EP1616619A1 - Dispositf et procédé ayant une influence sur bulles d'air dans une chambre hybridation - Google Patents

Dispositf et procédé ayant une influence sur bulles d'air dans une chambre hybridation Download PDF

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Publication number
EP1616619A1
EP1616619A1 EP05106505A EP05106505A EP1616619A1 EP 1616619 A1 EP1616619 A1 EP 1616619A1 EP 05106505 A EP05106505 A EP 05106505A EP 05106505 A EP05106505 A EP 05106505A EP 1616619 A1 EP1616619 A1 EP 1616619A1
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EP
European Patent Office
Prior art keywords
hybridization
air bubbles
slide
relief structures
space
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Withdrawn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
EP05106505A
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German (de)
English (en)
Inventor
Wolfgang Streit
Gyoergy Wenczel
Waltraud Lamprecht
Heribert Eglauer
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Tecan Trading AG
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Tecan Trading AG
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Publication date
Priority claimed from DE202004011272U external-priority patent/DE202004011272U1/de
Application filed by Tecan Trading AG filed Critical Tecan Trading AG
Publication of EP1616619A1 publication Critical patent/EP1616619A1/fr
Withdrawn legal-status Critical Current

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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01F—MIXING, e.g. DISSOLVING, EMULSIFYING OR DISPERSING
    • B01F25/00—Flow mixers; Mixers for falling materials, e.g. solid particles
    • B01F25/40—Static mixers
    • B01F25/42—Static mixers in which the mixing is affected by moving the components jointly in changing directions, e.g. in tubes provided with baffles or obstructions
    • B01F25/43—Mixing tubes, e.g. wherein the material is moved in a radial or partly reversed direction
    • B01F25/431—Straight mixing tubes with baffles or obstructions that do not cause substantial pressure drop; Baffles therefor
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L7/00—Heating or cooling apparatus; Heat insulating devices
    • B01L7/52—Heating or cooling apparatus; Heat insulating devices with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01F—MIXING, e.g. DISSOLVING, EMULSIFYING OR DISPERSING
    • B01F31/00—Mixers with shaking, oscillating, or vibrating mechanisms
    • B01F31/65—Mixers with shaking, oscillating, or vibrating mechanisms the materials to be mixed being directly submitted to a pulsating movement, e.g. by means of an oscillating piston or air column
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01F—MIXING, e.g. DISSOLVING, EMULSIFYING OR DISPERSING
    • B01F33/00—Other mixers; Mixing plants; Combinations of mixers
    • B01F33/30—Micromixers
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip
    • B01L3/502723—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip characterised by venting arrangements
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/06—Fluid handling related problems
    • B01L2200/0684—Venting, avoiding backpressure, avoid gas bubbles
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0809—Geometry, shape and general structure rectangular shaped
    • B01L2300/0822—Slides
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0861—Configuration of multiple channels and/or chambers in a single devices
    • B01L2300/0877—Flow chambers
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/18—Means for temperature control
    • B01L2300/1805—Conductive heating, heat from thermostatted solids is conducted to receptacles, e.g. heating plates, blocks
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0475—Moving fluids with specific forces or mechanical means specific mechanical means and fluid pressure
    • B01L2400/0481—Moving fluids with specific forces or mechanical means specific mechanical means and fluid pressure squeezing of channels or chambers
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0475—Moving fluids with specific forces or mechanical means specific mechanical means and fluid pressure
    • B01L2400/0487—Moving fluids with specific forces or mechanical means specific mechanical means and fluid pressure fluid pressure, pneumatics
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/08—Regulating or influencing the flow resistance
    • B01L2400/084—Passive control of flow resistance
    • B01L2400/086—Passive control of flow resistance using baffles or other fixed flow obstructions

Definitions

  • the invention relates to a device according to the preamble of independent claim 1 and to a corresponding process unit according to claim 15 and to an automatic system according to the preamble of claim 17 for the hybridization of nucleic acid samples, proteins and tissue sections.
