EP0891555A1 - Dispositif servant a compter de petites particules et appareil de tri comprenant ce dispositif - Google Patents

Dispositif servant a compter de petites particules et appareil de tri comprenant ce dispositif

Info

Publication number
EP0891555A1
EP0891555A1 EP97915420A EP97915420A EP0891555A1 EP 0891555 A1 EP0891555 A1 EP 0891555A1 EP 97915420 A EP97915420 A EP 97915420A EP 97915420 A EP97915420 A EP 97915420A EP 0891555 A1 EP0891555 A1 EP 0891555A1
Authority
EP
European Patent Office
Prior art keywords
light
capillary
optical waveguide
delivering
measuring light
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Withdrawn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
EP97915420A
Other languages
German (de)
English (en)
Inventor
Martin Hassler
Harald Danigel
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.)
Novartis AG
Original Assignee
Novartis AG
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 Novartis AG filed Critical Novartis AG
Priority to EP97915420A priority Critical patent/EP0891555A1/fr
Publication of EP0891555A1 publication Critical patent/EP0891555A1/fr
Withdrawn legal-status Critical Current

Links

Classifications

    • 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
    • G01N35/10—Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices
    • G01N35/1081—Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices characterised by the means for relatively moving the transfer device and the containers in an horizontal plane
    • G01N35/109—Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices characterised by the means for relatively moving the transfer device and the containers in an horizontal plane with two horizontal degrees of freedom
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J19/00—Chemical, physical or physico-chemical processes in general; Their relevant apparatus
    • B01J19/0046—Sequential or parallel reactions, e.g. for the synthesis of polypeptides or polynucleotides; Apparatus and devices for combinatorial chemistry or for making molecular arrays
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume, or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/01—Arrangements or apparatus for facilitating the optical investigation
    • G01N21/03—Cuvette constructions
    • G01N21/05—Flow-through cuvettes
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2219/00—Chemical, physical or physico-chemical processes in general; Their relevant apparatus
    • B01J2219/00274—Sequential or parallel reactions; Apparatus and devices for combinatorial chemistry or for making arrays; Chemical library technology
    • B01J2219/00277—Apparatus
    • B01J2219/00279—Features relating to reactor vessels
    • B01J2219/00306—Reactor vessels in a multiple arrangement
    • B01J2219/00313—Reactor vessels in a multiple arrangement the reactor vessels being formed by arrays of wells in blocks
    • B01J2219/00315—Microtiter plates
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2219/00—Chemical, physical or physico-chemical processes in general; Their relevant apparatus
    • B01J2219/00274—Sequential or parallel reactions; Apparatus and devices for combinatorial chemistry or for making arrays; Chemical library technology
    • B01J2219/00277—Apparatus
    • B01J2219/00457—Dispensing or evacuation of the solid phase support
    • B01J2219/00459—Beads
    • B01J2219/00468—Beads by manipulation of individual beads
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2219/00—Chemical, physical or physico-chemical processes in general; Their relevant apparatus
    • B01J2219/00274—Sequential or parallel reactions; Apparatus and devices for combinatorial chemistry or for making arrays; Chemical library technology
    • B01J2219/00277—Apparatus
    • B01J2219/00497—Features relating to the solid phase supports
    • B01J2219/005—Beads
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2219/00—Chemical, physical or physico-chemical processes in general; Their relevant apparatus
    • B01J2219/00274—Sequential or parallel reactions; Apparatus and devices for combinatorial chemistry or for making arrays; Chemical library technology
    • B01J2219/00583—Features relative to the processes being carried out
    • B01J2219/00603—Making arrays on substantially continuous surfaces
    • B01J2219/00659—Two-dimensional arrays
    • C—CHEMISTRY; METALLURGY
    • C40—COMBINATORIAL TECHNOLOGY
    • C40B—COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES
    • C40B60/00—Apparatus specially adapted for use in combinatorial chemistry or with libraries
    • C40B60/14—Apparatus specially adapted for use in combinatorial chemistry or with libraries for creating libraries
    • G01N15/149—
    • G01N2015/1024—
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume, or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Electro-optical investigation, e.g. flow cytometers
    • G01N2015/1486—Counting the particles
    • 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
    • G01N2035/00465—Separating and mixing arrangements
    • G01N2035/00564—Handling or washing solid phase elements, e.g. beads

