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WO2001031347A1 - Appareil modulaire automatisé de traitement d'échantillons - Google Patents

Appareil modulaire automatisé de traitement d'échantillons Download PDF

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Publication number
WO2001031347A1
WO2001031347A1 PCT/GB2000/004142 GB0004142W WO0131347A1 WO 2001031347 A1 WO2001031347 A1 WO 2001031347A1 GB 0004142 W GB0004142 W GB 0004142W WO 0131347 A1 WO0131347 A1 WO 0131347A1
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WO
WIPO (PCT)
Prior art keywords
fluid
chambers
sample
chamber
microarray
Prior art date
Application number
PCT/GB2000/004142
Other languages
English (en)
Inventor
Harry Aldridge
Colin Burns
Paul Lloyd
Paul Shaw
Sophie Wildsmith
Original Assignee
Smithkline Beecham Plc
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 Smithkline Beecham Plc filed Critical Smithkline Beecham Plc
Priority to JP2001533433A priority Critical patent/JP2003513246A/ja
Priority to EP00971597A priority patent/EP1228375A1/fr
Priority to AU10431/01A priority patent/AU1043101A/en
Publication of WO2001031347A1 publication Critical patent/WO2001031347A1/fr

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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
    • 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
    • B01J2219/00596—Solid-phase processes
    • 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
    • B01J2219/00605—Making arrays on substantially continuous surfaces the compounds being directly bound or immobilised to solid supports
    • 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
    • B01J2219/00605—Making arrays on substantially continuous surfaces the compounds being directly bound or immobilised to solid supports
    • B01J2219/00608—DNA chips
    • 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
    • B01J2219/00605—Making arrays on substantially continuous surfaces the compounds being directly bound or immobilised to solid supports
    • B01J2219/00612—Making arrays on substantially continuous surfaces the compounds being directly bound or immobilised to solid supports the surface being inorganic
    • 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
    • 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
    • B01J2219/0068—Means for controlling the apparatus of the process
    • B01J2219/00686—Automatic
    • B01J2219/00689—Automatic using computers
    • 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
    • B01J2219/0068—Means for controlling the apparatus of the process
    • B01J2219/00702—Processes involving means for analysing and characterising the products
    • B01J2219/00707—Processes involving means for analysing and characterising the products separated from the reactor apparatus
    • 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
    • B01J2219/00718—Type of compounds synthesised
    • B01J2219/0072—Organic compounds
    • B01J2219/00722—Nucleotides
    • 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
    • B01J2219/00718—Type of compounds synthesised
    • B01J2219/0072—Organic compounds
    • B01J2219/00725—Peptides
    • C—CHEMISTRY; METALLURGY
    • C40—COMBINATORIAL TECHNOLOGY
    • C40B—COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES
    • C40B40/00—Libraries per se, e.g. arrays, mixtures
    • C40B40/04—Libraries containing only organic compounds
    • C40B40/06—Libraries containing nucleotides or polynucleotides, or derivatives thereof
    • C—CHEMISTRY; METALLURGY
    • C40—COMBINATORIAL TECHNOLOGY
    • C40B—COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES
    • C40B40/00—Libraries per se, e.g. arrays, mixtures
    • C40B40/04—Libraries containing only organic compounds
    • C40B40/10—Libraries containing peptides or polypeptides, or derivatives thereof
    • 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
    • G01N2035/00178—Special arrangements of analysers
    • G01N2035/00326—Analysers with modular structure
    • 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
    • G01N35/1095—Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices for supplying the samples to flow-through analysers
    • G01N35/1097—Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices for supplying the samples to flow-through analysers characterised by the valves