  • the present invention also relates to a corresponding method.
  • DNA deoxyribose nucleic acid
  • microarrays of such samples provides research with an important technique for the simultaneous analysis of thousands of genes.
  • This technique involves the immobilization of DNA samples from many genes on a solid substrate surface, such as on a glass slide for a light microscope.
  • the DNA samples are preferably arranged in an array of sample spots or "spots", ie in a two-dimensional grid on the substrate, and it is possible later on - starting from a specific one Position within such an array back to the origin of the corresponding DNA sample.
  • RNA patterns are often provided with a so-called “tag” or “label”, ie a molecule which, for example, emits a fluorescent light having a specific wavelength.
  • Immobilized samples may also include amino acid-containing (eg, proteins, peptides) or nucleic acid-containing (eg, cDNA, RNA) samples. Samples added to the immobilized samples may comprise any molecules or chemical compounds that hybridize or otherwise associate with the immobilized samples.
  • RNA patterns hybridize or bind to immobilized DNA samples and together form hybrid DNA-RNA strands.
  • differences in the hybridization among the DNA samples can be determined by measuring the intensity and wavelength dependency of the fluorescence of each individual microarray element to determine whether the level of gene expression in the DNA sample varies the examined DNA samples.
  • DNA microarrays have established themselves as successful tools and devices for carrying out DNA hybridization have been continuously improved (cf., for example, US Pat. No. 6,238,910 or US 2003/0013184 A1 or EP 1 260 265 A1 of the present applicant).
  • These documents disclose a device for providing a hybridization space for the hybridization of nucleic acid samples on a slide, which is designed to be movable relative to this slide and an annular seal for closing the Hybridisierraums by applying a surface of this slide comprises.
  • the disclosed apparatus includes conduits for feeding media into and out of the media Hybridization in or out of the Hybridisierraum out and a sample feed.
  • the object of the present invention according to a first aspect relates to the provision of an alternative device and according to a second aspect the provision of a method with which or with which air bubbles in a hybridization chamber can be influenced in a simple manner.
  • This object according to the first aspect is achieved with the combination of features of independent claim 1 and is characterized in that the device delimits a slit-shaped hybridization space which on a surface of the device and / or the slide directed towards the interior of this hybridization space has relief structures for conducting and / or or blocking air bubbles.
  • This object according to the second aspect is achieved by moving and / or trapping air bubbles according to the features of independent claim 19 within the hybridization chamber.
  • the present invention is based on the consideration that on the one hand gas molecules of the ambient air can diffuse into the hybridization chamber and thus air bubbles are formed. On the other hand, for example, as a result of changes the solubility of gases, spontaneously form gas bubbles in Hybridisiermedium. In the context of this invention, therefore, all gas bubbles in the hybridization medium - regardless of the formation process in the hybridization chamber - are referred to as "air bubbles".
  • FIG. 1 shows a vertical longitudinal section, corresponding to FIG. 7 of US 2003/0013184 A1 or EP 1 260 265 A1 of the present applicant, through a device 1 for the hybridization of nucleic acid samples, proteins and tissue sections.
  • This device 1 is movable like a cover in relation to a slide 3 (in this case pivotable about an axis 34, so that the hybridization space 2 can be opened by a simple movement (compare Fig. 1A) and closed (see Fig. 1B).
  • An annular seal or sealing surface 4 is used to close the Hybridisierraums 2 by applying a surface 5 of this slide 3.
  • This sealing surface 4 may be a stepped surface or step 104 of the lid or the device 1, which is flat on the surface 5 of the slide 3 Alternatively, for example, a lip seal may also be used, but preference is given to an O-ring seal 103 as a sealing surface 4.
  • the arrangement comprises lines 6, 6 'for feeding or discharging media into or out of the hybridization space 2 the hybridization space 2.
  • Such media may contain reagents for carrying out the hybridization reaction, such as wash liquors or buffer solutions, but also inert gases (such as e.g. Nitrogen) for drying the hybridization products on the slides 3 or for blowing out the Hybridisierhuntn 2 and the media lines 6,6 'be.
