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WO2015079048A1 - Système d'électrophorèse - Google Patents

Système d'électrophorèse Download PDF

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Publication number
WO2015079048A1
WO2015079048A1 PCT/EP2014/076019 EP2014076019W WO2015079048A1 WO 2015079048 A1 WO2015079048 A1 WO 2015079048A1 EP 2014076019 W EP2014076019 W EP 2014076019W WO 2015079048 A1 WO2015079048 A1 WO 2015079048A1
Authority
WO
WIPO (PCT)
Prior art keywords
electrophoresis
cassette
gel
imaging
separation
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.)
Ceased
Application number
PCT/EP2014/076019
Other languages
English (en)
Inventor
Owe Salven
Stefan Sjolander
Tomas Agren
Hakan Erk ROOS
Tomas Haukkala
Urban Jonsson Axelsson
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.)
Global Life Sciences Solutions USA LLC
Original Assignee
GE Healthcare Bio Sciences Corp
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 GE Healthcare Bio Sciences Corp filed Critical GE Healthcare Bio Sciences Corp
Publication of WO2015079048A1 publication Critical patent/WO2015079048A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G01N27/416—Systems
    • G01N27/447—Systems using electrophoresis
    • G01N27/44704—Details; Accessories
    • G01N27/44717—Arrangements for investigating the separated zones, e.g. localising zones
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/34—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving hydrolase
    • C12Q1/37—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving hydrolase involving peptidase or proteinase
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G01N27/416—Systems
    • G01N27/447—Systems using electrophoresis
    • G01N27/44704—Details; Accessories
    • G01N27/44717—Arrangements for investigating the separated zones, e.g. localising zones
    • G01N27/44721—Arrangements for investigating the separated zones, e.g. localising zones by optical means

Definitions

  • the present invention relates to an electrophoresis system for electrophoresis experiments, and more particularly to a combined electrophoresis and scanning system for automated electrophoresis experiments and scanning using electrophoresis cassettes.
  • Electrophoresis is a commonly used method for analysis, wherein charged molecules and particles migrate in a separation medium, usually a gel, which is subjected to an electrical field between two electrodes. Separation of proteins may be by isoelectric point (pi), molecular weight, electric charge, or a combination of these factors.
  • the separation gel is usually placed on a support and two opposing ends of the gel are contacted with an electrode buffer in solution or rigid form.
  • the electrodes may be inserted in vessels containing the electrode buffers.
  • the buffer solutions from both the electrolytic medium and a reservoir for ions to keep the pH and other parameters constant.
  • the molecules are detected and identified in different manners: e.g. visually by staining the gel or by optical means such as scanning or imaging the stained gel or labelled samples by a laser scanner or the like.
  • Gel electrophoresis is today routinely used for separating biomolecules such as proteins, peptides, nucleic acids etc. Samples are handled in different types of screening, identifying (cell signaling, expression & purification) or in clinical tests. Protein samples can derivate from e.g. human, mammalian tissue, cell lysates or bacterial, insect or yeast cellular systems. The electrophoretic conditions for different types of molecules are different and have to be adapted in many cases. Thus, both the gel and the buffer solutions must often be chosen for each type of sample.
  • the preparation of the electrophoresis process includes several rather laborious steps.
  • a suitable gel is chosen and placed or molded on a support. The gel is contacted with the buffer solutions.
  • a common way is to have a gel slab in a cassette of glass or plastic in contact with the buffer solutions in buffer tanks. For each run the gel has to be placed on the support or the cassette be prepared. Then the buffer tanks are filled with buffer solutions and the samples are applied on the gel.
  • WO 87/04948 to incorporate the buffer substance in a gel material whereby the buffer is obtained in the form of a buffer strip.
  • US6368481 discloses a precast electrophoresis cassette wherein buffer strips are incorporated as an integral part of the cassette.
  • the proteins may be transferred to a membrane (typically nitrocellulose or PVDF), where they are probed (detected) using antibodies specific to the target protein, a process commonly referred to as western blotting or immunoblotting.
  • a membrane typically nitrocellulose or PVDF
  • the primary method for transferring the proteins to the membrane is referred to as electroblotting and uses an electric current to pull proteins from the gel into the membrane. The proteins move from within the gel onto the membrane while maintaining the
  • the proteins bind to the surface of the membrane due to its nonspecific protein binding properties (i.e. binds all proteins equally well). In order to avoid unspecific binding of probing antibodies, remaining binding sites on the membrane may be blocked.
  • the membrane with the transferred proteins is incubated with specific primary antibody directed towards the protein of interest and secondary antibody e.g. for the protein of interest with a modified antibody which is linked to a reporter enzyme; when exposed to an appropriate substrate this enzyme drives a colorimetric reaction and produces a colour or by fluorescently labelled targets (dyes), that may be detected by a suitable imaging technique.
  • specific primary antibody directed towards the protein of interest and secondary antibody e.g. for the protein of interest with a modified antibody which is linked to a reporter enzyme; when exposed to an appropriate substrate this enzyme drives a colorimetric reaction and produces a colour or by fluorescently labelled targets (dyes), that may be detected by a suitable imaging technique.
  • Electrophoresis and the following blotting step is traditionally characterized by a lot of manual handling of both gels and membranes, as well as a range of liquids, e.g. buffers, reagents, wash solutions etc.
  • Some attempts to facilitate and/or automate the workflow have been made in the past, but there are very few US5674006 discloses one example of an apparatus for efficiently circulating and moving a fluid across a workpiece.
  • the apparatus can provide for the automated handling of the fluids used, and is well suited for use in the staining and fixing of biological assays such as electrophoresis gels.