Definitions

  • Device for counting small particles and a sorting apparatus comprising such a device
  • the invention relates to a device for counting small particles according to the preamble of independent claim 1.
  • the invention relates especially to a device for counting carriers (beads) as are used in modern research in the synthesis of novel chemical substances.
  • the invention relates also to a sorting apparatus according to the preamble of claim 14 comprising such a device.
  • novel chemical substances is often guided by the objective of synthesizing a substance that has given physical, chemical or bioactive properties.
  • Such chemical substances may be, for example, plastics, colorants, aromatics, flavourings, bioactive materials or other compounds having useful properties.
  • Bioactive materials are, for example, pharmaceutical or therapeutic active ingredients, herbicides, pesticides, fungicides or active ingredients used in veterinary medicine.
  • combinatory technologies have often been used in the search for such novel chemical substances. These are based on synthesizing a very large number of different chemical compounds by adding together a predetermined number of reagents that serve as "building blocks" in combination to form one substance.
  • the "building blocks” may be amino acids, with which polypeptides of a predetermined length, that is to say a predetermined number of amino acids in a polypeptide, are synthesized.
  • a vastly large number of such polypeptides having different, predetermined lengths can be synthesized.
  • a method frequently used in combinatory technology is the synthesis of substances on carriers.
  • the carriers serve as foundations on which the "building blocks" are added together to form the novel substance.
  • the carriers are small, mostly spherical particles having a diameter of customarily less than 200 ⁇ m and are made, for example, of resin or polystyrene.
  • the carriers are hereinafter referred to as "beads".
  • the beads For that reason, a large number of up to several ten thousand beads having on them the generally different substances are customarily placed together in a vessel, for example on a microtitre plate, and are there studied for the desired property or action by means of an indicator medium. If one of the substances on the beads is then found to be active or to have the desired property, the beads must be divided into smaller portions, for example into the various vessels of a further microtitre plate or deep-well plate, and the detection experiment must be repeated. In analogous manner, the beads are divided into smaller and smaller portions. The purpose of that successive procedure is to isolate or detect the bead on which the active substance or substance having the desired property is located. Towards the end of that successive process, the vessel containing the beads to be distributed then contains only a very few beads, for example approximately one hundred beads, which are to be distributed into, for example, one hundred vessels, so that finally only one bead is present in each vessel.
  • the beads located in one vessel into a plurality of vessels, for example into the individual positions of a microtitre plate or deep-well plate.
  • the beads are first suspended in a solvent to form a suspension and that suspension is then distributed.
  • the distribution from one vessel into a plurality of other vessels is customarily carried out using syringes or pipettes.
  • Devices are also known in which the principle of a water jet pump is used to divide the beads, the beads being sucked by a conditioned jet of a solvent from a reservoir into the jet. The resulting suspension is then separated into droplets which are then distributed into the various vessels.
  • a disadvantage of such and similar apparatuses is, however, that the beads must first be transferred from the vessel containing them into a reservoir. Individual beads may be lost in the process, the risk being that the very bead that is lost is one that carries an active substance.
  • the known devices prove to be highly problematic especially in cases in which a very small number of beads is to be divided into still smaller portions, where each portion is to contain only from one bead to a few dozen beads. With traditional pipettes or syringes it is possible to determine only the volume of distributed suspension, or the droplets of suspension introduced can be counted. The disadvantage here is that the exact number of distributed beads is unknown.
  • a problem underlying the invention is therefore to provide a device for counting small particles, especially beads. With the device it is to be possible to count beads in the form of a suspension as far as possible without error. The loss of individual beads in the course of the counting process is to be avoided.
  • a further problem underlying the invention is to provide a sorting apparatus with which small particles, for example beads in the form of a suspension, can be removed from one vessel and distributed into other vessels, it being possible to control and determine the number of particles introduced into a vessel. With such a sorting apparatus, it is to be possible, especially, to divide small amounts of, for example, up to a few hundred particles into still smaller portions of from one particle to a few dozen such particles each.
  • the apparatus is also to enable a portion of beads to be distributed, in a controlled manner, into the various positions of a microtitre plate or deep- well plate so that, for example, there is subsequently a maximum of one bead in each position. Furthermore, with such a sorting apparatus, the loss of particles or beads is to be avoided as far as possible.
  • Fig. 1 shows a longitudinal section through a first embodiment of the device according to the invention for counting small particles
  • Fig. 2 shows a cross-section through a second embodiment of the device according to the invention
  • Fig. 3 is a diagram showing signal intensities of the measuring light as a function of the wavelength of the measuring light
  • Fig. 4 shows a schematic representation of an embodiment of the sorting apparatus according to the invention showing the essential parts.
  • the small particles to be counted or sorted are, by way of example, beads in the form of a suspension.
  • the term "bead” denotes small carriers that, typically, are essentially spherical in shape and have a diameter of customarily up to 200 ⁇ m. They are made, for example, of resin or polystyrene.
  • they are often used as solid bodies on which the chemical substances to be synthesized are built up successively from reagents that serve as building blocks.
  • Such beads which are often used in solid phase synthesis, are sufficiently well known and therefore require no further explanation.
  • the invention is not limited to applications in which beads are counted or sorted.
  • the particles to be counted or sorted may equally well be other small particles, for example small particles that are not spherical in shape. They may be, for example, relatively large colloids or other small-volume physical structures.
  • the device according to the present invention is also suitable for counting or sorting animal varieties for example small insects, their eggs or larvae, especially in high-throughput- screen (HTS) systems.
  • animal varieties for example small insects, their eggs or larvae
  • HTS high-throughput- screen
  • plants or plant parts are infested with a predetermined number of such animal varieties. These are then treated with a bioactive compound and the resulting effect is determined by comparision with an untreated standard.