  • the assembly also includes a sealable pattern feeder 7 through which RNA-containing liquids or other sample liquids can be pipetted by hand.
  • the pattern feeder 7 is preferably closed with a plastic plug (not shown).
  • an automatic or robotized pattern feeder can be provided, as disclosed in different embodiments in US 2003/0013184 Al or EP 1 260 265 A1 of the present applicant, to which reference is expressly made.
  • the devices 1 are arranged parallel to each other and in a group of four, because this arrangement just allows a measure of a temperature control plate 20, on which a frame 21 in the size of a microplate with four parallel to each other arranged slides 3 fits.
  • Each of these groups of four is associated with a temperature control plate 20 connected to a temperature controller.
  • a temperature control plate 20 is designed for the areal reception of a frame 21 carrying four slides 3. Because the slides 3 are held softly in the frame 21 and because the temperature control plate 20 is formed so that the frame can be slightly lowered towards it, the slides 3 lie directly on the surface of the temperature control plate 20.
  • Each group of four of a processing unit 18 comprises one an axle 34 pivotable and relative to a base plate 35 lockable bracket 36 with four seats 37, wherein in each of these seats 37, a device 1 is inserted.
  • Each processing unit 18 also includes a terminal plate 22 for sealingly connecting the unit lines 23, 23 ', 23 "of the system 38 to the lines 6, 6', 6" of the devices 1. O-rings are preferred as seals for these connections (not shown) ).
  • the arrangement preferably comprises a media-separating agitation device 8 for moving liquids against nucleic acid- or amino acid-containing samples or tissue sections immobilized on the surface 5 of the slides 3.
  • the agitation device 8 of the arrangement comprises a membrane 9. This membrane 9 separates a pressure chamber 10, which is designed to be filled with a pressure fluid (gas or liquid) via a pressure line 6 ", from an agitation space 11 which passes over an agitation conduit 12 is connected to the hybridization space 2.
  • RNA sample liquid and sealing the pattern feed 7 air or other gas is preferably delivered via the pressure conduit 6 "(but it could also be a liquid) brought into the pressure chamber 10 in blocks (Überduckversion) or sucked from it (vacuum version), so that the membrane 9 bends in the same rhythm and correspondingly reduces or increases the agitation space 11.
  • the sample liquid is moved in the same rhythm of overpressure or underpressure and relaxation in Hybridisierraum 2 against one or the other end, preferably located on the directed against the interior of the Hybridisierraums 2 surface 14 of the lid or the device 1 each one Querströmkanal 15,15 'is located.
  • these transverse flow channels 15, 15 ' facilitate the transverse distribution of RNA molecules contained in the sample solution. This has the effect that the sample liquid or the washing liquids are distributed homogeneously over the entire volume present in the hybridization chamber 2.
  • the cross-flow channels 15, 15 ' also serve as a liquid reservoir, so that the oscillating motion of the model solution generated by the agitation device 8 built into the device does not cause parts of the hybridization chamber 2 to be unintentionally dry.
  • a second agitation space 11 ' is connected to the hybridization space 2 via a second agitation conduit 12 '. If a pressure surge delivered to the pressure chamber 10 now presses the first diaphragm 9 into the first agitation space 11, this pulse is transmitted via the first agitation conduit 12 to the sample fluid in the hybridization space 2.
  • the pattern liquid differs somewhat from the second agitation conduit 12 '(and may even partially fill it) and increases the pressure in the second agitation space 11'.
  • the second membrane 9 ' bends upwards and is elastically stretched.
  • a sample liquid having a minimum volume in the range of approximately 100 ⁇ l
  • a negative pressure in the pressure chamber 10 is generated, so that the backward movement of the sample liquid in the hybridization space 2, which is opposite to the preceding pressure surge, is enhanced.