  • the object of the invention is to provide a new electrophoresis system, which electrophoresis system overcomes one or more drawbacks of the prior art. This is achieved by the electrophoresis system as defined in the independent claim.
  • electrophoresis system provides for high resolution electrophoresis experiments and imaging in one combined unit.
  • Another advantage is that it is capable of providing high quality imaging of immunoblotting membranes, and wherein acquired images are aligned with corresponding electrophoresis gel images.
  • Fig 1 is a schematic perspective view of an electrophoresis cassette according to one embodiment.
  • Figs. 2a to 2f show components of the electrophoresis cassette of fig. 1.
  • Fig 3 shows an electrophoresis gel unit with a gel member attached to the top face of a support frame.
  • Fig. 4 shows an enlarged section of the lower end of the cassette according to one embodiment.
  • Figs 5 a to 5 c illustrates the step of preparing the cassette for performing an electrophoresis experiment.
  • Figs. 6a and 6b show perspective views of an electrophoresis tray that is compatible with the electrophoresis cassette for running electrophoresis experiments.
  • Figs 7a to 7c show a schematic view of one embodiment of a buffer pad unit for use with an electrophoresis tray of fig. 6.
  • Figs 8a to 8f show schematic views of the interaction between the
  • electrophoresis tray the buffer pad units and an electrophoresis cassette.
  • Figs. 9a to 9e show another embodiment of a buffer pad unit.
  • Figs 10a to lOf show schematic cross-sectional views of the interaction between the electrophoresis tray, the buffer pad units and an electrophoresis cassette.
  • Figs. 1 la to 11c show an example of an electrophoresis system
  • Fig 12 shows details of one embodiment of an electrophoresis system.
  • Fig. 13 shows another example of an electrophoresis system
  • Figs 14a to 14d show one embodiment of a sealing member in accordance with one embodiment.
  • Fig 15 shows a membrane unit for immunob lotting
  • Figs. 16a to 16d disclose one embodiment of a membrane unit holder, arranged to hold a membrane unit in position for imaging. .
  • Figs 17a- 17c show a schematic cross sectional view of the membrane unit holder
  • Figs. 18a and 18 b disclose one embodiment of a support surface of
  • Fig. 19 shows another embodiment of a buffer pad unit.
  • the separation-zone of an electrophoresis gel is defined as the part of the gel wherein the separated species of the sample are located after a completed electrophoresis run.
  • PCT/SE2013/050633 PCT/SE2013/050634, PCT/SE2013/050635 and
  • Fig. 1 is a perspective view of an electrophoresis cassette 10 according one schematic embodiment.
  • the cassette 10 comprises a cassette housing 20, a detachable gel support frame 30, a section-wise removable backing film 40 and a removable sample well cover 50.
  • Fig. 1 shows the electrophoresis cassette in assembled state.
  • the gel cassette 10 defines therein a gel compartment for molding a flat gel member 36 for electrophoretic separation.
  • the electrophoresis cassette 10 is a precast cassette, but alternatively, the cassette 10 may be empty and ready for molding of a custom gel in the gel compartment, e.g. by the end customer.
  • Figs. 2a and 2b shows the cassette housing 20 with the other components of the cassette 10 removed.
  • Fig. 2a is a top view whereas fig. 2b shows the cassette housing 20 from below.
  • the cassette housing 20 is generally comprised of a thin upper wall 60 with an upper face 65 and a lower face 66, and a rim 70 that projects downwards from the upper wall 60 around its periphery with a bottom face 80 and an inner wall 75.
  • the lower face 66 of the upper wall 60 and the inner wall 75 of the rim 70 essentially defines the gel compartment, which may be closed from below by attaching the support frame 30 and the removable backing film 40 to the lower face 80 of the rim 70, as is shown in fig 1 and will be discussed in more detail below.
  • the thickness of a gel member 36 (as schematically disclosed in fig. 3), molded in the cassette 10 will be essentially the same as the height of the inner wall 75 of the rim.
  • the upper wall 60 is of uniform thickness whereby the gel member 36 also will be of uniform thickness, provided that the support frame 30 and the removable backing film 40 are flat as in the disclosed embodiment.
  • the thickness of the gel is preferably adapted to the specific gel type and the buffer system used, as well on the desired currents involved in the electrophoresis step.
  • features of the cassette housing 20 may be formed to provide for a gel member 36 of different thickness in different sections thereof.
  • the cassette housing 20 should provide a rigid structure to the cassette 10 during storage and use, it should be made of a suitably rigid material. Moreover, as will be disclosed in detail below, the cassette 10 is designed for running electrophoresis separation, therefore the cassette housing 20 should be electrically insulating.
  • the cassette material may be selected so as to not essentially degrade or get discolored by UV radiation in doses corresponding to polymerization. Moreover the cassette material may be selected so as to not hinder polymerization of the gel, and depending on the design of the cassette 10 the material may be selected so as to exhibit a suitable adhesion to the gel, e.g.
  • the cassette 10 is further designed to be used in a combined electrophoresis and fluorescence imaging apparatus wherein the gel member 36 may be imaged during or after the electrophoresis step while still in the cassette, as will be disclosed in detail below. Therefore, at least the section of the upper wall 60 covering the separation-zone of the gel member 36 should be sufficiently transparent to electromagnetic radiation of relevant wavelengths.
  • the whole cassette housing 20 is injection molded in the same material. Moreover, all components of the cassette 10 may be selected so as to be non/low fluorescent.