  • the device according to the invention for counting small particles, especially carriers (beads), is based on an optical method. It comprises, typically, a capillary 2 (Fig. 1) for receiving the particles to be counted, a light source 3 for emitting measuring light, which is indicated symbolically in Fig. 1 by the arrows denoted by the reference character M, and also light-delivering means which deliver the measuring light M emitted by the light source 3 to the capillary 2, a photoelectric detector 5 for receiving the measuring light M, and light- removing means which deliver the measuring light M coming from the capillary 2 to the photoelectric detector 5.
  • a capillary 2 Fig. 1
  • a light source 3 for emitting measuring light, which is indicated symbolically in Fig. 1 by the arrows denoted by the reference character M
  • light-delivering means which deliver the measuring light M emitted by the light source 3 to the capillary 2
  • a photoelectric detector 5 for receiving the measuring light M
  • light- removing means which deliver the measuring light M coming from
  • the intemal diameter ID of the capillary 2 is such that the individual particles can enter the capillary 2 only one behind the other and can move along inside it only one behind the other.
  • the light-delivering means irradiate the capillary 2 with the measuring light M transversely to the longitudinal axis thereof, the measuring light M being in the form of a light beam, the width of which is such that only one of the particles can be completely in the light beam at any one time.
  • the light beam is indicated symbolically in the region of the capillary 2 by the two rays M1 and M2.
  • capillary is used to mean generally tube-like structures, the cross-section of which is small in comparison with their longitudinal extent, and comprises, for example, also pipettes, micropipettes and syringes.
  • the device according to the invention operates as follows: the measuring light M emitted by the light source 3 passes through the light-delivering means to the capillary 2 and irradiates the capillary 2 with a light beam transversely to the longitudinal direction of the capillary 2. After passing through the capillary, the measuring light M passes through the light-removing means to the photoelectric detector which converts the measuring light M into an electrical signal which is available for further processing and evaluation. If the capillary 2 is then immersed in a container 20 that contains, for example, a suspension 21 of a solvent and beads 22, a drop of the suspension 21 moves into the capillary 2 and moves along inside it. The drop may contain no beads, one bead or a plurality of beads.
  • the internal diameter of the capillary 2 is such that the beads can move along in the capillary 2 only one behind the other but not next to one another, and furthermore, since the width of the light beam passing through the capillary 2 is such that only one of the beads can be completely in the light beam at any one time, the beads must cross the light beam one after the other.
  • a bead crosses the light beam, changes occur in the measuring light M received by the photoelectric detector. By reference to those changes, it is possible to count the individual beads that may be present in the drop.
  • Fig. 1 shows a longitudinal section of a first preferred embodiment of the device according to the invention for counting small particles, especially beads. That Figure shows the device in its normal position of use. Terms such as “upper” and “lower” used hereinafter relate to that position of use.
  • the device according to the invention is denoted as a whole by the reference numeral 1.
  • the capillary 2 is in the form of a hollow cylinder having a longitudinal axis A and an intemal diameter ID.
  • the light-delivering means preferably comprise a delivering optical waveguide 4 which has a casing 41 and also at least one optical fibre 42, for example a glass fibre.
  • the light-removing means preferably comprise a removing optical waveguide 6 which has a casing 61 and also at least one optical fibre 62, for example a glass fibre.
  • the arrangement of the light-delivering and light-removing means with the optical waveguides 4, 6 is preferred because a simple optical connection for guiding the measuring light M between the light source 3 and the capillary 2, and between the capillary 2 and the photoelectric detector 5, can thus be obtained.
  • the capillary 2, with the optical waveguides 4, 6 fixed in relation thereto can be moved independently of the light source 3 and the photoelectric detector 5 without thereby impairing the propagation of the measuring light M along the light path from the light source 3 through the capillary 2 to the detector 5.
  • the delivering optical waveguide 4 and the removing optical waveguide 6 are arranged - as shown in Fig. 1 - parallel to the longitudinal axis A of the capillary 2.
  • a first reflecting surface 10 which is so arranged that it deflects the emerging measuring light M towards the capillary 2.
  • a second reflecting surface 11 is provided in the region of the light-entry surface 63 of the removing optical waveguide 6 a second reflecting surface 11 which is so arranged that it deflects the measuring light M emerging from the capillary 2 into the removing optical waveguide 6.
  • the representation of the light source 3 and of the photoelectric detector 5 in Fig. 1 is to be understood as being symbolic.
  • the light source 3 may comprise, for example, a deuterium lamp, a light-emitting diode or a laser diode.
  • the photoelectric detector may comprise, for example, a photoelement, a photocell, a photomultiplier, a photodiode or a diode array.
  • a sleeve 8 which surrounds the capillary 2 closely is also provided.
  • a carrier element 9 having a cavity which receives the sleeve 8 with the capillary 2 and also the two optical waveguides 4, 6 arranged parallel to the longitudinal axis A of the capillary 2.
  • the cavity is substantially cylindrical in its upper region. In the region of its lower end, the cavity narrows by an angle of approximately 45° and merges into a substantially circular bore, the diameter of which corresponds substantially to the external diameter of the capillary 2.
  • the capillary 2 extends beyond the lower end of the carrier element 9.
  • the sleeve 8 extends approximately to the start of the narrowing of the cavity in the carrier element 9.
  • the two optical waveguides 4, 6 terminate above the lower end of the sleeve 8.
  • the carrier element 9 may be mounted, for example, in the region of its upper end on a holding device (not shown) which allows the carrier element 9 and also, of course, the capillary 2 to be moved up and down.
  • the sleeve 8 is connected securely to the carrier element 9 by way of the enclosed optical waveguides 4, 6. That can be achieved, for example, by filling the remaining free space of the cavity with an adhesive so that the optical waveguides are cast in between the sleeve 8 and the carrier element 9 in a mechanically stable manner.
  • Suitable adhesives are, for example, epoxy- resin-based adhesives.
  • a transparent mounting 7 is fitted into the region of the narrowing portion of the cavity of the carrier element 9.
  • transparent means that the mounting 7 is substantially transparent at least for the wavelength or the wave range of the measuring light M used for the detection.
  • the transparent mounting 7 also extends into the cylindrical region of the cavity as far as the lower ends of the optical waveguides 4, 6 and substantially completely fills the space between the lower ends of the optical waveguides 4, 6, the sleeve 8 or capillary 2 and the wall that bounds the cavity.
  • the portion of the transparent mounting 7 that is in the cylindrical region of the cavity is, as a result, annular in shape and surrounds the sleeve 8 together with the capillary 2 inside it.
  • the portion of the transparent mounting 7 that is in the narrowing region of the cavity is of substantially annular but in this embodiment conically tapering shape and surrounds the capillary 2 directly so that its bounding face that faces the capillary 2 forms both a light-exit window 71 for the measuring light M coming from the delivering optical waveguide 4 and a light-entry window 72 for the measuring light M coming from the capillary.