  • FIG. 1A illustrates an open holder 36
  • FIG. 1B shows a vertical section through a processing unit 18 for hybridizing nucleic acid samples, proteins or tissue slices with closed holder 36. All four hybridization spaces 2 of these groups of four are associated with a temperature control plate 20 connected to a temperature controller. To ensure that the devices 1 are placed plane-parallel to the slides 3
  • the holder 36 also has a central joint (not shown) with a mobility parallel to the axis 34.
  • an additional pressure is exerted on the devices 1 via the holder 36; this can be accomplished via screws, rocker arms or similar known devices (not shown).
  • Each processing unit 18 also includes a terminal plate 22 for sealingly connecting the unit conduits 23, 23 ', 23 "to the conduits 6, 6', 6" of the devices 1. As seals for these compounds, O-rings are preferred (not shown).
  • Figures 2 and 3 show schematic views of the closed arrangement of Figure 1B, seen from below.
  • the gap-shaped hybridization chamber 2 between a device 1 and a slide 3, on which this device 1 is placed defined in its volume.
  • the O-ring seal 103 laterally delimits the hybridization space 2, which preferably has a respective transverse flow channel 15, 15 'at its opposite ends, which are provided as a depression in the surface 14 of the cover or device 1.
  • the slide 3 here a glass slide for light microscopy
  • its optional handle and / or barcode field 33 are shown in dashed lines.
  • Clearly visible is also a Abdrückfeder 17, which presses on the handle panel 33 of the slide 3. It can also be provided several Abdschreibfedern (not shown).
  • 1 centering springs may be arranged (preferably not shown) on the side edges of the device, with which slides with special formats are centered on the device 1, so that maximum utilization of the slide surface is ensured.
  • this Abdgurfeder 17 facilitates the automatic separation of the slide 3 from the lid or of the device 1. Also visible are the openings of the inlet conduit 6 and the outlet conduit 6 'and the pattern supply 7, in one of the two Querströmkanäle 15, 15 'lead.
  • FIG. 2A shows a device 1 with an arrangement of relief structures 101 for blocking air bubbles 102 according to a first variant.
  • These relief structures 101 are formed here as depressions 105 and arranged in boundary regions 106 of the hybridization space 2 along at least part of the sealing surface 4. As shown, no lines 6,6 ', 7 open into these recesses 105.
  • a plurality of depressions are arranged in a row to the left and right of the hybridization chamber 2, these rows extending over the entire boundary region 106 between the two terminal transverse flow channels 15 , 15 'extend.
  • the depressions 105 are in the immediate vicinity of the O-ring 103, they are all the same length and interrupted by narrow webs 107, so that the air bubbles 102 are trapped in the respective recess 105 and do not connect to the slide 3 and thus a hybridization prevent or obstruct the underlying samples. In addition, air bubbles 102 trapped in the depressions 105 can not spread over the hybridization space 2.
  • FIG. 2B shows an arrangement of relief structures 101 for blocking air bubbles 102 according to a second and third variant.
  • the recesses 105 are in the immediate vicinity of the O-ring 103, they are all the same length and interrupted by narrow webs 107, so that the air bubbles 102 are trapped in the respective recess 105.
  • the second variant left only a few, long recesses 105 are provided; it could also be arranged only a single recess on each side of the hybridization chamber 2 (not shown), it is important that a web 107 these wells 105 from the Querströmkanälen 15,15 'separates.
  • the third variant (right), a large number of short (for example round) depressions 105 are provided. These have the advantage that many individual compartments for blocking air bubbles 102 are created.
  • FIG. 2C shows an arrangement of relief structures 101 for blocking air bubbles 102 according to a fourth and fifth variant.
  • the recesses 105 are in the immediate vicinity of the O-ring 103, they have a different length and are interrupted by narrow webs 107, so that the air bubbles 102 are trapped in the respective recess 105.
  • the fourth variant left
  • only a few, long depressions 105 are provided, between which short (eg round) depressions can be arranged.
  • the fifth variant (right) several recesses 105 are provided with increasing length.
