US3853711A - Installation for automation of microbiological work techniques - Google Patents

Installation for automation of microbiological work techniques Download PDF

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Publication number
US3853711A
US3853711A US00153286A US15328671A US3853711A US 3853711 A US3853711 A US 3853711A US 00153286 A US00153286 A US 00153286A US 15328671 A US15328671 A US 15328671A US 3853711 A US3853711 A US 3853711A
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invention according
gel
strips
strip
glass
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US00153286A
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English (en)
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C Heden
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Biotec AB
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Biotec AB
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Priority claimed from SE8592/70A external-priority patent/SE344368B/xx
Priority claimed from SE7107264A external-priority patent/SE374433B/xx
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/02—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor using a plurality of sample containers moved by a conveyor system past one or more treatment or analysis stations
    • G01N35/028—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor using a plurality of sample containers moved by a conveyor system past one or more treatment or analysis stations having reaction cells in the form of microtitration plates

Definitions

  • the present invention relates to a method for automating microbiological analysis of samples on gel substrates and to an installation for the automation of microbiological work technique.
  • the method is mainly characterized by separately putting each substrate into a protective tube or cassette after the addition of active substance and/or sample, whereby the substrate is so shaped that diffusion, growth or other spreading in the substrate essentially occurs in the longitudinal direction of said substrate, and that the environmental conditions in the protective tubes are individually selected for each tube.
  • the installation is mainly characterized in that it constitutes an interconnection of two or more of the following components as modules in the modifiable installation: a cutter, an incubating station, a thermostatic chamber, a refrigerated storage chamber and a reading station.
  • Such an installation for use in the hospital most often consists of a chemical central processing unit which performs the actual analytical work, an electronic control unit which precisely determines all the work stages, an amplifier which transmits the results as electric impulses to a computer, and an electric teleprinter equipped with punched tape.
  • a chemical central processing unit which performs the actual analytical work
  • an electronic control unit which precisely determines all the work stages
  • an amplifier which transmits the results as electric impulses to a computer
  • an electric teleprinter equipped with punched tape included in the central unit is a system for mechanically conveying samples and reaction solutions, pneumatic control of the table movement and the pipettes, reactant liquid supply, heating and cooling and colorimetric measuring of the mechanism of reaction, apparatus ventilation and washing of test tubes, pipettes and measuring heads.
  • One model of the chemical central unit has 24 solid analysis channels and a maximum capacity of about 135 samples per hour, and a channel which makes possible at least 3,000 analyses per hour. The measuring occurs in a photometer in principle a 2-way photocell colorimeter having
  • the present invention relates to a method for automating microbiological analysis of samples on gel substrates and is characterized in that each substrate is separately introduced into a protective tube after an active substance and/or sample has been added.
  • the substrate is so shaped that diffusion, growth or other spreading therein basically occurs in its longitudinal direction.
  • the environmental conditions in the protective tubes can be individually selected in each tube.
  • the invention also relates to an installation for automation of microbiological work technique, and consists of an interconnection ofa cutter for making agar strips, a machine for making spirals which transmit small amounts of liquid, an incubating station, a thermostatic chamber, a refrigerated storage chamber of agar strips and a reading station which possibly includes a computer and recorder.
  • FIG. 1 is a block diagram of an embodiment of an installation according to the invention.
  • FIG. 2 shows protective tubes containing samples, and means for supplying a special atmosphere.
  • FIG. 3 is a cross section along the line 3-3 in FIG. 2.
  • FIG. 4 shows means for feeding different gel substrates
  • FIG. 5 shows a cutting machine for making gel layers from gel blocks.
  • the basic unit in the automation technique consists of long, narrow strips or shallow troughs of glass which are 2 X 50 cm and have a thin substrate layer l-2 mm thick.
  • the glass strips are kept in sterile cassettes or tubes which can be provided with individual gas milieus, e.g., N air, 10% CO in air, etc.
  • a module system has been worked out to extend from there. Said system is designed to perform the following functions:
  • A. Distribution which aims at producing separate colonies from the micro-organisms in a mixture. From here, for example, the bacteria in a clinical sample can be isolated; micro-organisms in a fluid impinger used for testing the air can be isolated; and mutants from an irradiated, or an otherwise genetically manipulated population, can be isolated.
  • the substrate strip consisting of a gel hereafter called agar" is injected into an insulating tube, and the material from a testing wire is spread out along the middle line of said strip.