  • the bounding face, remote from the capillary 2, of the conical portion of the transparent mounting 7 is preferably in the form of a reflecting surface, so that the first and second reflecting surfaces 10, 11 form part of the transparent mounting 7.
  • the reflecting surface can be produced, for example, by applying a metal layer, for example by vacuum metallisation.
  • the inner wall of the carrier element 9 that bounds the narrowing region of the cavity is in the form of a reflecting surface, or the total intemal reflection effect is used.
  • the reflecting surfaces 10, 11 may also be in the form of a curved surface, for example a truncated conical surface and/or a hemispherical surface. As a result, for example, the divergence of the measuring light striking the surfaces 10, 11 can at least be reduced.
  • the transparent mounting 7 can be manufactured, for example, from glass, quartz or sapphire. It forms an optical connection between the optical waveguides 4, 6 and the capillary 2.
  • the lower ends of the optical waveguides 4, 6 are each connected to the transparent mounting 7 in a mechanically stable manner.
  • the connection may be effected, for example, by bonding with a transparent adhesive 13. It is especially advantageous if the adhesive 13 has a refractive index that corresponds substantially to the refractive index of the optical fibres 42, 62 or to the refractive index of the transparent mounting 7. As a result, undesirable reflections and scattered light can be reduced.
  • Suitable adhesives are, for example, two-component epoxy-resin-based adhesives.
  • a seal 14 for example an O- ring, between the outer wall of the capillary 2 and the carrier element 9.
  • the seal 14 prevents solvent or portions of the suspension from penetrating past the capillary 2 into the inside of the carrier element 9 as the capillary 2 is immersed in the container 20 and, for example, contaminating the light-exit window 71 or the light-entry window 72.
  • the described arrangement of the first embodiment with the sleeve 8, the carrier element 9 and the transparent mounting 7 has a number of advantages.
  • the form of the device 1 according to the first embodiment is very compact and space-saving.
  • movements or vibrations of the device do not impair the propagation of the measuring light M along the optical light path.
  • the bounding face of the transparent mounting 7 facing the capillary 2, which bounding face contains the light-exit window 71, is curved and thus has the property of at least reducing the divergence with which the measuring light M customarily emerges from the optical fibre 42 of the delivering optical waveguide 4. Furthermore, there is no mechanical connection between the capillary 2 and the transparent mounting 7.
  • the capillary 2 can thus be removed in simple manner from the sleeve 8 that surrounds it closely without it being necessary to manipulate the optical elements of the device for that purpose. That is especially advantageous because, for example in the case of contamination, the capillary 2 can be exchanged for a new capillary rapidly and without great expense. In particular, complicated adjustment of the other parts of the device is not necessary after such an exchange.
  • the upper end of the capillary 2 may be connected by a tube 15 to a syringe or pump device (not shown) in order, on the one hand, actively to suck the suspension out of the container 20 into the capillary 2 or, on the other hand, to eject drops in the capillary in an active and purposeful manner.
  • a syringe or pump device not shown
  • the optical waveguides 4, 6 with their longitudinal direction at an angle, especially approximately at right angles, to the longitudinal axis A of the capillary 2. In that case it is possible to dispense with the reflecting surfaces 10 and 11 because the light-exit surface 43 of the delivering optical waveguide 4 is already so oriented that the measuring light M emerging therefrom irradiates the capillary transversely to the longitudinal axis A thereof.
  • Fig. 2 shows a cross-section of a second embodiment of the device according to the invention.
  • the second embodiment differs from the first essentially in that the light- delivering means and the light-removing means each comprise a plurality of delivering optical waveguides 4a, 4b, 4c and removing optical waveguides 6a, 6b, 6c. If that second embodiment were to be shown in longitudinal section, the drawing would be substantially the same as in Fig. 1.
  • the drawing in Fig. 2 corresponds to a section perpendicular to the longitudinal axis A of the capillary 2 and above the transparent mounting 7.
  • each of the delivering optical waveguides 4a, 4b, 4c and each of the removing optical waveguides 6a, 6b, 6c is arranged parallel to the longitudinal axis A of the capillary 2.
  • the optical waveguides 4a, 4b, 4c, 6a, 6b, 6c are arranged in a ring around the capillary 2 between the outer wall of the sleeve 8 and the wall bounding the cavity in the carrier element 9.
  • the optical waveguides 4a, 4b, 4c, 6a, 6b, 6c all terminate at the same height relative to the capillary 2, as shown, for example, also in Fig. 1.
  • the light-exit surfaces of all the delivering optical waveguides 4a, 4b, 4c and the light-entry surfaces of all the removing optical waveguides 6a, 6b, 6c all lie in one plane, which is substantially perpendicular to the longitudinal axis A of the capillary 2. That arrangement has the advantage that the light path for the measuring light is symmetrical with respect to the longitudinal axis A of the capillary 2. As shown in Fig.
  • the optical waveguides 4a, 4b, 4c, 6a, 6b, 6c are, moreover, so arranged around the capillary 2 that each of the delivering optical waveguides 4a, 4b and 4c and a respective one of the removing optical waveguides 6a, 6b and 6c lie in a pair diametrically opposite one another with respect to the capillary 2.
  • each light-exit surface of one of the delivering optical waveguides 4a, 4b, 4c, with respect to the longitudinal axis A of the capillary 2 there is located one of the light-entry surfaces of the removing optical waveguides 6a, 6b, 6c.
  • Each delivering optical waveguide thus has an associated removing optical waveguide: the removing optical waveguide 6a is associated with the delivering optical waveguide 4a, the removing optical waveguide 6b is associated with the delivering optical waveguide 4b and the removing optical waveguide 6c is associated with the delivering optical waveguide 4c.
  • the delivering optical waveguides 4a, 4b, 4c may all be connected to the same light source 3 (Fig. 1).
  • the light source comprises a beam-splitting device, known perse, that splits the measuring light M into the different delivering optical waveguides 4a, 4b, 4c.
  • a separate light source it is also possible for a separate light source to be associated with each of the delivering optical waveguides 4a, 4b, 4c, in a manner analogous to that shown in Fig. 1 for one optical waveguide.
  • the latter variant with the separate light sources is preferred because a higher intensity of the measuring light M inside the capillary can thus be produced, as a result of which the sensitivity of detection of particles crossing the measuring light beam in the capillary 2 is increased.
  • a plurality of different light sources may be used as separate light sources. Those light sources may differ, for example, in the wavelength or wave range of the measuring light emitted. It is also advantageous, with a view to as great as possible sensitivity, if a separate photoelectric detector is associated with each of the removing optical waveguides 6a, 6b, 6c, in a manner analogous to that shown in Fig. 1 for one removing optical waveguide.