  • the arrangements shown here have the advantage that compartments to block air bubbles 102 which are individually distributed according to certain process parameters or chamber characteristics. For example, it is thus possible to provide recesses which are arranged correspondingly to the gradient occurring in the experience (in the hybridization medium and / or in the appearance of the air bubbles).
  • relief structures 101 formed as depressions 105 are particularly suitable for blocking air bubbles 102 produced by diffusion through the O-ring 103, namely in the immediate vicinity of their point of origin.
  • the variation in the width and / or depth of such depressions 105 is also within the scope of the present invention.
  • catch channels for air bubbles along the lateral O-rings.
  • Such collecting channels are then formed as a single depression, which is only separated by a thin web of the two Querströmkanälen 15,15 '.
  • the recesses 105 according to the invention can also be used in hybridization chambers 2, which does not comprise an agitation device.
  • FIG. 3A shows an arrangement of relief structures 101 for conducting air bubbles 102 according to a first variant.
  • the relief structures 101 are here designed as elevations 108 for conducting air bubbles 102 and distributed substantially over the entire hybridization space 2.
  • the elevations 108 are arranged regularly and in an entangled, orthogonal grid 109.
  • an entangled, orthogonal grid 109 which is arranged rotated by 45 ° to a flow axis 110. Deviations from the orthogonal arrangement are possible, so that by a smaller angle to the direction of flow than 45 ° (steeper arrangement) the detachment is facilitated.
  • steereper arrangement steereper arrangement
  • a greater distance is achieved when pushing away.
  • FIG. 3B shows an arrangement of relief structures 101 for conducting and blocking air bubbles 102.
  • the blocking relief structures 101 are in the form of depressions 105 is formed and arranged in boundary regions 106 of the hybridization space 2 along at least part of the O-ring 103 (see FIG.
  • the conductive relief structures 101 are formed as elevations 108 for conducting air bubbles 102 and distributed substantially over the entire hybridization space 2 (see Fig. 3A).
  • the combination of blocking and conductive relief structures 101 also makes it possible to combine the already described positive effects of these relief structures.
  • the arrangement and the size of all relief structures 101 used for influencing air bubbles 102 are preferably adapted to one another.
  • the volume of the hybridization chamber preferably about 60 .mu.l
  • the stroke of the agitation device preferably about 5 .mu.l
  • the length of the hybridization chamber preferably about 50 mm
  • the pendulum path of the hybridization chamber resulting from the agitation Hybridization media over the immobilized samples preferably about 5 mm.
  • the frequency of Agitationszyklen is adjustable. This means that different periods of agitation are alternated with different long times without agitation. However, the agitation times are preferably chosen so long that results in a large number of agitation movements per minute and thus a multiple mixing of the chamber volume. Mixing tests with dyes have shown that after just a few minutes a virtually 100% thorough mixing takes place.
  • Figure 3C shows an alternative arrangement of relief structures for guiding and blocking air bubbles.
  • the blocking relief structures 101 are formed as smaller depressions 105 (see FIG. 3B).
  • the conductive relief structures 101 are formed as finer and more closely spaced raised bumps 108 for directing air bubbles 102 (see Fig. 3B).
  • FIG. 4 shows vertical partial sections through an arrangement according to FIG. 1B with the device 1 folded down or the hybridization chamber 2 closed. Arrows indicate the direction of flow of the sample fluid relative to the samples immobilized on the slide 3. This movement of the sample liquid, in interaction with the relief structures 101 shown, causes the air bubbles 102 to be displaced away from the actual flow direction or flow axis 110.
  • FIG. 4A shows an embodiment of relief structures 101 for conducting air bubbles 102 according to a first variant.
  • This relief structure is formed as a recess 105, which has a flat and a steep edge.
  • the flanks of the depression 105 each form a line of intersection with the surface 14 of the device 1.
  • these intersecting lines run parallel to one another (compare FIG. 5).
  • FIG. 5 shows an embodiment of relief structures 101 for conducting air bubbles 102 according to a first variant.
  • This relief structure is formed as a recess 105, which has a flat and a steep edge.
