent.
  • bacterial species can for example be serotyped and examples of bacterial species that can be classified by means of a serologic pattern are salmonella, shigella and streptococci. Similarly it is possible to determine the amount of antibodies in a biological material by using an antigen-latex conjugate.
  • one can determine the amount of an active substance that may inhibit an immunologic reaction. Determination of antibiotics in serum can be carried out by this technique.
  • a definite but varying amounts of an antibiotic-antibody conjugate are applied in the reagent zones a suitable carrier.
  • the serum sample or a dilution thereof, the corresponding antibiotic concentration of which is to be determined, is uniformly distributed in the two zones and mixed by means of rotation of the device.
  • a definite amount cf the specific antibiotic-protein conjugate bonded to latex particles (antigen unit) is thereafter applied to the reagent zone and the device is further rotated for the antigen-antibody reaction.
  • the reaction between antigen and antibody units results in agglutination.
  • the agglutination is inhibited in the different wells up to the maximum concentration of antibody units to which the antibiotic sample can be totally bonded.
  • this concentration of antibody units is exceeded the free antibody units now remaining react with the antigen units and result in agglutination.
  • the result thereof is compared to e.g. a standard curve which has been obtained by determining in advance the minimum amount of antibiotic inhibiting the different concentrations of antibody units by using standard solutions of this antibiotic.
  • Active substances in biologic liquides that can be quantified by means of an immunologic technique are antimicrobial substances, antibodies, antigens such as toxins and polysaccharides from microorganisms.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Immunology (AREA)
  • Organic Chemistry (AREA)
  • Molecular Biology (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Biotechnology (AREA)
  • Urology & Nephrology (AREA)
  • Microbiology (AREA)
  • Analytical Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Wood Science & Technology (AREA)
  • Zoology (AREA)
  • Hematology (AREA)
  • Biomedical Technology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Genetics & Genomics (AREA)
  • Food Science & Technology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Cell Biology (AREA)
  • Toxicology (AREA)
  • Biophysics (AREA)
  • General Engineering & Computer Science (AREA)
  • Medicinal Chemistry (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)
  • Automatic Analysis And Handling Materials Therefor (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
  • Investigating Or Analysing Biological Materials (AREA)
EP87906634A 1986-04-04 1987-09-28 Test device and method of characterizing biological material Withdrawn EP0335866A1 (en)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
SE8601511A SE460472B (sv) 1986-04-04 1986-04-04 Testbricka och saett foer att karaktaerisera biologiska material
PCT/SE1987/000439 WO1989002927A1 (en) 1986-04-04 1987-09-28 Test device and method of characterizing biological material

Publications (1)

Publication Number Publication Date
EP0335866A1 true EP0335866A1 (en) 1989-10-11

Family

ID=26659316

Family Applications (1)

Application Number Title Priority Date Filing Date
EP87906634A Withdrawn EP0335866A1 (en) 1986-04-04 1987-09-28 Test device and method of characterizing biological material

Country Status (8)

Country Link
EP (1) EP0335866A1 (da)
JP (1) JPH02501349A (da)
AU (1) AU8037187A (da)
DK (1) DK252489D0 (da)
FI (1) FI892489A7 (da)
NO (1) NO891892L (da)
SE (1) SE460472B (da)
WO (1) WO1989002927A1 (da)

Families Citing this family (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
GB8915938D0 (en) * 1989-07-12 1989-08-31 Proteus Biotech Ltd Apparatus for microbiological testing
JPH0832922A (ja) * 1994-07-14 1996-02-02 Hitachi Ltd 磁気記録再生装置
KR101711105B1 (ko) 2016-04-21 2017-03-06 주식회사 퀀타매트릭스 신속한 항생제 감수성 검사를 위한 멀티-웰 기반의 세포배양검사 장치
JP7076203B2 (ja) * 2017-12-13 2022-05-27 株式会社日立ハイテク 細菌検査用抗菌剤導入プレート、および透明プレート

Family Cites Families (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
DK97670C (da) * 1959-03-13 1964-01-06 Warner Lambert Pharmaceutical Indikator til kvantitativ bestemmelse af mængden af et biokemisk stof i en legemsvæske.
FR1356317A (fr) * 1963-02-06 1964-03-27 Dispositif et procédé pour l'identification de bactéries isolées
GB1006774A (en) * 1963-10-07 1965-10-06 Promoveo Improvements in devices for the study of the fermentative characteristics of living cells
FR2029138A5 (en) * 1969-01-14 1970-10-16 Colobert Louis Chemical reaction indication for biochemical - reactions
SE343950B (da) * 1970-07-22 1972-03-20 S Johansson
GB1390766A (en) * 1971-04-26 1975-04-16 Beecham Group Ltd Bacteriological test method
FI57128C (fi) * 1978-11-21 1980-06-10 Orion Yhtymae Oy Saett att identifiera mikroorganismer

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO8902927A1 *

Also Published As

Publication number Publication date
DK252489A (da) 1989-05-24
FI892489A0 (fi) 1989-05-23
AU8037187A (en) 1989-04-18
WO1989002927A1 (en) 1989-04-06
DK252489D0 (da) 1989-05-24
NO891892D0 (no) 1989-05-09
NO891892L (no) 1989-05-09
FI892489L (fi) 1989-05-23
JPH02501349A (ja) 1990-05-17
SE460472B (sv) 1989-10-16
SE8601511D0 (sv) 1986-04-04
FI892489A7 (fi) 1989-05-23
SE8601511L (sv) 1987-10-05

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