  • the second embodiment has the advantage that the inside of the capillary 2 can be better and more homogeneously illuminated because the delivering optical waveguides 4a, 4b, 4c irradiate the capillary 2 with the measuring light from different sides.
  • the sensitivity of detection and consequently the reliability with which the individual particles are counted can thus be increased further.
  • the number of the delivering and removing optical waveguides 4a, 4b, 4c and 6a, 6b, 6c shown in Fig. 2 (in each case three) is given by way of example. Arrangements are also possible in which the light-delivering means and the light-removing means each comprise only two or, alternatively, more than three optical waveguides.
  • the internal diameter of the capillary 2 is such that the individual particles to be counted can enter the capillary 2 only one behind the other and can move along inside it only one behind the other and moreover, the width of the measuring light beam that irradiates the capillary 2 is such that only one of the particles can be completely in the light beam at any one time. It is especially preferred that those features be obtained as a result of the diameter of each optical waveguide not exceeding the diameter of the capillary 2.
  • the diameter of the optical waveguide is intended to mean the diameter of the light-guiding fibres or, in the case of a plurality of fibres in an optical waveguide, their total diameter.
  • the internal diameter of the capillary 2 is especially less than 300 ⁇ m and more especially less than 160 ⁇ m. That dimension ensures that, for the beads customarily used today, the beads can move along inside the capillary 2 only one behind the other and not side by side.
  • the invention is not limited to the use of optical waveguides. It is also possible for the light-delivering means and light-removing means to comprise different means for guiding the light, for example graduated index lenses (GRIN lenses).
  • GRIN lenses graduated index lenses
  • the light source 3 (Fig. 1 ) emits the measuring light M, which propagates through the delivering optical waveguide 4 and, via the light-exit surface thereof, enters the transparent mounting 7.
  • the measuring light M is then reflected at the first reflecting surface 10, passes through the light-exit window 71 and irradiates the capillary 2 with the light beam transversely to the longitudinal axis A thereof, the light beam being indicated symbolically in Fig. 1 by the light rays M1 and M2.
  • the distance between the rays M1 and M2 indicates the width of the light beam.
  • the divergence of the measuring light M is at least reduced by the curved light-exit window 71.
  • the measuring light After passing through the capillary 2, the measuring light passes through the light-entry window 72 into the transparent mounting 7, is reflected once again at the second reflecting surface 11 and, via the removing optical waveguide 6, reaches the photoelectric detector 5 which converts it into an electrical signal.
  • the signal is supplied to an evaluating unit (not shown) for further processing.
  • the carrier element 9 together with the parts contained therein is then lowered either by motorized means or by hand until the lower end of the capillary 2 comes into contact with the suspension 21 , containing the beads to be counted, in the container 20.
  • a drop of the suspension 21 is then sucked up, for example by means of the syringe or pump device (not shown) which is connected to the tube 15 at the upper end of the capillary 2, and enters the capillary 2, in which it moves upwards.
  • the drop which either consists entirely of solvent or contains at least one bead, enters the measuring light beam, this produces changes in the measuring light M received by the photoelectric detector.
  • That occurrence is shown in Fig. 3. That graph shows the intensity of the measuring light received by the photoelectric detector as a function of the wavelength of the measuring - 15 -
  • the wavelength of the measuring light is plotted in nanometers on the axis labelled ⁇ and the absorption of the measuring light is plotted in units of absorption on the axis labelled AS.
  • the zero point of absorption is set as the absorption measured by the photoelectric detector 5 when there is only solvent and no beads inside the measuring light beam in the capillary 2. That scale is, of course, entirely arbitrary.
  • Ethanol is used as solvent in the graph according to Fig. 3.
  • the curve labelled L shows the absorption of the measuring light measured in the case where air is present in the measuring light beam and the curve labelled B shows the absorption of the measuring light in the case where there is one bead in the measuring light beam.
  • the graph in Fig. 3 shows that, by reference to the changes in the intensity of the measuring light received by the photoelectric detector 5, all the possible interfaces and transitions, namely air/solvent, solvent/bead, bead/solvent and solvent/air can be detected beyond doubt.
  • the graph in Fig. 3 also shows that the differences in the absorption of the measuring light between curve B and curve L are dependent on the wavelength. In the application of the device according to the invention, the wavelengths used for the measuring light will, of course, be those with which the differences are most strongly pronounced.
  • those wavelengths may be in the infrared range, in the visible range or in the ultraviolet range.
  • a very wide wave range is shown in Fig. 3 is purely illustrative. In practical application, it is sufficient to detect the measuring light in a distinctly narrower wave range, or alternatively to use monochromatic measuring light.
  • the choice of suitable wavelength depends on the beads and on the solvent used. In principle, the evaluation can be carried out monochromatically, polychromatically or spectrally (see Fig. 3).
  • the photoelectric detector connected to the evaluating unit (not shown) registers first the air/solvent transition, and thus the "start" of the drop, by reference to the change in the measuring light it receives. If a bead is present in the drop, the solvent/bead transition is registered next. The bead/solvent transition is then registered and it is thus known that a bead is present in the suspension drop. If further beads are present in the drop, next a solvent/bead transition and then again a bead/solvent transition will be registered, and so on. If, however, the "end" of the drop passes through the measuring light beam, then a solvent/air transition is registered.
  • the dimensions of the capillary 2 are such that there is room inside it for two beads only one behind the other and not side by side, and additionally that the width of the measuring light beam is such that only one bead can be completely in the measuring light beam at any one time, each individual bead is registered and, as a result, the beads can thus be counted without error. It is, therefore, not the volume of the suspension that is measured or registered but rather the individual bead directly.
  • the difference in the intensity of the measuring light between the case in which there is a bead in the measuring light beam and the case in which there is only solvent in the measuring light beam is so great that it is also possible to detect when two beads are sticking to one another, that is to say are not fully separated from one another by solvent.
  • a particular advantage of the device according to the invention is, moreover, that it is not limited to the use of a specific solvent, but is suitable for all solvents. Moreover, the device does not require the supply of additional solvent and also does not require an additional reservoir for the beads. This reduces the risk of losing individual beads quite considerably.
  • the present invention relates also to a sorting apparatus for distributing small particles, especially carriers (beads), which comprises a device according to the invention for counting small particles.