  • the flanks of the depression 105 each form a line of intersection with the surface 14 of the device 1.
  • FIG. 5 With increasing height difference and constant slope of the depression 105 diverge these intersections; For example, a fish scale-like relief on the surface 14 of the device 1 can be generated.
  • the inclination angle of the steep flank and the angle between the flow axis 110 and the intersection between the steep flank and the surface 14 of the device 1 together determine the influence on an air bubble 102 which is flushed against this steep flank.
  • these two angles are selected such that the flow resistance of the air bubble 102 is greater in relation to the steep flank in the direction of the flow axis 110 than in the direction of the relief structure 101.
  • the air bubble 102 is forced away from the general direction of flow.
  • This displacement is additionally supported by the, in the same direction deflected, local microflow caused by the relief structure 101.
  • These microcurrents occur at all relief structures 101 arranged at an angle to the flow axis 110 and additionally support a more effective mixing of the sample liquid.
  • a further preferred variant comprises the formation of conductive recesses 105, which begin at one end in the depressions 105 for blocking the air bubbles 102 and which extend beyond the middle of the hybridization space 2 (by those shown in FIGS. 2 and 3) drawn file 110 marked).
  • These conductive recesses 105 are also preferably oriented at an angle of 45 ° to the flow axis 110 and arranged alternately starting from the left or right side of the Hybridisierhunt 2 and lying at 90 ° to each other.
  • These conductive recesses 105 may be formed consistently deep or tapered to the center of the Hybridisierraums 2.
  • FIG. 4B shows an embodiment of relief structures 101 for conducting air bubbles 102 according to a second variant.
  • This relief structure is formed as a survey 108, which has a flat and a steep edge.
  • FIG. 4C shows an embodiment of relief structures 101 for conducting air bubbles 102 according to a third variant.
  • This relief structure is partially formed as a recess 105 and partially as a survey 108 and has two steep and a flat edge. All comments on FIG. 4A also apply mutatis mutandis to FIGS. 4B and 4C.
  • FIG. 4D shows an embodiment of relief structures 101 for conducting air bubbles 102 according to a fourth variant.
  • This relief structure is formed as an elevation 108 and has two steep flanks and a flat (raised) surface. All comments on Figure 4A apply mutatis mutandis here.
  • Another variant (not shown) comprises the formation of relief structures 101 for conducting air bubbles 102, in which this relief structure 101 is formed as a depression 105 and has two steep flanks and a flat (recessed) surface.
  • Both variants have in common that the lines of intersection of the steep flanks with the surface 14 of the device 1 can have a parallel course, so that the height difference of these relief structures 101 is constant (shown). However, both variants also have in common that the intersection lines of the steep flanks with the surface 14 of the device 1 may have a divergent course, and that the height difference of these relief structures 101 is nonetheless constant (not shown).
  • relief structures 101 formed as elevations 108 with a dome-shaped cross section are also preferred (not shown). With such elevations 108, air bubbles 102 having a diameter of at most about 50 ⁇ m are more likely to only make a point contact. Thus, even with relatively little agitation, the air bubbles do not remain attached to the elevations 108 and are also further flushed into the transverse flow channels 15, 15 '. If the air bubbles 102 do not remain trapped in recesses 105 on the way, they are collected in the crossflow channels 15, 15 '.
  • depressions 105 is in most cases preferred to the elevations 108, because wells always also mean an increase in the height of the preferably about 60-70 ⁇ m high hybridization chamber 2. This reduces the risk that an air bubble will affect the hybridization of a sample.
  • Particularly preferred is an at least approximate, volumetric compensation of recesses 105 and elevations 108, so that a defined volume for the gap between the device 1 and the slide 3 and thus a defined volume for the Hybridisierhunt 2 can be maintained.
  • the blocking of air bubbles 102 with devices 1 according to the invention basically functions in systems equipped with or without agitation means for moving the sample solution with respect to the immobilized samples.
  • a system 38 for hybridizing nucleic acid samples, proteins or tissue sections is equipped with an agitation device (for example as shown and described in FIG. 1), a pendulum movement of the sample fluid relative to the samples immobilized on the slide 3 occurs.