Definitions

  • the present invention relates to a method for processing samples, particularly biological samples on solid supports such as microarrays of genes, and to apparatus adapted for use in this method.
  • Genomics particularly in the area of gene expression, is used increasingly in the pharmaceutical and biotechnology sector. Rapid technological developments have enabled gene expression monitoring, see Brown et al, Nature Genetics, 1999, 21, 33-37; Watson et al, Current opinion in Biotechnology, 1998, 9, 609-614; and Bowtell et al, Nature Genetics supplement, 1999, 21, 25-32, notably using microarrays.
  • the level of expression of thousands of genes in one sample can be measured at one time, see Marton et al, Nature Medicine, 1998, 4, 11, 293-1301; De Risi et al, Science, 1997, 278, 680-686; and Lashkari et al, 1997, 94, 3057-13062.
  • microarrays and their potential for screening has been identified, see DeBouck et al, Nature, 1999, yet the automation of their use leaves a loc to be desired. There is currently no commercial system which will automate the process from sample loading right through to the stage where a microarray which can be imaged is obtained.
  • the initial stages of making microarrays, by stamping onto grids has been automated, with a number of manufacturers producing commercially available "gridding" machines.
  • the final steps of using microarrays have also received attention and there are a number of imaging instruments on the market, some with carousels which can automatically load a number of slides for imaging.
  • this system has a number of drawbacks: it is only available for 2 slides at any one time; there are only 3 positions for solutions, which limits the number of washes which can be performed, and means that only the hybridisation step, and not the slide preparation step, can be automated; the processing solutions and sample are heated whilst in contact with the slide; the sample injection step is manual; and the system is based on a vacuum which prevents even removal of the solutions, resulting in streaks and preventing the slides from being read accurately.
  • the present invention relates to an apparatus which allows the automation of slide preparation, sample preparation, sample loading, hybridisation, washing and drying; and to a method of processing samples using this apparatus.
  • the apparatus and the method of the invention are convenient to use and give reproducibly high quality microarrays.
  • a particularly surprising finding is that the apparatus and method of the invention significantly reduces or eliminates the phenomenon of "streaking", whereby a portion of the sample that is not firmly adhered to the solid support may spread beyond the bounds of the original spots, commonly seen in prior art methods.
  • a further significant advantage over prior art methods is the reproducibility of the resultant microarrays, improved reproducibility arises largely because the of the greater consistency in timing of the processing steps compared to a manual method.
  • an automated sample processing apparatus comprising: a plurality of support blocks each adapted to receive a solid support bearing the sample to be processed and/or reagent, such that in use the support block and the solid support form a substantially enclosed chamber; means for introducing fluid into the chambers and removing fluid from the chambers; and means for heating the chambers and/or the fluid.
  • a method for automatically processing a sample comprising: inserting a solid support bearing a sample to be processed and/or reagent into one of a plurality of support blocks such that the support block and the solid support form a substantially enclosed chamber; and processing a sample within the chamber using a predetermined protocol comprising steps involving the introduction of fluid into the chamber, the removal of fluid from the chamber, and the heating of the chamber and/or the fluid.
  • the solid support bearing the sample to be processesd and/or reagent is preferably in the form of a solid support bearing an immobilised material which is capable of interacting with a fluid introduced into the chamber.
  • the immobilised material may comprise DNA, protein or protein recognition units e.g.
  • the immobilised material may also be a tissue section.
  • the fluid which is introduced into the chamber and which is capable of interacting with the immobilised material may contain a biological sample e.g. a tissue or a cell extract. Substances present in the biological sample will prefeably bind or hybridise to the immobilised material. Binding or hybridisation may be visualised in a processed by staining or other methods such as radiolabelling or chemical labelling, either before or after the binding step. Biotinylation, fluorescence, chemiluminescence or other labelling techniques may also be used. Analysis of the resultant binding pattern may be undertaken using an appropriate detection system such as optical or laser scanning technology and a comparison of results between, for example, drug treated and untreated control tissues can be undertaken using suitable computer software.
  • a biological sample to be processed using the method of the invention will preferably be from a mammal treated with a chemical compound of therapeutic potential.
  • the method of the invention provides a fast, reproducible and robust methods for determining e.g. nucleotide or protein expression pattern of tissues or cell types, i.e. genomics or proteomics.
  • the method may be applied in, for example, toxicology, clinical trials, process monitoring, biotechnology and pharmaceutics, either as a research or diagnostic tool.
  • the method may be used for analyzing the spatial and temporal changes in the expression of proteins in mammalian tissues and cell types when exposed to environmental stimuli such as pharmaceutical reagents, as well as other environmental stimuli or factors.
  • the method allows for the comparison of targets treated with therapeutic drugs and untreated controls to provide a quantitative and/or semi-qualitative analysis of treated and untreated (control) target systems e.g. cells, tissues or biological organisms.
  • the method may also be adapted for use in, for example, drug manufacturing, purity analysis (e.g. in food and agriculture) and for diagnostics, e.g. for detecting proteins in urine or blood.
  • the invention comprises a method for automatically processing a microarray comprising: inserting a solid support bearing a microarray into one of a plurality of support blocks such that the support block and the solid support form a substantially enclosed chamber; and processing the microarray within the chamber using a predetermined protocol comprising steps involving: heating the microarray; introducing fluid into the microarray comprising a biological sample capable of interacting with the microarray; removal of the fluid from the chamber; washing the microarray; and drying the microarray.