  • Sorting apparatuses perse are known from the prior art. For example, in the context of combinatory technology, sorting apparatuses for distributing beads are often used, for which the English term "autosampler" has been adopted.
  • the sorting apparatus according to the invention is improved in that it comprises a device according to the invention for - 17 -
  • Fig. 4 shows an embodiment of the sorting apparatus according to the invention in highly schematic form showing the essential parts.
  • the sorting apparatus as a whole has been given the reference numeral 100 and comprises a counting device of the invention according to the first embodiment described above (see Fig. 1).
  • a counting device of the invention according to the second embodiment described above is, of course, also suitable for integration into a sorting apparatus according to the invention.
  • the embodiment of the sorting apparatus according to the invention shown in Fig. 4 also comprises a base plate 101 and a wall element 102 which is fixed to the side of the base plate 101. Movably attached to the wall surface 102 is an arm 103 which extends over the base plate 101 substantially parallel thereto. The arm 103 is movable along the wall surface 102, as shown symbolically in Fig. 4 by the double arrow X. On the arm 103 there is provided a holding and lifting device 104 which extends from the arm 103 towards the base plate 101. The holding and lifting device 104 is movable along the longitudinal direction of the arm 103, as shown symbolically by the double arrow Y.
  • the direction of movement X of the arm 103 and the direction of movement Y of the holding and lifting device 104 are thus perpendicular to one another.
  • the holding and lifting device 104 can be extended and retracted, for example in telescopic manner, in the direction of the base plate 101. That direction of movement is indicated symbolically in Fig. 4 by the double arrow Z.
  • Drive means (not shown), for example motorized drive means, are provided for moving the arm 103 and the holding and lifting device 104.
  • the drive means have not been shown in Fig. 4.
  • the carrier element 9 of the counting device which carrier element contains the capillary 2, the sleeve 8 (Fig. 1 ), the transparent mounting 7 (Fig. 1 ) and also the ends of the delivering and removing optical waveguides 4 and 6.
  • the delivering optical waveguide 4 is connected to the light source 3 and the removing optical waveguide 6 is connected to the photoelectric detector 5.
  • the end of the capillary 2 remote from the base plate 101 is connected via a tube 106 to a pump device 105 which can produce a subatmospheric pressure or a superatmospheric pressure in the capillary 2.
  • the pump device 105 may, for example, comprise a syringe, a micropump or a different pump.
  • a control and evaluating unit 107 which, on the one hand, controls the drive devices (not shown) for moving the arm 103 and the holding and lifting device 104, as indicated symbolically by a signal line 108, and which, on the other hand, receives signals from the photoelectric detector 5 via a further signal line 109, which signals are processed further and evaluated by the control and evaluating unit 107 to count the beads or the drops.
  • the unit 107 also controls the pump device 105 by way of a connecting line 110.
  • a suspension of beads and a solvent is present in an initial container 120.
  • the beads are to be distributed into various target vessels 122, for example the various positions of a microtitre plate 121 or a deep-well plate, in such a manner that a maximum of one bead is then present in each of the target vessels 122.
  • the initial container 120 and the microtitre plate 121 with the target vessels 122 are placed on the base plate 101.
  • the arm 103 and the holding and lifting device 104 are so moved that the capillary 2 is above the initial container 120.
  • the carrier element 9 is then lowered by means of the holding and lifting device so that the capillary 2 comes into contact with the suspension in the initial container 120 and sucks up a drop from the suspension by means of the pump device 105.
  • the drop rises up the capillary 2 the number of beads contained in the drop is counted, as already described in detail above.
  • the drop does not contain a bead or contains more than one bead, it is ejected back into the initial container 120 by means of a pressure pulse from the pump device 105 and a new drop is then sucked up. If the drop contains precisely one bead, the carrier element 9 with the capillary 2 is raised by the holding and lifting device 104. The holding and lifting device 104 is then moved over an as yet unoccupied target vessel 122 and the carrier element with the capillary 2 is lowered again. Finally, the drop with the bead contained therein is ejected, for example by means of compressed air, by the pump device 105 from the capillary 2 into the target vessel 122.
  • the holding and lifting device 104 then again raises the carrier element 9 with the capillary 2 and is moved back over the initial container 120, from which a new drop can be removed. In that manner it is possible to distribute the beads contained in the initial container 120 individually into the various target vessels 122.
  • the counting device makes it possible not only to count the beads contained in one drop but also to determine the number of drops sucked into the capillary 2, a plurality of drops can be sucked into the capillary 2 from the initial container 120 in one suction operation. If, for example, three drops are already present in the capillary 2, each one of which contains precisely one bead, and if, for example, two beads are detected in the fourth drop that is sucked in, then it is possible, by a metered supply of pressure, to eject only the fourth drop back into the initial container 120 and to keep the other three drops in the capillary 2. In that manner it is possible to suck up a plurality of drops into the capillary 2, each one of which contains precisely one bead.
  • the capillary 2 is then moved over an as yet empty target vessel 122. By a metered supply of pressure, precisely one of the drops in the capillary 2 is then ejected into the target vessel 122. The capillary is then moved over a different, as yet empty target vessel and the next drop is ejected. That operation is repeated until there are no more drops in the capillary 2. The capillary 2 can then be moved back to the initial container 120 again to take up new drops.
  • the possibility of sucking up a plurality of drops into the capillary is especially advantageous because the sorting process can, as a result, be carried out markedly more rapidly.
  • the sorting apparatus is not limited to applications in which the beads are to be distributed individually into target vessels. It is, of course, also possible to distribute the beads in larger portions. It may, for example, be desirable to distribute the beads in such a manner that not more than a predetermined number of beads is present in each of the target vessels. That predetermined number may be up to several dozens.
  • the counting device then registers whether the number of beads sucked into the capillary is greater than the predetermined number. If that is the case, the drops with the beads are ejected back into the initial container 120. If, on the other hand, the number of beads sucked up is smaller than or equal to the predetermined number, then the beads are ejected into an empty target vessel 122.
  • the sorting apparatus thus allows small particles, for example beads, to be distributed into target vessels 122 in such a manner that the number of particles introduced into the target vessel 122 can be controlled and determined.
  • the particles can be removed directly from the initial container 120, so that no further reservoirs are necessary. As a result, the risk of losing one of the particles during the distribution operation is reduced quite considerably.