  • Figure 5 shows a schematic partial view of a Hybridisierhunt 2 between a device 1 and a slide 3, on which this device 1 is placed.
  • the device 1 has an arrangement of relief structures 101 for guiding and blocking air bubbles 102 according to a respective first variant (see Fig. 3B) and an embodiment of these relief structures 101 as elevations 108 according to the fourth variant (compare Fig. 4D) ,
  • the track which the Tracing pattern molecules on average during a pendulum motion generated by an agitator is marked with two opposing arrows. This distance can also be much longer but not much shorter than outlined in the schematic diagram.
  • a distance of two behind each other in the direction of agitation the same direction relief structures of about 2/3 of the pendulum motion or about 4 mm.
  • the typical path of an air bubble 102 begins at an arbitrary position (1) in the Hybridisierraum 2.
  • This air bubble 102 with a diameter of at most about 50 microns (with a chamber height or gap height of preferably 30-200 microns) of the (in the sketch detected upward flow) and moved substantially in a straight line until it abuts a trained as a survey 108, the first relief structure 101 (2).
  • the resistance to escape of the air bubble 102 (here: below 45 ° to the top right) remains considerably smaller than standing, it is pulled along by the flow as far as the end of the relief structure 101 (3, 4).
  • the air bubble 102 then continues to move in a straight line until it encounters a second relief structure 101 (FIG. 5), is distracted therefrom (here: at 45 ° to the top left) and stops after breaking off the flow (6).
  • the flow sets in the opposite direction and pulls the air bubble 102 until it hits the back of the first relief structure 101 (FIG. 7), is deflected again by it (here: at 45 ° to the bottom left) and into the depression 105 formed relief structure 101 enters (8).
  • the air bubble 102 is trapped and can only follow the pendulum flow within these narrow limits (9). If other air bubbles are already blocked in the depression 105, an aggregation of the existing air bubbles or even a fusion thereof is often observed. These two processes additionally increase the probability that none of the air bubbles can leave the depression 105 again.
  • the relief structures 101 according to the invention (elevations and depressions) on the cover or on the device 1 preferably have a height difference from the otherwise flat surface 14 of the lid or device 1 of at most 1/3 of the preferred gap height of 30 .mu.m to 200 .mu.m. Especially preferred are differences in height of 30 microns at wells and 20 microns at elevations.
  • These relief structures 101 may enter the device 1 during the manufacturing process (eg injection molding, processing of bar material) are formed. Recesses can also be incorporated later (eg by milling). Also surveys can be subsequently applied to the actual manufacturing process of the device 1 (eg by gluing or depositing of resins). This can be done for example by means of a template or prefabricated on adhesive strips semi-finished products.
  • Object slides 3 can also be provided with relief structures 101, be these recesses 105 and / or elevations 108.
  • the relief structures 101 are preferably arranged on the object carriers 101 on the surface 5 corresponding to the expected sample distributions. Thus, e.g. square so-called "low density arrays" be separated by channel-like depressions.
  • the slides 3 may include a seal that is part of or disposed on the slide. Such seals may e.g. be produced in a two-component injection molding process with plastic slides. The spraying of such, preferably soft seals 4 on glass slides is also possible.
  • the device 1, or the lid closing off the hybridization space 2 may comprise a hard seal 4 for maintaining the defined gap width or height of the hybridization space.
  • the use of e.g. composite gaskets produced by the two-component injection molding method with a soft (sealing) component and a hard component (defining the gap height) are also conceivable.
  • the material of the device 1 preferably polysulfone
  • Each annular assembly or ring with sealing function is referred to in the context of the present invention as an O-ring.
  • Such an O-ring may optionally have a e.g. have round, elliptical or polygonal cross-section or be formed as a sealing lip.
  • the surface 5 of the object carrier 3 is arranged on the surface plane-parallel to the surface 14 of the device 1.