  • D Quantitative microbiological analysis with the help of bacteria embedded in the agar whose purpose is to determine, by a measurement of the size of the growth zone or the inhibition zone, unknown substances concentration in solutions with the effect on selected micro-organisms.
  • This type of standardized diffusion analysis can be used for the quantitative determination of amino acids, vitamins and antibiotics in solutions.
  • the procedure takes place as follows: Agar strips containing a suitable concentration of sample organisms are gradually fed into an insulating tube at the same time as small liquid samples sucked up in a suitable carrier (spiral) are applied.
  • auxoautotrophic bacterial strains i.e., micro-organisms are used, the growth of which is dependent on the substance to be determined. This is of course excluded from the agar used.
  • micro-organisms which produce microbiologically active factors the purpose of which is to select the colonies from a mixture of micro organisms which are capable of producing a certain active factor.
  • micro-organisms can be obtained which are capable of producing amino acids or antibiotics.
  • agar strips containing sample organisms in a suitable medium are injected into an insulating tube and the material with microorganisms from a testing wire is spread along the middle line of the strip.
  • a sample consisting of microorganisms is added to the substrate, as is also the case in paragraph B.
  • paragraphs C and D a certain microorganism is spread over the surface of the agar medium or embedded within the medium, and the sample consists of some microbiologically active substance which is added.
  • paragraph E one microorganism is embedded in the substrate, and another microorganism is added as sample.
  • the sample which is added to initiate one of the various microbiological reactions to be studied by the present method and installation, is hereafter generally referred to as microbiologically active substance.
  • this expression is directed towards a culture of a microorganism or mixture of microorganisms, as well as a solution of some microbiologically active substance.
  • FIG. I shows an embodiment of an installation according to the invention consisting of a. a cutter for making agar strips from agar units which simultaneously lays said agar strips on the glass strips,
  • a reading station possibly including manipulative operations, dependent on the type of analysis.
  • the cutting machine according to the invention is shown in greater detail on FIG. 5.
  • the agar block 40 is embedded in a glass cassette 41 and is then laid on the base 48 of the cutting machine; the feeding belt from the piston 45 is coupled to the feeding mechanism 44.
  • the desired strip thickness for the agar is set and the cutting machine is turned on.
  • the agar block is then fed forward an amount equal to the desired thickness and a knife 42, operated by a drive mechanism 43, cuts off an agar sliver 47 which falls down onto the glass strip 46.
  • the sample is transmitted with the help of the spiral or thread from the cutter to the agar strips.
  • Said agar strips are then fed into insulating tubes which have a rubber membrane on the bottom.
  • the insulating tubes are fed lying laterally onto the conveyor and are then moved in their longitudinal direction a few mm to the side of a guide rail.
  • the lid which remains in the conveying direction thereby becomes free of the tube.
  • the tube is then swung by another rail up 45, and the glass strip with the agar strip is allowed to slide into the insulating tube which then swings back. If the cultivation is to take place in a normal atmosphere, the tube is laterally moved back so that the lid is automatically put on.
  • the tube is instead moved further to the side a few cm, and away from the lid. In this way a cannula penetrates through the rubber bottom of the insulating tube.
  • the desired gas e.g,, nitrogen, carbon dioxide, etc.
  • the desired gas e.g, nitrogen, carbon dioxide, etc.
  • the tube is laterally displaced back. With the return of the insulating tube, the cannula is pulled out, the bottom seals and the lid is reset automatically.
  • the tube becomes axially fixed, and instead the lid and cannula are maneuvered e.g., hydraulically, pneumatically or electrically.
  • FIG. 2 illustrates the supply of controlled gas with a movable cannula 31 operated by a hydraulic cylinder 37.
  • the gas is fed to the cannula through a pipe 30, and the gas supply after the penetration of the cannula 31 into the membrane 32 is controlled by a controlling device 38.
  • the insulating tube 33 is fixed in tube brackets 34.
  • FIG. 3 is a section illustrating how the layer of agar 36 on the glass strip 35 is placed in the insulating tube 33.
  • the cassettes or tubes are incubated in the thermostatic chamber during the continuous feed through operation.
  • the capacity is adjustable, and with a feeding rate of one cassette per minute, the installation has a capacity of 1,500 samples per 24 hours.
  • the preincubated agar strips are kept in the refrigerated storage chamber.
  • the reading of the samples is finally taken in the reading station which possibly has optional work operations.
  • the values are then transmitted to the computer to be registered; the net values are then assessed for final feeding as completed analysis results.
  • Supply and circulation lines are also included in the installation.
  • a chamber for rustproof wire for feeding said wire to the spiral machine and a washing station for cassettes from the reading station. Said cassettes are then fed to a chamber for renewed use. After the reading station, the agar strips are further scraped off from the glass strips. Said glass strips are washed, move on to a chamber and are then fed anew into the cutter.