  • the cassette housing 20 is made of a rigid polymer, such as Cyclo Olefin Polymer (COP), Cyclic Olefin Copolymer (COC), polypropylene (PP), Polyethylene terephthalate (PET) , polycarbonate, polymethyl methacrylate (PMMA), combinations, variants thereof or the like.
  • COP Cyclo Olefin Polymer
  • COC Cyclic Olefin Copolymer
  • PP polypropylene
  • PET Polyethylene terephthalate
  • PMMA polymethyl methacrylate
  • a transverse wall 90 arranged to divide the gel compartment into an electrophoresis compartment and an over- fill chamber 100 arranged to receive excess gel solution during the step of molding the gel member 36.
  • a fill port 120 at the opposite end of the electrophoresis compartment with respect to the over- fill chamber 100, and an air vent 130 in the over- fill chamber 100.
  • the disclosed cassette 10 is provided with 16 sample well openings 1 10 for enabling loading of sample onto the gel member 36 for separation, each sample well opening 110 corresponding to one electrophoresis lane during separation.
  • the number and shape of sample well openings 110 may vary depending on the actual dimensions of the electrophoresis cassette, the type of separation and the electrophoresis gel type etc.. There may be any suitable number of sample well openings 110 between 1 and e.g. 100.
  • the cassette is provided with one wide sample loading opening extending essentially across the full width of the gel member, replacing the individual sample well openings.
  • the user may e.g form wells directly in the gel using a well-comb or the like, or there may be provided one or more sample loading cups that may be attached to the cassette 10 in contact with the gel member 36 for providing a flexible number of separation lanes.
  • the sample well openings 110 are covered by a removable sample well cover 50 which is disclosed in more detail in figs 2c and 2d.
  • the sample well cover 50 is arranged to fit over the well openings 110 and to keep them closed during the molding process and storage. Before sample is to be loaded into the sample wells 110, the well cover 50 is removed to open the sample wells 110.
  • the well cover 50 comprises well forming protrusions 52 that are formed to fit in a mating relationship in the sample well openings 110 to essentially provide a sealing interaction therewith to avoid leakage of gel solution during molding and air into the cassette during storage.
  • the well forming protrusions 52 are designed to extend below the lower face 66 of the upper wall 60 into the gel member 36 to form sample wells extending into the gel member 36 when removed.
  • the well forming protrusions 52 are designed such that they are flush with the lower face 66 of the upper wall 60 to provide an essentially flat surface of the gel member 36 and wherein sample wells are formed by the sample well openings 110.
  • the sample well cover 50 is arranged to seal against the upper face 65 of the upper wall 60 or a combination thereof. To facilitate removal while providing sufficiently efficient sealing, the sample well cover 50 is made of a suitable elastic material, such as e.g.
  • the sample well cover 50 may be co-molded with the cassette housing 20, such that the cassette housing 20 is molded in a first step in a first rigid material where after the sample well cover 50 is molded in a second step in a second, elastic material where the cassette housing partly acts as mold.
  • the co-molded sample well cover 50 can be made selectively removable.
  • an intermediate material providing for suitable adhesion characteristics may be provided in between the cassette housing 20 and the sample well cover 50, e.g. a thermoplastic material with low melting temperature or the like.
  • the detachable gel support frame 30 is detachably attached to the bottom face 80 of the rim 70 and the section-wise removable backing film 40 is in turn attached/laminated to the bottom of the gel support frame 30.
  • the gel support frame 30 and the backing film 40 together provide a lower wall that closes the electrophoresis compartment and the over- fill chamber 100 for molding and storage.
  • the disclosed embodiment of the gel support frame 30 comprises two buffer buffer-slits 150a and 150b and a separation zone window 160 each covered from below by a respective removable section 210a-c of the backing film 40, shown in fig. 2f.
  • the backing film may 40 be laminated onto the bottom face of the gel support frame 30 such that the respective sections 210a-c can be removed e.g. by an operator grabbing and pulling a respective peel tab 21 la-c.
  • the sections 210a and 210b of the backing film 40 are removed in order to place the gel in contact with respective buffer sources, e.g. buffer pads in an electrophoresis apparatus.
  • the section 210b is removed to uncover the bottom face of a gel member 36 through the separation-zone window 160.
  • At least the sections of the film 40 being in direct contact with the gel should have sufficiently low surface adhesion with the same.
  • Low surface adhesion may be achieved by selecting suitable material and surface properties for the whole film and/or modifying the surface properties at the specific interaction-zones, e.g. surface roughness, surface coating, laminating other material to said zones or the like.
  • the gel support frame 30 is comprised of a rigid polymer film with adhesive layers applied to both faces thereof and the backing film 40 is comprised of a plain polymer film bonded to the rigid polymer film by the adhesive layer.
  • the adhesive layer on the housing side of the gel support frame 30 may be arranged to provide a removable but essentially airtight bond to the cassette housing 20, and to provide a high gel adhesion compared to the gel adhesion of the cassette housing 20 and the gel adhesion of the polymer film of the backing film 40.
  • the rigid polymer film of the of the gel support frame 30 may be a Polyethylene terephthalate (PET) film with adhesive layers applied to both faces in the form of a melt-adhesive, such as ethylene- vinyl acetate (EVA) or another adhesive with suitable properties for pealable bonding
  • the polymer film of the backing film 40 may be a PET film.
  • the support frame 30 with the adhesive layers only covers parts of the gel member 36 that do not need to be accessible from the bottom thereof and hence has openings which correspond to the respective removable sections of the backing film 40.