  • the apparatus of the invention may contain any number of support blocks from two upwards.
  • the apparatus preferably comprises at least five support blocks.
  • the support blocks may be present in, for example, banks of 5, 8 or 10 blocks. It will be readily apparent to those skilled in the art how further blocks may be added to the apparatus.
  • the support blocks used in the invention may wholly enclose the solid support, e.g. a glass slide, bearing the sample to be processed and/or reagent, i.e. the support block provides an enclosed, fluid tight container for the solid support.
  • the support block preferably comprises top and bottom members which may be separated, thus allowing easy insertion and removal of the solid support.
  • part of the internal surface of the substantially enclosed chamber is defined by the sample and/or reagent bearing surface of the solid support.
  • the support block preferably comprises top and bottom members which may be separated, thus allowing easy insertion and removal of the solid support.
  • the support block comprises top and bottom members they preferably comprise sealing means to assist in the provision of an enclosed, fluid tight chamber.
  • the sealing means may comprise clamping means such as screws.
  • the sealing of the block may be further assisted by the provision of a gasket, e.g. an O-ring, between the top and bottom members, or between one or both of the top and bottom members and the solid support, this gasket may be made of any resilient material that is inert to the processing conditions employed, e.g. silicon rubber.
  • a gasket e.g. an O-ring
  • the support block is preferably substantially rectangular in cross section, however other shapes may be employed depending on the configuration of the solid supports.
  • the support blocks may be made from any substantially rigid material that is inert to the processing conditions employed, for examples metal or plastics.
  • the means for introducing fluid into the chambers and removing fluid from the chambers preferably comprise a fluid inlet and a fluid outlet in the body of the sample block.
  • the fluid inlet and fluid outlet will be in contact with the interior of the chamber in use.
  • the apparatus preferably comprises fluid storage means in connection with the means for introducing fluid into the chambers, the storage means being adapted to contain the fluid that is introduced into the chambers during processing.
  • the fluid inlet is preferably interchangeably connected to a plurality of fluid storage means thus allowing several fluids to be used in the processing of the sample. Fluids used in processing of the sample include liquids and gases.
  • the apparatus also preferably contains means for directing fluid that is removed from the chamber to waste.
  • the configuration of the means for introducing fluid into the chambers and removing fluid from the chambers are preferably such that a fluid can be repeatedly circulated through the chamber.
  • a means of circulating fluid over the solid support is particularly advantageous when the processing involves the hybridisation of a sample to a microarray since it results in more uniform hybridisation than can be achieved when the fluid containing the hybridisation sample is static.
  • the circulation of fluid through the chamber may continue for e.g. up to 24 hours, suitably for 2-16 hours.
  • the rate of circulation of fluid is preferably controllable, fluid is preferably circulated slowly.
  • Fluid connections used in the apparatus of the invention e.g. tubing connecting the means for introducing fluid into the chambers/removing fluid from the chambers and the fluid storage means/waste, is preferably narrow bore in order to minimise the quantity of reagent/samples/fluids that are required in the sample processing. Fluids are preferably introduced and removed from the chambers by positive pressure, e.g. a pumping mechanism.
  • Fluids that are typically used in the process of the invention include hot water which is used to prepare solid supports, e.g. glass slides, solvents used for support preparation, samples, wash solutions and gases, e.g. air for drying solid supports received in the support blocks.
  • the solid supports are preferably dried in a manner which does not leave any residue or "streaks" on supports, such that e.g. in the processing of microarrays there is no interference with the imaging of the spots.
  • the solid supports are preferably dried using a flow of gas e.g. air, the gas may be at an elevated temperature e.g. about 50°C.
  • the support blocks and the means for introducing fluid into the chambers and removing fluid from the chambers are preferably maintained in a thermally controlled environment. This is particularly advantageous when the processing of the sample involves the circulation of sample/reagent through the chamber, since no cooling/heating of the sample will occur when it is outside the chamber.
  • fluids may be introduced into the chamber at a chosen, e.g. elevated temperature.
  • the apparatus is preferably adapted to allow easy cleaning such that there is no risk of contamination between different runs of the apparatus.
  • the solid supports preferably bear microarrays of reagents which are used in the processing of a sample, and the sample is introduced as a fluid into the chamber.
  • the various steps involved in the processing of a sample according to the invention can preferably be programmed e.g. as to time and temperature. Different support blocks in the apparatus according to the invention can preferably be controlled independently thus allowing different sample processing protocols to be used simultaneously in a single apparatus.
  • the method of the invention is preferably performed under computer control.
  • the computer control means may comprise a personal computer of may be integrated with the apparatus adapted to perform the claimed method.
  • the computer control means preferably controls one or more of the following parameters: temperature, fluid volume and flow rate and timing of various stages in the sample processing.
  • a typical automated procedure which may be performed using the apparatus of the invention is as follows: 1. Insert slide into support chamber.
  • Figure 1 is a schematic representation of an apparatus according to the invention, comprising five support blocks and illustrating the status of fluid inlets and outlets during a sample processing protocol similar to that described above.