  • the free content of seven different amino acids in the urine has been determined. Furthermore, eight different B-vitamins have been determined by using the same basic medium as for the amino acids. This method of analysis has shown itself to be so sensitive that the percentage of free biotine in rat plasma could be determined to l nanogram/ml.
  • test tube racks are preferably in the form of a plastics strip with indented cavities having a capacity of 0.1 or 1 ml, even if the size of the tubes makes it possible to use significantly larger cavities.
  • each cavity has a capacity of 0.1 ml, there will be room for 99 cavities in each rack, a number which allows for a large series of tests in a quite limited space.
  • Separate colonies can e.g., first be isolated from the sample by cultivation on agar strips. Material fromthe colonies can then be easily transferred to liquid substrate portions in the test tube rack cavities which are then sealed with tape or, in the alternative, rice paper or fibre glass paper. A similar technique may be used in genetic selection work.
  • filter paper slips may be used which have been impregnated with the liquid to be supplied to the gel.
  • the slips could be secured on an adhesive tape with only surface contact, where the sticky surface is of such nature that it yields neither vapors nor fluids.
  • the sticky surface is preferably made ofa known per se, uniform, solid, sticky substance such as a high-molecular, sticky polymer.
  • the slips may be punched out of prepared filter paper and either remain stored in the hollow punchers or brought into contact with adhesive tape in special tubes.
  • tubes with slips containing separate preparations are set up beside each other at a distance of about 15-30 mm with the opening facing up.
  • the slips are forced up against the opening with a spring while they are prevented from being squeezed out by a tongue or guard in the mouth of the tube.
  • a strip of tape is placed above the mouth of the tube, the slips stick when contacted with the sticky surface and the top slip in each tube is lifted up and is fixed onto the strip.
  • the next portion of the tape can then be similarly handled, etc.
  • the tape is manually rolled up into a roll containing a large number of batches of prepared slips protected from the air. The tape may then be stored for future use.
  • the tape strip can e.g., be perforated with holes corresponding in number to the serum samples which are suspected to contain a certain antigen (e.g., Au in hepatitus).
  • a certain antigen e.g., Au in hepatitus
  • On the sides of each hole are a slip prepared with antibodies (Anti Au) and a slip containing antigen (-l-Au).
  • Anti Au antibodies
  • -l-Au a slip prepared with antibodies
  • -l-Au a slip containing antigen
  • the tape with the sticky surface and the slips are laid down on an agar strip made e.g., as described above. This is done either manually or automatically. Incubation then occurs in a normal manner in a thermostat cabinet for a suitable period of time, e.g., 24 hours.
  • the method is not confined in principle to these loose slips but, according to another embodiment, narrow strips of filter paper magazined in rolls are prepared instead. These strips are then pulled out parallel to each other from the rolls at a distance from each other of 15-30 mm. The tape is placed perpendicular to the direction of the strips and the latter are then lopped off in line with the tape edge.
  • the method may also be modified in numerous other ways, e.g., with a wide tape which is lopped off together with the strips to a narrower width by supplying the small round filter paper slips with automatic applications, etc.
  • Registration and evaluation of the samples then take place in a normal manner by measuring the stain formed around each slip where the growth is hindered or stimulated, respectively, or by studying the position and dispersion of the precipitate which may occur with slips impregnated with different antigens and antibodies.
  • the tape covers the agar, as a rule enough oxygen penetrates, due to the agar being strip-like, so that even aerobic bacteria will be able to grow.
  • oxygen diffusion can be further facilitated by making the surface of the agar layer serrated. This is most easily done when the cutting in the cutter is effected with a thin wire or edge which, during the actual slicing operation, is made to oscillate in a known per se manner by mechanically, electrically or otherwise produced vibrations.
  • bands or wires prepared with various substances are rolled up on spools mounted beside each other on a common shaft.
  • the length of the shaft corresponds approximately to that of the agar strip.
  • the narrow bands or wires are then simultaneously pulled out right across the agar strip and are then cut on both sides of said strip to lie in intimate contact with the surface.
  • a number of various agar types can be arranged in a magazine, a certain arbitrary number of which can be coupled in.
  • the agar block is propelled forwards a bit, corresponding to the desired thickness of the preparation, and is then sliced with transverse knife.
  • FIG. 4 shows an arrangement where there is a row 1 of spools with rolled-up bands or wires 2 on a shaft.
  • the row of spools lies with its axis parallel to the agar strip and the bands are pulled out directly over said strip by means of a grip.
  • the bands or wires are then cut off by a cutter which may consist of either long knives being of the same length as the agar strip or slitting knives or the like which run on a guide along the side edge of the agar strip.
  • the row 1 of spools is placed above a stack with six cassettes 3-8 containing different types of agar.
  • a suitable length of agar is pushed out from the desired cassette by e.g., a hydraulic cylinder, and a slice is cut off with a knife 9.
  • the slice is laid on a glass strip which rests on a hoist 10.