Abstract

Dispositif servant à compter de petites particules, particulièrement des perles servant à synthétiser des substances chimiques, et comprenant un capillaire (2) servant à recevoir les particules à compter, une source de lumière (3) servant à émettre la lumière de mesure (M) et un détecteur photoélectrique (5) servant à recevoir la lumière de mesure (M). L'invention concerne également des moyens de transmission de la lumière qui transmettent la lumière de mesure (M) émise par la source de lumière (3) vers le capillaire (2), ainsi que des moyens de suppression de la lumière transmettant la lumière de mesure (M) provenant du capillaire (2) au détecteur photoélectrique (5). Le diamètre intérieur (D) du capillaire (2) est conçu de telle façon que les particules individuelles ne peuvent pénétrer dans le capillaire que l'une derrière l'autre et ne peuvent se déplacer le long du capillaire que l'une derrière l'autre. Les moyens de transmission de la lumière irradient le capillaire (2) par la lumière de mesure (M) dans un sens transversal par rapport à l'axe de la longueur (A) dudit capillaire, la lumière de mesure (M) étant sous forme d'un faisceau lumineux, dont la largeur est telle qu'une seule des particules peut se trouver en totalité dans le faisceau lumineux à la fois. Un appareil de tri servant à répartir ces petites particules comprend ce dispositif.
EP97915420A 1996-04-04 1997-03-24 Dispositif servant a compter de petites particules et appareil de tri comprenant ce dispositif Withdrawn EP0891555A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
EP97915420A EP0891555A1 (fr) 1996-04-04 1997-03-24 Dispositif servant a compter de petites particules et appareil de tri comprenant ce dispositif

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
EP96810215 1996-04-04
EP96810215 1996-04-04
EP97915420A EP0891555A1 (fr) 1996-04-04 1997-03-24 Dispositif servant a compter de petites particules et appareil de tri comprenant ce dispositif
PCT/EP1997/001479 WO1997038318A1 (fr) 1996-04-04 1997-03-24 Dispositif servant a compter de petites particules et appareil de tri comprenant ce dispositif

Publications (1)

Publication Number Publication Date
EP0891555A1 true EP0891555A1 (fr) 1999-01-20

Family

ID=8225581

Family Applications (1)

Application Number Title Priority Date Filing Date
EP97915420A Withdrawn EP0891555A1 (fr) 1996-04-04 1997-03-24 Dispositif servant a compter de petites particules et appareil de tri comprenant ce dispositif

Country Status (3)

Country Link
EP (1) EP0891555A1 (fr)
AU (1) AU2290897A (fr)
WO (1) WO1997038318A1 (fr)