  • the support frame 30 has a thin adhesive layer of a material, e.g EVA or another pealable adhesive, which melts at a lower temperature than the PET foil itself and hence the backing film 40 and the support frame 30 can be laminated together using a heat lamination process and finally releasably attached to the cassette housing 20 by a heat bonding process or the like. It has been experimentally verified that an EVA layer meet the crucial property of high gel adhesion for tested gel compositions, without disturbing the gel polymerization or other characteristics which are necessary in this concept.
  • EVA e.g EVA or another pealable adhesive
  • the stack of foils is laminated at approximately 100 - 115°C and this procedure should result in a flat, not creased or wrinkled, foil.
  • the backing film 40 may be thick enough to give a stable feeling, i.e. not too elastic or flimsy, but also thin enough to allow cooling during electrophoresis as will be disclosed in more detail below.
  • the backing film 40 may be from e.g. 0.1 to 0.4 mm of thick or any value there between depending on the material of the film.
  • Adhesion to cassette must be strong enough to prevent leakage but must also allow opening of the foil by hand with little force.
  • the gel support frame 30 is designed to stay attached to the gel member 36 after removal from the cassette 10.
  • the support frame 30 is formed of a suitably rigid material to preserve the shape of the gel and to facilitate handling of the gel member 36 by providing accessible gripping portions that are not covered by the gel member. After removal of the section 210c of the backing film 40 the lower face of the separation zone of the gel member 36 is accessible through the separation-zone window 160.
  • the gel member 36 In order to achieve proper attachment of the gel member 36 to the support frame 30 it should be designed with high surface adhesion to the gel member. This may be achieved by selecting suitable material properties and/or by surface modification e.g. surface roughness, surface coating or the like as discussed above.
  • the support frame 30 is attached to the bottom face 80 of the rim 70 such that it is easily detachable, but still provides adequate sealing around the rim 70 to keep the gel compartment sealed during molding and storage. This may e.g. be achieved by selection of suitable material parameters and e.g. use of adhesive, or heat welding.
  • the cassette housing 20 is made of a rigid polymer and the support frame 30 of a rigid polymer film rigid polymer film with adhesive layers applied to both faces.
  • the support frame 30 is provided with at least one peel tab 170 for pulling the support frame 30 to detach it from the cassette housing 20 together with the gel member.
  • the support frame 30 comprises one or more reinforcement layers (not shown) at exposed sections, like peel tabs or the like.
  • At least the inner walls of the cassette housing 20 should have low surface adhesion with the gel.
  • Low surface adhesion may be achieved by selecting suitable material and surface properties for the whole film and/or modifying the surface properties, e.g. low surface roughness, surface coating, or the like as discussed above.
  • the shape of certain features in the gel compartment may be designed to avoid attachment of the gel thereto to further facilitate release of the gel member, e.g. rounded corners, non-vertical walls and openings etc.
  • the support frame 30 further comprises an alignment tag 180 with a predefined alignment structure defining a positional reference for alignment of the support frame 30.
  • the alignment structure is provided in the form of two alignment holes 190a and 190b, arranged to ensure that the cassette 10 and/or the support frame 30 is properly aligned with respect to a complementary alignment structure e.g. comprising 2 pins, in an electrophoresis apparatus or the like.
  • a complementary alignment structure e.g. comprising 2 pins
  • the alignment structure 190a-b is provided as a part of the support frame 30 to which the gel member 36 is attached also after the electrophoresis run and in the following transfer step, repeatable positioning of the gel may be achieved which may be very valuable in many situations as will be disclosed in more detail below.
  • the alignment structure may be asymmetrical in a way that it can only be fitted into a complementary alignment structure of an instrument or the like in one unique orientation, whereby, it cannot be inserted in the wrong way, upside down or the like.
  • the support frame 30 is suitably provided with an identification code 200 or the like which will make it possible to read the identity of the gel member 36 also after it has been removed from the cassette 10 in a secure way.
  • the identification code 200 may e.g. be a machine readable code as a bar-code, matrix-code or the like, and provide the user and/or instruments with relevant information.
  • the gel support frame 30 is comprised of a rigid film of an electrically insulating material, e.g of a polymer material.
  • the term rigid refers to the film being much more rigid compared to the gel, and especially in the plane to avoid distortion of the gel outline.
  • the film may be quite flexible and bendable in other directions (which is a common characteristic for a film) and it should not be brittle, as it has to be possible to release the gel member 36 from the cassette housing by pulling the peal tab 170 of the support frame 30.
  • the support frame 30 is flexible in the out of plane direction as it then will facilitate removal of the gel member 36, by applying the release force mainly along the extension of the film to gradually release the gel from the cassette housing 20.
  • the support frame 30 may be of a more frame-like rigid structure, defining a substantial part of the gel compartment and the upper and lower walls 60 and 40 being removable from the rim of the rim 70 of the frame-like rigid structure.
  • Fig. 4 shows an enlarged section of the lower end of the cassette 10 according to one embodiment, wherein the fill port 120 comprises a membrane section 122, e.g. a septa, arranged to allow penetration of a fill nozzle, e.g. in the form of a syringe needle or the like, thereto for feeding gel solution into the cassette 10, but which effectively prevents the injected solution from leaking out when the fill nozzle has been removed and air from entering into the gel compartment.
  • the fill port 120 and the membrane section is comprised of an elastic material.
  • the fill port 120 may be co-molded in the same step as the sample well cover 50, with the main difference that the fill port 120 is designed to be permanently attached to the cassette housing 20 whereas the sample well cover is pealable.
  • the fill port 120 may be formed to be retained on the cassette housing 20 by mechanical means, such as by undercutting the opening into which the port 120 is molded, or alternatively by modifying the surface of the cassette housing 20 to increase adhesion.