  • Figure 2 shows a plan view of the inner surface, end and cross-sectional views of the bottom member of a two member support block (la) of the type wherein in use part of the internal surface of the substantially enclosed chamber is defined by the sample and/or reagent bearing surface of the solid support.
  • a fluid inlet and fluid outlet (2a, 2b) are shown together with a groove (3) for retaining a O-ring used to enhance the seal between the support block and solid support.
  • Figure 3 shows a plan view of the outer surface, a plan view of the inner surface and a cross-sectional view of the top member of a two member support block (lb) of the type wherein in use part of the internal surface of the substantially enclosed chamber is defined by the sample and/or reagent bearing surface of the solid support.
  • the inner surface and the cross-section show a flange (4) adapted to retain the solid support.
  • Four clamping screws (5) are illustrated in the view of the outer surface.
  • Figure 4 shows side and top views of a clamping screw for use with the support block of Figures 2 and 3.
  • Figure 5 shows a perspective view of the assembled support block (1) comprising bottom (la) and top members (lb), and a solid support (6) held in position in the block.
  • the O-ring which is not visible in this figure forms a seal between the sample and/or reagent bearing surface of the solid support and the inner surface of the bottom member of the support block (la).
  • Figure 6 shows a photograph of a single assembled support block and solid support during the processing of a sample.
  • Figure 7 shows a perspective view of an apparatus according to the invention, comprising ten support blocks.
  • Figures 8, 9 and 10 are visual inspection images of microarrays produced in Example 1, viz: Figure 8 - 600 ⁇ l, Without accelerator, control manual method; Figure 9 - 600 ⁇ l, Without accelerator, automated method of invention; Figure 10 - 600 ⁇ l, With accelerator, automated method of invention.
  • Figure 11 shows the correlation between the right hand side (RHS) and left hand side (LHS) of the 600 ⁇ l without accelerator, control manual slide.
  • Figure 12 shows the correlation between the RHS and LHS of the 600 ⁇ l without accelerator, automated method of the invention slide.
  • Figure 13 shows the correlation between the RHS and LHS of the 600 ⁇ l with accelerator, automated method of the invention slide.
  • Figure 14 is a bar chart representing the solidity of the average spot as a percentage of the maximum solidity value for the three tests. Accompanying this bar chart is the relevant data.
  • Figure 15 is a bar chart representing the Roundness of the average spot as a percentage of the maximum Roundness value for the three tests. Accompanying this bar chart is the relevant data.
  • Figure 16 is a bar chart representing the Circularity of the average spot as a percentage of the maximum Circularity value for the three tests. Accompanying this bar chart is the relevant data.
  • Figure 17 is a bar chart representing the average Artefact levels of each spot.
  • cDNA was reverse transcribed for all three experiments (lOug total R A per hybridisation).
  • the cDNA was made from rat liver and fluorescently labelled during the transcription reaction by a method similar to that described in DeRisi et al, 1997, Science, 278, 680-686, except that Cy3 was replaced with Alexa 546.
  • the hybridisation solution was made up as follows: labelled cDNA lOO ⁇ l
  • the hybridisation buffer used was lOxSSC, 0.2% SDS.
  • the accelerator used was 60% polyethylene glycol (PEG). BDH Laboratory Supplies.
  • Hybridisation and wash conditions 600ul of hybridisation solution (see above) was heated to 100°C for 3 minutes before pipetting into the hybridisation chamber (see figures). Using the manual method the chamber was sealed and rotated overnight in an incubator for 16 hours at 50°C. The slide was then removed from the chamber and washed for 10 minutes in 2 x SSC (Sigma), 0.1% saline followed by a 5 minute wash in 0.1% SSC, 0.1% saline (both at 40°C ). The slide was centrifuged in order to remove any traces of buffer.
  • 2 x SSC Sigma
  • 0.1% saline followed by a 5 minute wash in 0.1% SSC, 0.1% saline (both at 40°C ).
  • the slide was centrifuged in order to remove any traces of buffer.
  • Bottles of the appropriate reagents were replenished, including wash buffers (see above for composition) and boiling water.
  • Freshly printed slides were assembled in hybridisation chambers on the support block of the automated machine. Sample was added to the sample block and the slides were automatically pre-rinsed, hybridised, washed and dried according to the following protocol:
  • the 'Final Wash' step incorporates two washes (low and high stringency) both run for the time stated.
  • the temperature was kept at 50°C for the duration of the experiment, but the sample holding block was heated to 80-100°C before sample was applied to the slide.
  • Imaging was carried out using a Molecular Dynamics Generation III Scanner (Amersham Pharmacia Biotech, UK) at 550-600nm. Images were analysed using a proprietary software package that provides data comprising: spot volume (with and without dust/artefact) spot circularity (a measure of how even the circumference of the spot is) spot roundness (a measure of how misshapen the spot is) spot solidity (a measure of how even the signal intensity is across the spot) These criteria were examined in order to compare the total signal, the reproducibility of signal, the quality of hybridisation to spots and the amount of artefact in the hybridisations.
  • FIG. 10-12 show the correlations of left and right hand sides of the slides for the manual method and the automated method (with and without accelerator). The correlations are 0.59, 0.76 and 0.95 respectively, indicating that the automated method increases the reproducibility of duplicate spots within a slide.
  • Spot quality Figures 13-15 show graphs of the roundness, circularity and solidity of spots. These criteria can be used to measure spot quality, since rounder, more circular and even spots facilitate image analysis and indicate even hybridisation. These criteria were measured for each spot and then averaged for one side of a microarray slide. In all cases the automated method gave rise to more solid, more circular, rounder spots than the manual method. The addition of accelerator also improved these results.
  • the processed slides have a low background and there is no smearing or streaking of wash solutions.
  • the automatically processed slides are easier to analyse.
  • This method of the invention therefore offers improvements in time, labour, reproducibility, quality and accuracy of microarray experiments when compared to manual methods of sample processing.