Families Citing this family (19)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
GB2316081B (en) * 1996-08-12 2001-05-09 Bio Dot Ltd Dispensing of particles
JPH11328554A (ja) * 1998-03-24 1999-11-30 Pittway Corp 粒子センサ―を有する煙検出器
DE60030957T2 (de) 1999-02-16 2007-06-14 Applera Corp., Foster City Kugelabgabesystem
WO2001069202A2 (fr) * 2000-03-16 2001-09-20 Ulrik Darling Larsen Capteurs pour appareil de caracterisation de particules
US7968287B2 (en) 2004-10-08 2011-06-28 Medical Research Council Harvard University In vitro evolution in microfluidic systems
US20100137163A1 (en) 2006-01-11 2010-06-03 Link Darren R Microfluidic Devices and Methods of Use in The Formation and Control of Nanoreactors
EP2530168B1 (fr) 2006-05-11 2015-09-16 Raindance Technologies, Inc. Dispositifs microfluidiques
US8772046B2 (en) 2007-02-06 2014-07-08 Brandeis University Manipulation of fluids and reactions in microfluidic systems
WO2008130623A1 (fr) 2007-04-19 2008-10-30 Brandeis University Manipulation de fluides, composants fluidiques et réactions dans des systèmes microfluidiques
EP4047367A1 (fr) 2008-07-18 2022-08-24 Bio-Rad Laboratories, Inc. Procedé de détection d'analytes cibles au moyens des bibliothèques de gouttelettes
US9399797B2 (en) 2010-02-12 2016-07-26 Raindance Technologies, Inc. Digital analyte analysis
EP3392349A1 (fr) 2010-02-12 2018-10-24 Raindance Technologies, Inc. Analyse numérique d'analytes
WO2012045012A2 (fr) 2010-09-30 2012-04-05 Raindance Technologies, Inc. Dosages sandwich dans des gouttelettes
EP3412778A1 (fr) 2011-02-11 2018-12-12 Raindance Technologies, Inc. Procédés permettant de former des gouttelettes mélangées
EP3736281A1 (fr) 2011-02-18 2020-11-11 Bio-Rad Laboratories, Inc. Compositions et méthodes de marquage moléculaire
EP2714970B1 (fr) 2011-06-02 2017-04-19 Raindance Technologies, Inc. Quantification d'enzyme
US8658430B2 (en) 2011-07-20 2014-02-25 Raindance Technologies, Inc. Manipulating droplet size
US11901041B2 (en) 2013-10-04 2024-02-13 Bio-Rad Laboratories, Inc. Digital analysis of nucleic acid modification
US9944977B2 (en) 2013-12-12 2018-04-17 Raindance Technologies, Inc. Distinguishing rare variations in a nucleic acid sequence from a sample

Family Cites Families (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPS6085355A (ja) * 1983-10-17 1985-05-14 Union Giken:Kk 光学的検出器
JPH0781996B2 (ja) * 1988-08-27 1995-09-06 株式会社日立製作所 オートサンプラ
US5007740A (en) * 1989-03-30 1991-04-16 The Foxboro Company Optical probe for fluid light transmission properties
JPH0518967A (ja) * 1991-07-09 1993-01-26 Kyoto Daiichi Kagaku:Kk Ｐｓｐ排泄率検査装置
FR2719902B1 (fr) * 1994-05-11 1996-07-05 Pasteur Sanofi Diagnostics Système d'analyse optique d'un échantillon de mélange réactionnel.

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO9738318A1 *

Also Published As

Publication number Publication date
AU2290897A (en) 1997-10-29
WO1997038318A1 (fr) 1997-10-16

Similar Documents

Publication Publication Date Title
WO1997038318A1 (fr) Dispositif servant a compter de petites particules et appareil de tri comprenant ce dispositif
US7030988B2 (en) Measuring apparatus and measuring chip
US7724364B2 (en) Small object identifying device and its identifying method
JP6258353B2 (ja) 液滴内に含有されるサンプル分析のための光学測定装置及び方法
CN1973195B (zh) 试样的分离、识别、分注方法和其装置及解析装置
JP3139770B2 (ja) チップレベル導波管センサ
JP2969935B2 (ja) 液定量取出し装置
US7271885B2 (en) Plasmon resonance measuring method and apparatus
US8553229B2 (en) Fine particle optical measuring method in fluidic channels
JP7134251B2 (ja) 少なくとも１つの細胞および／または少なくとも１つの粒子を含有する液体を分配するためのディスペンサーを含む分配装置
US20070041875A1 (en) Flow analysis dispensing needles and nozzles
US6495812B1 (en) Apparatus and method for analyzing an object of interest having a pivotably movable source and detector
US20070059184A1 (en) Flow optical analysis for peristaltic and other rotary pumps
EP0047612A1 (fr) Appareil pour la détection de défauts dans les revêtements internes de récipients ouverts à une extrémité et entraînés en mouvement
CN116359128A (zh) 分配检测装置
JP2003254905A (ja) 測定装置
KR20210013058A (ko) 적어도 하나의 세포 및/또는 적어도 하나의 입자를 포함하는 액체를 검사하는 방법
US8049888B2 (en) Device for measuring light emitted by microscopically small particles or biological cells
JP2002277390A (ja) 測定チップ
JP2003344427A (ja) 分注装置
CN112986236A (zh) 用于确定液滴的位置和/或范围的装置和方法
EP4190450A1 (fr) Appareil d'imagerie d'une section de buse d'un dispositif de distribution de gouttelettes, distributeur comprenant l'appareil d'imagerie et applications associées
JP3024890B2 (ja) 使い捨てチップ及びそのチップを用いる分注装置
KR20230140568A (ko) 담체 상에 액체를 침착하기 위한 장치 및 방법
WO1999041589A1 (fr) Mesure d'absorption optique avec cavite d'integration de lumiere

Legal Events

Date Code Title Description
PUAI Public reference made under article 153(3) epc to a published international application that has entered the european phase

Free format text: ORIGINAL CODE: 0009012

17P Request for examination filed

Effective date: 19980817

AK Designated contracting states

Kind code of ref document: A1

Designated state(s): CH DE DK FR GB IT LI NL SE

STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: THE APPLICATION IS DEEMED TO BE WITHDRAWN

18D Application deemed to be withdrawn

Effective date: 20000901