  • the fill port 120 and the sample well cover 50 may be molded using the same injection port, and the structures being linked by a resin flow channel leaving a connection member 121 there between.
  • the connection member may e.g. be formed to break upon removal of the sample well cover 50 or may be cut before removal of the sample well cover 50.
  • Figs. 5 a-c illustrates the step of preparing the cassette 10 for performing an
  • FIG. 5b show the bottom side of cassette 10, and in fig. 5c it is schematically shown that removable sections 210a and 210b of the backing film 40 are removed from the cassette 10 by pulling peal tabs 211a and 211b respectively, whereby the gel member 36 becomes exposed through the buffer slits 150a and 150b of the support frame 30 respectively.
  • Fig 6a shows a perspective view of an electrophoresis tray 300 that is compatible with the electrophoresis cassette 10 for running electrophoresis experiments using the same.
  • the tray 300 is disclosed as a separate feature, but as will be disclosed below it may be an integral part of an electrophoresis system.
  • the tray 300 comprises a cassette support surface 310 for supporting at least the separation zone of an electrophoresis cassette 10 during electrophoresis.
  • the cassette support surface 310 is flanked by a pair of buffer holders 320a and 320b respectively, each one arranged to hold a buffer pad unit 322 (e.g. as is shown in fig.
  • the tray 300 comprises a heat transfer unit (not shown) connected to the cassette support surface 310 to control the temperature the electrophoresis cassette 10 during electrophoresis by heat transfer contact with a section of the back surface of the electrophoresis cassette 10.
  • the tray 300 comprises a flat top surface with two buffer holders 320a and 320b formed as two separate recesses therein, and an alignment structure 330 that is formed to be complementary to the alignment tag 180 of the support frame 30 to ensure proper orientation of the cassette 10 on the tray.
  • the alignment structure 330 is comprised of an elongated pin 340a, a circular pin 340b.
  • the alignment structure is made asymmetric, whereby proper orientation of the alignment tag 180 and the cassette 10 is ensured.
  • the buffer slits 150a and 150b of the support frame 30 are positioned at the respective buffer holders 320a and 320b to enable mating contact between the gel exposed through the buffer slits 150a and 150b (with the respective removable section 210a and 210b of the backing film 40 removed) and a buffer pad unit 322, schematically shown in figs 7a, 7b and 7c, placed in the respective buffer pad holders 320a and 320b, as is schematically shown in figs 8a, 8b and 8c.
  • the tray 300 is further provided with a lid 400 arranged to be closed over the cassette 10 during operation of the electrophoresis system. The details and functions of the lid 400 will be disclosed in more details below.
  • the buffer pad unit 322 schematically disclosed in figs. 7a, 7b and 7c comprises a cup 323 housing a buffer strip 324 in a buffer strip compartment 329 and an electrode arrangement 325.
  • Fig. 7b shows a top view of fig 7a with the buffer strip 324 placed in the buffer cup 323, whereas fig. 7c shows the bottom of the buffer cup 323.
  • the cup further comprises an external electrical connector 326 for connecting the electrode arrangement 325 to a power source of the electrophoresis apparatus.
  • the electrical connector 326 is provided as bias springs arranged both to provide electrical connection and to bias the buffer strip 324 against the back face of the gel member in cassette 10 when arranged in position as will be disclosed more in detail below.
  • the tray 300 is provided with complementary electrical connectors 327.
  • the cup 323 is formed to fit into the buffer pad holders 320a and 320b so that the top portion of the buffer strip 324 can be placed in contact with the gel in a cassette placed on the tray 300.
  • they are preferably keyed with respect to each other, and likewise the buffer strip 325 may be keyed with respect to the buffer cup 323.
  • the buffer cup 323 is of generally rectangular shape when viewed from above as in fig. 7b, but comprises a key flange 332 complementary to a recess in the buffer pad holders 320a and 320b.
  • the buffer strip compartment 329 of the buffer cup 323 comprises a section 334 that is complementary to a protrusion 333 on the buffer strip 324.
  • the gel strip 324 may be formed with a raised section 328 to facilitate contact to the gel in the cassette 10.
  • the disclosed buffer cup 323 is further provided with grooves 335 arranged to provide disposal of gas that may be generated in buffer strip compartment 329 of the buffer cup 323.
  • Figs. 9a to 9e show another embodiment of a buffer pad unit 322, similar to the one of figs 7a-c.
  • the keying between both the buffer cup 323and the buffer pad holders 320a and 320b as well as between the buffer cup 323and the buffer strip 325 is accomplished by a unitary structure 334 of the buffer cup 323 and complementary structures of the buffer holders 320a-b and the buffer strip 324 respectively.
  • the grooves 335 in the buffer strip compartment 329 of the buffer cup 323 are provided also in the bottom of compartment 329, whereby they will serve to accommodate waste liquid that may be produced at the buffer strip compartment 329 of the buffer cup 323, while the grooves 335 in the side walls of the compartment 329 enables disposal of gas (and will accommodate liquid in case the grooves in the bottom are flooded.).
  • the area of the electrode 325 should not be too small, in the disclosed embodiment, the electrode 325 is formed to cover a substantial section of the bottom area of the buffer strip compartment 329, e.g. nearly half the bottom area.
  • the electrode 329 is preferably arranged at a
  • the buffer strip 324 may be comprised of a buffer substance incorporated in a gel material e.g. the type disclosed in WO 87/04948. By placing the buffer strip 324 in a cup 323, changing the buffer media between electrophoresis runs is greatly facilitated, e.g. compared to placing gel strips directly in the buffer recess.