  • the glass strip is held in a magazine 12 and is fed from there on a track 11 to the hoist 10 which is controlled by e.g., a hydraulic cylinder 13 and is provided with a driver 14 that, at a determined height, may either feed or extract agar strips to or from protective tubes 15.
  • the arrangement is designed to keep agar strips without protective tubes in sterile, well moistened air. But, in certain cases, it may be necessary to have a protective gas, a special atmosphere, a temperature other than room temperature, etc. Therefore, the agar strips, at this stage, may even be fed out for e.g., treatment with trichloracetic acid, reaction with fluorescent antibodies or to storage at 37C in special protective tubes.
  • Procedures are taken hereunder for the substances, with which the band is prepared, to diffuse out into the agar.
  • the strip is transferred by a driver 18 to a downwards moving conveyor belt where inoculation is carried out by spreading the bacteria strains and the like, present here, as a film over the agar surface.
  • the bands set on can be removed.
  • the strip is fed up a certain height to an upwards moving conveyor chain where it finally reaches the level 20.
  • a carrier 21 is fed by means of a carrier 21 to a table 22 where it may either be carried to the left for optical reading at the arrow 23, or to the right for treatment with trichloroacetic acid in a protective tube before continuing on to optical reading.
  • Apparatus for automatic performance of microbiological analyses of samples applied to a gel substrate layer comprising a gel substrate preparation station, an inoculating station, an incubating station, a reading station capable of detecting microbiol growth, and conveyor means, said gel substrate preparation station comprising a solid block of said gel having a generally uniform elongated rectangular transverse cross-section with respect to at least one axis, cutter means for slicing successive transverse thin strips of gel from said block, a supply of similar glass strips; each having an area sufficient to provide a support for at least one of said thin sliced strips of gel, said conveyor means including means to relatively position glass strips with respect to sliced strips of gel for deposit of the slices of gel on successive glass strips at a first position and to move said glass strips in a sequential manner to other said stations after receiving a slice of gel.
  • said block of gel comprises a rectangular slab the dimensions in cross-section in one plane having a ratio greater than l: 1 and said apparatus includes means to move said slab towards said cutter means.
  • said plurality of stations includes a station for inoculating the gel strip prior to a station for inserting a glass strip and inoculated gel layer into a protective tube.
  • one of said plurality of stations includes means to introduce a gas into the interior of a protective tube containing an inoculated gel layer.
  • said protective tube includes an rubber membrane and said means to introduce said gas includes a cannula for penetrating said membrane.
  • said conveyor means includes two adjacent sections, each of said sections being movable in opposite directions, and transfer means to shift a glass strip between one section and the other section to move a glass strip between two of said stations.
  • said conveyor means includes a plurality of endless chain means disposed generally in longitudinal alignment and said plurality of stations are disposed generally at locations in planes spaced from each other in the direction of said alignment.
  • said apparatus also includes means to store a plurality of gel blocks generally in the form of rectangular thin slabs in stacked superposed relation to each other, means to selectively position a glass strip adjacent the end of any one of said slabs and cutter means to'slice a strip from said one end of a slab for placement on said strip.
  • said apparatus includes means to supply a plurality of microbiologically active substances to slice of gel'on a glass strip at one end of said plurality of stations.
  • said means to supply a plurality of microbiologically active substances includes a wire coil, the active substance being contained within said coil, and means to sever the wire coil in a length proportional to the amount of substance desired.
  • said means to supply a plurality of microbiologically active substances includes a porous band having a substance contained therein.
  • said means to supply a plurality of microbiologically active substances includes adhesive tape, and filter paper adhered to said tape, the active substance being contained by said filter paper.
  • said station for the receipt of a microbiologically active substance includes means to supply a plurality of said substances, said means to supply a plurality of active substances including a horizontally aligned group of individual supplies of said substances to be dispensed to various locations along the length of a slice of gel.
  • said means to supply said microbiologically active substances includes a plurality of revoluble spools mounted in axial alignment on an axis parallel to the length of said slice of gel.
  • said apparatus includes means to store a plurality of gel blocks generally in the form of rectangular thin slabs, conveyor means to support a glass strip adjacent one end of any one of said slabs, and cutter means to slice off a thin strip of said one end of said slab for placement on said glass strip.
  • said apparatus also includes means to store a plurality of said glass strips and means to successively position said strips to receive a thin strip of gel thereon.
  • said conveyor means includes a second two series of vertically spaced elements movable concurrently in parallel paths adjacent the paths of said first mentioned two series of elements, and transfer means to move glass strips horizontally between said first and second two series of elements.