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  • Physics & Mathematics (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Pathology (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)
  • Sampling And Sample Adjustment (AREA)

Abstract

L'invention concerne un appareil automatisé de traitement d'échantillons, qui comporte plusieurs blocs supports, chacun étant adapté pour recevoir un support solide portant l'échantillon à traiter et/ou un réactif, de façon qu'en cours d'utilisation, le bloc support et le support solide forment une chambre sensiblement fermée. L'appareil comporte également un dispositif d'introduction de fluide dans la chambre et d'extraction de fluide de la chambre, et un dispositif permettant de chauffer la chambre et/ou le fluide, ainsi qu'un procédé de traitement d'échantillons reposant sur l'utilisation du présent appareil.
PCT/GB2000/004142 1999-10-29 2000-10-27 Appareil modulaire automatisé de traitement d'échantillons WO2001031347A1 (fr)

Priority Applications (3)

Application Number Priority Date Filing Date Title
JP2001533433A JP2003513246A (ja) 1999-10-29 2000-10-27 モジュールの自動サンプル処理装置
EP00971597A EP1228375A1 (fr) 1999-10-29 2000-10-27 Appareil modulaire automatic de traitement de chantillons
AU10431/01A AU1043101A (en) 1999-10-29 2000-10-27 Modular automated sample processing apparatus

Applications Claiming Priority (2)

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GB9925679.4 1999-10-29
GBGB9925679.4A GB9925679D0 (en) 1999-10-29 1999-10-29 Novel process and apparatus

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WO2001031347A1 true WO2001031347A1 (fr) 2001-05-03

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EP (1) EP1228375A1 (fr)
JP (1) JP2003513246A (fr)
AU (1) AU1043101A (fr)
GB (1) GB9925679D0 (fr)
WO (1) WO2001031347A1 (fr)