  • the buffer pad unit 322 is formed as a disposable unit potentially packed together with the cassette 10, but in another embodiment, the cup 323 including the electrode 325 are intended for reuse with disposable buffer strips 324 that are replaced after use.
  • the buffer pads are integrated with the electrophoresis cassette like in US6368481, which is incorporated herein by reference.
  • Fig 6b shows an enlarged view of the lid 400 of the tray 300 in fig 6a.
  • the lid 400 is pivotally connected to the tray 300 by hinges 402 and is arranged to be closed over a cassette 10 (or transfer membrane) placed on the tray 300 by a pivotal movement to a horizontal position.
  • a locking mechanism 404 arranged to interact with a complementary locking structure on the tray 300.
  • the present electrophoresis system is arranged to enable imaging of the electrophoresis gel in the same system. Therefore, the lid is provided with an imaging window 406 covered by a transparent window pane 407 of glass or the like.
  • the window pane should be selected to be of low fluorescent type.
  • a suitable material is a low- fluorescent Borofloat glass.
  • the window pane may be provided with an anti-reflex coating .
  • the lid is further provide with a sample well window 408.
  • the lid 400 is provided to secure the cassette 10 to the tray 300, in order to provide improved thermal contact between the cassette 10 and the cassette support surface 310 to control the temperature the electrophoresis cassette 10 during electrophoresis by heat transfer contact. During electrophoresis, uniform cooling over the separation-zone cassette 10 is desirable in order to provide high quality results. Improved thermal contact is partially achieved in that the lid 400 mechanically pushes the cassette 10 against the cassette support surface.
  • the lid 400 is further provided with a sealing member 410 that extends around the window 406 to provide a closed compartment covering the separation-zone of the cassette 10. Further thermal contact may then be achieved by pressurizing the closed compartment as will be disclosed in more detail below.
  • a gas supply connection 418 in communication with a gas port 420 in the inner rim of the compartment.
  • a suitable source of pressurized gas e.g. pressurized air or any other suitable gas
  • the compartment may be pressurized to a suitable pressure.
  • the pressure is set to be greater than 0.05 bar, or greater than 0.1, 0.2, 0.3 bar and.
  • the sealing member 410 may be a lip type seal or the like, and it may be provided with a reinforced section in between the sample well window 408 and the imaging window 406.
  • the purpose of the reinforced section is to apply a higher pressure on the cassette 10 in this region, partly to prevent potential spillage from the sample wells to enter the closed compartment over the separation-zone and partly to provide an improved support for the cassette 10 when the well cover 50 is removed from the wells.
  • Figs 8a to 8f schematically show the steps of loading a cassette 10 on the tray 300 for performing an electrophoresis separation experiment. The individual order of some steps may vary.
  • Buffer strips 324 are placed in buffer cups 323 to form two buffer pad units 322 to be placed in buffer pad holders 320a and 320b in the tray 300.
  • Fig 8b shows the tray 300 with the buffer pad units 322 in position.
  • Figs. 8c and 8d shows positioning of a cassette 10 in position on the tray 300, wherein the position of the cassette 10 is uniquely determined by the alignment structures 340a and 340b.
  • the removable sections 210a and 210b of the backing film 40 should be removed by pulling peal tabs 211a and 211b respectively, whereby the gel member 36 becomes exposed through the buffer slits 150a and 150b of the support frame 30 respectively.
  • the lid 400 When the cassette 10 is placed in position on the tray 300, the lid 400 may be closed as shown in figs. 8e and 8f. • As mentioned, the sample well window 408 of the lid 400 is arranged to provide access to the well cover 50, and to provide support for the cassette 10 when the well cover 50 is removed from the wells as indicated in fig 8f. When the well cover 50 has been removed, the wells 110 are accessible in the sample well window for supplying sample, e.g by a pipette or the like.
  • Figs. 10a to lOf show a schematic cross-sectional view of a tray 300 arranged on a tray loading mechanism 502 of an electrophoresis system 500, wherein some details have been simplified or moved into the cross-sectional plane for illustrative purposes.
  • buffer strips 324 and buffer cups 323 are not yet placed in position in the buffer pad holders 320a and 320b.
  • the buffer cups 323 are shown with the bias spring 326 attached to the bottom of the cup and the electrical connection to the electrode 325 has been left out.
  • the buffer cups 323 are provided with an inclined bottom surface, which serves to position the buffer strip closer to the wall in the direction of inclination, further the buffer cups 323 as well as the buffer strips 325 are arranged with the corresponding wall closest to the cassette support surface 310 to create a symmetrical arrangement.
  • the electrophoresis cassette 10 is shown elevated above the cassette support surface 310 of the tray 300 in position to be docked onto the tray 300.
  • the mating of the buffer pads and the buffer connection sections may be biased to some degree. This may be especially important for some gel/pad compositions wherein one may get mass transfer of e.g.
  • the buffer pad unit 322 By biasing the buffer pad unit 322 against the gel member 36 such situations may be accomplished for.
  • suitable material properties for the gel component of the buffer pads 322 they may be comprised of a suitable resilient material capable of at least partially providing the biased mating.
  • the biased mating may be achieved by providing buffer strips 324 of specific shape that allow a certain degree of compression due to its shape.
  • the bias spring 326 is arranged to provide for the biased mating in combination with the material characteristics of the buffer strip 324 and the shape of the same. In figs.
  • the tray 300 and the lid 400 and their respective components are schematically disclosed in cross-section.