  • the hybridization space 2 is not dependent on an approximately rectangular base area, as in FIGS. 2 and 3 will be shown.
  • the recesses 105 can be arranged according to the seal 4.
  • bumps 108 may be arranged according to the present invention in an eg tangential direction of agitation.
  • the device 1 for influencing the air bubbles need not necessarily be transparent or have transparent parts.
  • the device 1, the slide 3, or even both of these components may have relief structures 101, be these depressions 105 and / or elevations 108.

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  • Chemical & Material Sciences (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Health & Medical Sciences (AREA)
  • Dispersion Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Clinical Laboratory Science (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Biochemistry (AREA)
  • Molecular Biology (AREA)
  • Analytical Chemistry (AREA)
  • Hematology (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)
EP05106505A 2004-07-17 2005-07-15 Dispositf et procédé ayant une influence sur bulles d'air dans une chambre hybridation Withdrawn EP1616619A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
DE202004011272U DE202004011272U1 (de) 2004-07-17 2004-07-17 Vorrichtung zum Bereitstellen einer Hybridisierkammer und zum Beeinflussen von Luftblasen in derselben
US10/909,521 US7459306B2 (en) 2004-07-17 2004-08-02 Device and method for providing a hybridization chamber and for influencing air bubbles in the same

Publications (1)

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EP1616619A1 true EP1616619A1 (fr) 2006-01-18

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Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
DE202008009556U1 (de) * 2008-07-16 2009-12-03 Eppendorf Ag Vorrichtung zum Temperieren wenigstens einer Probe
WO2013004673A1 (fr) * 2011-07-05 2013-01-10 Boehringer Ingelheim Microparts Gmbh Structure microfluidique comportant des creux
WO2013127990A1 (fr) * 2012-03-01 2013-09-06 Victorious Medical Systems Aps Procédé et système de répartition et d'agitation d'une quantité de liquide sur une lame de microscope
EP2899283A4 (fr) * 2012-09-24 2016-05-04 Toyo Seikan Group Holdings Ltd Procédé d'élimination des bulles et dispositif associé

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0785433A2 (fr) * 1996-01-17 1997-07-23 bioMerieux Vitek, Inc. Carte test pour analyses
US5951952A (en) * 1995-05-31 1999-09-14 Biomerieux, Inc. Test sample card
EP1260265A1 (fr) * 2001-05-25 2002-11-27 Tecan Trading AG Appareil pour la préparation d'une chambre d'hybridation, set de traitament et système pour l'hybridation d'échantillons d'acide nucléique, protéines et tissus

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5951952A (en) * 1995-05-31 1999-09-14 Biomerieux, Inc. Test sample card
EP0785433A2 (fr) * 1996-01-17 1997-07-23 bioMerieux Vitek, Inc. Carte test pour analyses
EP1260265A1 (fr) * 2001-05-25 2002-11-27 Tecan Trading AG Appareil pour la préparation d'une chambre d'hybridation, set de traitament et système pour l'hybridation d'échantillons d'acide nucléique, protéines et tissus

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
DE202008009556U1 (de) * 2008-07-16 2009-12-03 Eppendorf Ag Vorrichtung zum Temperieren wenigstens einer Probe
WO2013004673A1 (fr) * 2011-07-05 2013-01-10 Boehringer Ingelheim Microparts Gmbh Structure microfluidique comportant des creux
US9409171B2 (en) 2011-07-05 2016-08-09 Boehringer Ingelheim Microparts Gmbh Microfluidic structure having recesses
WO2013127990A1 (fr) * 2012-03-01 2013-09-06 Victorious Medical Systems Aps Procédé et système de répartition et d'agitation d'une quantité de liquide sur une lame de microscope
EP2899283A4 (fr) * 2012-09-24 2016-05-04 Toyo Seikan Group Holdings Ltd Procédé d'élimination des bulles et dispositif associé
US10077124B2 (en) 2012-09-24 2018-09-18 Toyo Seikan Group Holdings, Ltd. Bubble removal method and bubble removal device

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