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  • Chemical & Material Sciences (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Physics & Mathematics (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Pathology (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
  • Investigating Or Analysing Biological Materials (AREA)
US00153286A 1970-06-22 1971-06-15 Installation for automation of microbiological work techniques Expired - Lifetime US3853711A (en)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
SE8592/70A SE344368B (fr) 1970-06-22 1970-06-22
SE7107264A SE374433B (fr) 1971-06-04 1971-06-04

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Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4160699A (en) * 1976-11-09 1979-07-10 Olympus Optical Co., Ltd. Drive system for automatic culture apparatus
US4219529A (en) * 1977-11-28 1980-08-26 Eastman Kodak Company Incubator for chemical analyzer
EP0539888A1 (fr) * 1991-10-30 1993-05-05 Shimadzu Corporation Dispositif pour la sélection des cellules et similaires
US5403735A (en) * 1990-07-25 1995-04-04 Hitachi, Ltd. Method and apparatus for investigating and controlling an object
US6534017B1 (en) * 1997-04-11 2003-03-18 Roche Diagnostics Gmbh Test element storage device

Citations (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3065150A (en) * 1960-11-21 1962-11-20 Kravitz Harvey Bacteriologic culture apparatus
US3205151A (en) * 1962-04-17 1965-09-07 Hollister Inc Inoculation device and method
US3416998A (en) * 1967-10-11 1968-12-17 Research Corp Method of detecting or classifying microorganisms using agar reagent sheets
US3616264A (en) * 1969-06-30 1971-10-26 Beckman Instruments Inc Temperature-controlled discrete sample analyzer
US3661718A (en) * 1970-01-26 1972-05-09 Birko Chemical Corp Method for taking and transferring bacteria samples
US3728277A (en) * 1970-01-12 1973-04-17 Witco Chemical Corp Stable water-in-oil emulsions
US3772154A (en) * 1971-05-03 1973-11-13 Technicon Instr Method and apparatus for automated antibiotic susceptibility analysis of bacteria samples

Patent Citations (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3065150A (en) * 1960-11-21 1962-11-20 Kravitz Harvey Bacteriologic culture apparatus
US3205151A (en) * 1962-04-17 1965-09-07 Hollister Inc Inoculation device and method
US3416998A (en) * 1967-10-11 1968-12-17 Research Corp Method of detecting or classifying microorganisms using agar reagent sheets
US3616264A (en) * 1969-06-30 1971-10-26 Beckman Instruments Inc Temperature-controlled discrete sample analyzer
US3728277A (en) * 1970-01-12 1973-04-17 Witco Chemical Corp Stable water-in-oil emulsions
US3661718A (en) * 1970-01-26 1972-05-09 Birko Chemical Corp Method for taking and transferring bacteria samples
US3772154A (en) * 1971-05-03 1973-11-13 Technicon Instr Method and apparatus for automated antibiotic susceptibility analysis of bacteria samples

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
L.S. Gall, Partially Automated System for Microbiological Analysis, Dev. in Industrial Microbiology, Vol. 11, pp. 460 469. *
Oxoid Ltd., Food Engineering, August, 1967, p. 138. *

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4160699A (en) * 1976-11-09 1979-07-10 Olympus Optical Co., Ltd. Drive system for automatic culture apparatus
US4219529A (en) * 1977-11-28 1980-08-26 Eastman Kodak Company Incubator for chemical analyzer
US5403735A (en) * 1990-07-25 1995-04-04 Hitachi, Ltd. Method and apparatus for investigating and controlling an object
EP0539888A1 (fr) * 1991-10-30 1993-05-05 Shimadzu Corporation Dispositif pour la sélection des cellules et similaires
US5348883A (en) * 1991-10-30 1994-09-20 Shimadzu Corporation Selecting device for cells and the like
US6534017B1 (en) * 1997-04-11 2003-03-18 Roche Diagnostics Gmbh Test element storage device

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DE2130705A1 (de) 1971-12-23
JPS535397B1 (fr) 1978-02-27
DE2130705B2 (de) 1976-08-19
GB1360622A (en) 1974-07-17

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