  • the gas supply connection 418 is visualized as a flexible gas conduit connected to the lid 400, and the gas port 420 is disclosed as a bore connecting the gas conduit to the compartment covering the separation-zone of the cassette 10, as is specifically show in figs. 10c- lOf.
  • the buffer pads 322 are slightly biased towards the back of the gel cassette 10 and the gel member 36 by bias springs 326, and when the lid 400 is closed, as is disclosed in figs.
  • the cassette 10 is pressed further down towards the support surface 310 of the tray 300 whereby the buffer pads are biased against the gel cassette 10 and the gel member 36.
  • the well cover 50 is still accessible through window 408 and the lid 400 serves to keep the cassette 10 in place when the well cover 50 is removed.
  • sample may be added in the sample wells 110 using a pipette 506 or the like, whereby the
  • electrophoresis system 500 is ready for running electrophoresis and scanning operations.
  • the electrophoresis system 500 may further comprise a door 504 to be closed when the electrophoresis tray has been transferred into the housing of the system 500 by the tray loading mechanism 502.
  • Figs lOe and lOf schematically show the function of the tray loading mechanism 502 and the door 504 of the system 500.
  • Figs. 1 la to 11c schematically discloses one embodiment of an electrophoresis system 500 arranged to perform both electrophoresis and imaging of the electrophoresis sample.
  • the electrophoresis system 500 comprises a fluorescence imaging unit for imaging the result of the separation directly in the apparatus.
  • the electrophoresis cassette 10 need not to be moved to a separate imaging unit following the separation.
  • the disclosed cassette 10 may be designed for imaging, by proper materials selection and design to avoid undesirable optical effects such as fluorescence emitted by parts of the cassette, image distortion etc.
  • electrophoresis tray 300 with buffer pads 322 recessed in the tray 300 is that the resulting electrophoresis set up is of low profile, whereby the imaging unit may operate in the close vicinity of the gel to increase sensitivity and resolution, and to avoid negative optical effects.
  • the electrophoresis tray 300 is shown in essentially horizontal position with the gel cassette 10 arranged on top thereof.
  • the electrophoresis tray 300 as well as the gel cassette 10 may be arranged for use in other orientations such as vertical or even upside down by making suitable adaptions.
  • the electrophoresis system 500 may comprise any suitable components for performing combined electrophoresis and imaging of the separated sample in the gel.
  • the system comprises a control unit 510, a power supply 520 for driving the electrophoresis separation, a heat transfer unit 525, a gas pressurizing source 530, and an imaging unit 540.
  • the control unit 510 is arranged to control the general operation of the system 500 and the components thereof.
  • the power supply 520 may be any suitable power supply capable of providing required power for driving the electrophoresis separation in the cassette 10.
  • the system may be connected to an external computer (not shown) for further processing and analysis of acquired images, as well as setting control parameters or the like for the electrophoresis and image acquisition processes.
  • the system may be connected to the external computer through any suitable communication interface, and user interaction may be performed through any suitable user interface on the computer.
  • the system 500 comprises an integrated user communication interface.
  • the gas pressurizing source 530 may e.g be a pump, or a valve connected to an external pressure source, capable of providing the desired over pressure in the compartment defined by window 406 covering the separation-zone of the cassette 10.
  • the imaging unit 540 is schematically shown as a scanning module with an imaging controller 542 and a scan head 544.
  • the scanning module may be either a point type scanner arranged to be translated both in the X and Y directions with respect to the cassette 10, or it may be a line type scanner wherein it only has to be translated in one direction.
  • the tray loading mechanism 502 is arranged to move the sample tray and this the cassette 10 in the Y direction with respect to the scan head 544, whereby the scan head 544 need only to be translated in the X direction in order to scan the full width of the separation zone.
  • Fig 11c shows an enlarged view of the imaging unit 540 wherein the scan head 544 comprises a housing 545, an excitation light source 546, a beam splitter 548, a beam focusing lens 550, an optical filter 552, a detector lens 554, a detector 556 and a X- translation member 558.
  • the scan head 544 is designed with a focal plane in the middle of the gel member 36.
  • the separation-zone of the cassette 10 is forced against the support surface by providing an over-pressure in the compartment 406 formed by the lid with the window pane 407.
  • Fig. 12 schematically discloses a cross-sectional view of one embodiment of the tray loading mechanism 502 transverse to the view in figs. 1 la-1 lc.
  • the tray loading mechanism 502 there are two trays 300 arranged on the tray loading mechanism.
  • an imaging system 540 with a double beam scan head 544 enabling simultaneous scanning at two different wavelengths.
  • a schematic X-translation mechanism 558 comprising a slide bar 560, and a drive belt 562.
  • the scan head(s) may be tilted with respect to the direction normal to the tray 300. By selecting a suitable tilt angle, reflections from the window pane 407 etc. may be suppressed.
  • Fig 13 schematically discloses an alternative embodiment of the imaging system 565, based on a 2 dimensional camera detector 566 capable of capturing the whole separation-zone in one single image.
  • the imaging system 565 further comprising an illumination source 567, a filter 568 and a lens unit 569 (schematically shown as one single lens for illustrative purposes, but it may be comprised of any number of optical elements required to project an image of the separation-zone.
  • the combined electrophoresis and imaging system provides several opportunities to follow the electrophoresis process or in other ways use the imaging to verify operation or the like of the electrophoresis separation.
  • the routines for following or verifying the operation of the electrophoresis process may be implemented by software code that may be executed by a processor unit in the control unit 510, the imaging unit 540 and/or an external computer.
  • the combined electrophoresis and imaging system may be arranged to detect the separation front of the electrophoretic separation in order to automatically verify when the separation process is finished and the sample is separated.