Cited By (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2003029845A3 (fr) * 2001-10-01 2003-05-15 Vision Biosystems Ltd Traitement histologique d'echantillons tissulaires
WO2003106033A1 (fr) * 2002-06-13 2003-12-24 Millenium Biologix Ag Chambre de reaction
EP1403644A1 (fr) * 2002-09-25 2004-03-31 Agilent Technologies Inc. a Delaware Corporation Dispositif d'analyse de fluides
WO2004072620A1 (fr) * 2003-02-06 2004-08-26 Applera Corporation Dispositifs et procedes de preparation d'echantillons biologiques
WO2004108268A1 (fr) * 2003-05-30 2004-12-16 Applera Corporation Appareil et procede d'hybridation et de detection spr
EP1605245A3 (fr) * 2004-06-08 2006-07-19 Agilent Technologies, Inc. Méthode de lavage pour matrices d'échantillons
US7517498B2 (en) 2003-08-19 2009-04-14 Agilent Technologies, Inc. Apparatus for substrate handling
US8554372B2 (en) 2003-09-29 2013-10-08 Leica Biosystems Melbourne Pty Ltd System and method for histological tissue specimen processing
DE102014001481A1 (de) * 2013-10-28 2015-04-30 Euroimmun Medizinische Labordiagnostika Ag Verbesserte Vorrichtung und Verfahren für Reaktionen zwischen einer festen und einer flüssigen Phase

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5139743A (en) * 1987-09-24 1992-08-18 Fuji Photo Film Co., Ltd. Biochemical analysis apparatus
US5382511A (en) * 1988-08-02 1995-01-17 Gene Tec Corporation Method for studying nucleic acids within immobilized specimens
US5451500A (en) * 1989-11-17 1995-09-19 Gene Tec Corporation Device for processing biological specimens for analysis of nucleic acids

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5139743A (en) * 1987-09-24 1992-08-18 Fuji Photo Film Co., Ltd. Biochemical analysis apparatus
US5382511A (en) * 1988-08-02 1995-01-17 Gene Tec Corporation Method for studying nucleic acids within immobilized specimens
US5451500A (en) * 1989-11-17 1995-09-19 Gene Tec Corporation Device for processing biological specimens for analysis of nucleic acids

Cited By (13)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7371346B2 (en) 2001-10-01 2008-05-13 Leica Biosystems Melbourne Pty Ltd Histological tissue specimen treatment
WO2003029845A3 (fr) * 2001-10-01 2003-05-15 Vision Biosystems Ltd Traitement histologique d'echantillons tissulaires
US8394322B2 (en) 2001-10-01 2013-03-12 Leica Biosystems Melbourne Pty Ltd Histological tissue specimen treatment
WO2003106033A1 (fr) * 2002-06-13 2003-12-24 Millenium Biologix Ag Chambre de reaction
EP1403644A1 (fr) * 2002-09-25 2004-03-31 Agilent Technologies Inc. a Delaware Corporation Dispositif d'analyse de fluides
WO2004072620A1 (fr) * 2003-02-06 2004-08-26 Applera Corporation Dispositifs et procedes de preparation d'echantillons biologiques
KR100706464B1 (ko) * 2003-05-30 2007-04-10 애플라 코포레이션 혼성화 및 에스피알 검출을 위한 장치 및 방법
WO2004108268A1 (fr) * 2003-05-30 2004-12-16 Applera Corporation Appareil et procede d'hybridation et de detection spr
US7517498B2 (en) 2003-08-19 2009-04-14 Agilent Technologies, Inc. Apparatus for substrate handling
US8554372B2 (en) 2003-09-29 2013-10-08 Leica Biosystems Melbourne Pty Ltd System and method for histological tissue specimen processing
EP1605245A3 (fr) * 2004-06-08 2006-07-19 Agilent Technologies, Inc. Méthode de lavage pour matrices d'échantillons
DE102014001481A1 (de) * 2013-10-28 2015-04-30 Euroimmun Medizinische Labordiagnostika Ag Verbesserte Vorrichtung und Verfahren für Reaktionen zwischen einer festen und einer flüssigen Phase
US9977040B2 (en) 2013-10-28 2018-05-22 Euroimmun Medizinische Labordiagnostika Ag Device and method for reactions between a solid and a liquid phase

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JP2003513246A (ja) 2003-04-08
EP1228375A1 (fr) 2002-08-07
AU1043101A (en) 2001-05-08
GB9925679D0 (en) 1999-12-29

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