  • the imaging unit is arranged to detect the presence or not of dyed sample at a predetermined position with respect to the separation-zone and the system may be arranged to end the separation process in response to the detection of the separation front.
  • the imaging unit is arranged to follow the separation front in order to provide a real time status of the separation process.
  • the combined electrophoresis and imaging system may be arranged to register the presence of dye or not in the sample wells by scanning the sample wells prior to the electrophoresis process is initiated. If the sample fluid is transparent, it may be difficult for a user to see if the sample was actually applied in the sample well. In addition, if no dye or a dye not matching the laser wavelengths of the instrument is used, this can be detected before the run. This will save the user a lot of time since he/she will not need to make a complete run with a inevitable failure.
  • the scanning device should be positioned over the sample wells.
  • the laser should be lit and the result analysed. If the sample fluoresces, the sample should be assumed to be correct. If there is no fluorescence in a well, the user should be asked to check if a sample is loaded and if a correct dye is used.
  • the system can determine the highest possible intensity that may be achieved by the sample. Using this value to the adjust the amplification (PMT) for maximum signal to this level, the system get the full dynamic capacity for the scanning of the final separation pattern.
  • PMT adjust the amplification
  • This signal can also be used for normalisation between the lanes and the final bands. Today you may have to make a pre-scan the gel on the area with the most intense bands to determine the right amplification to get a good image out of the gel. According to one embodiment this signal may be used for normalisation of sample application between bands.
  • electrophoresis gel using an electrophoresis system comprising the steps:
  • the signal from the most intense well is then used to determine an amplification setting to reach the highest possible signal level at this intensity.
  • this amplification setting is used to scan the complete gel. This setting will in practice give an image where all bands are within range and that the dynamic capacity is used in an optimal way. (In theory a band could have higher signal that the sample well since we use stacking to concentrate the sample. )
  • sample well signal for normalisation of sample application variation and as standards for band quantitation during evaluation of the final image.
  • the sealing member 410 may be a lip type seal or the like.
  • Figs 14a to 14d show one embodiment of a sealing member 410 of lip type more in detail.
  • the lip of the present seal 410 is inclined towards the pressurizing compartment whereby the over pressure in the compartment will force the lip against the cassette top surface and thus improve the sealing effect.
  • the lip seal 410 may be provided with a reinforced section in between the sample well window 408 and the imaging window 406.
  • the lip seal is molded using the lid 400 as a part of the mold, whereby the seal is permanently attached to the lid by the molding process.
  • One additional advantage compared to a normal lip seal is that it has a lower press force and it is also more predictable regarding the force applied onto the cassette 10. Further, the seal has a wider flexible dynamic range, whereby it is not so tolerance sensible, and it is possible to manufacture this type of mold tool only with 5- axis machining technology.
  • Fig 15 shows a membrane unit 600 for immunob lotting comprised of a membrane 610 that is attached to a rigid blot frame 620.
  • the rigid blot frame 620 is designed to stay attached to the membrane 610 throughout the process steps.
  • the rigid blot frame 620 is formed of a suitably rigid material to preserve the shape of the membrane 610 and to facilitate handling of the membrane 610 by providing accessible gripping portions outside of the transfer-zone.
  • the rigid blot frame 620 further comprises an alignment tag 640 with a predefined alignment structure in the form of two alignment holes 650a and 650b, arranged to ensure that the membrane unit 600 is properly aligned with respect to a complementary alignment structure e.g. comprising 2 pins, in an transfer unit, a scanner or the like.
  • the alignment structure 650a and b is compatible with or essentially identical with the alignment structure 190a and b of the gel support frame.
  • suitable alignment means the gel member 36 and the membrane unit 600 may be aligned during the transfer process to create a known geometrical relationship between the bands of the electrophoresis gel and the transferred bands.
  • the membrane unit 600 is provided without a rigid blot frame 620.
  • the electrophoresis system of the present invention is arranged to enable scanning of membrane units 600 following the transfer process. However, as the membrane 610 of the membrane unit 600 is substantially thinner than the gel member 36 of the cassette 10, the focal plane of the imaging unit 540 will not coincide with the membrane 610 if the membrane unit would be positioned directly on the support surface 310 of the tray 300.
  • the imaging system 540 is arranged to automatically adjust the focal plane accordingly.
  • a spacer unit providing a support surface for the membrane 610 at the correct focal plane.
  • the membrane may not be easily placed on a support surface in a way that it is preserved flat enough to enable high quality imaging.
  • Fig 16a-16d disclose one embodiment of a membrane unit holder 700, arranged to position the membrane 610 in the desired position with respect to the focal plane of the imaging system as well as holding the membrane 610 flat against the support surface.
  • the membrane unit holder 700 comprises a spacer member 710 of predetermined thickness selected to position a membrane 610 positioned thereon at the desired focal plane, and a cover frame 720 of a predetermined thickness selected so that the total thickness of the membrane unit holder 700 with a membrane unit 600 placed therein essentially corresponds to the height of a cassette 10.
  • the cover frame 720 comprises a window 730 corresponding to the separation-zone of the cassette 10 and thus the membrane section 610 of the membrane unit.
  • both the spacer member 710 and the cover frame 720 are provided with a predefined alignment structure in the form of two alignment holes 740a, 740b and 741a , 741b respectively to provide proper aligned with respect to a complementary alignment structure e.g.
  • the alignment structure is optional for at least the spacer member 710.
  • the spacer member 710 and the cover frame 720 are optionally connected by a hinge member 750.
  • Figs. 16 b-16d show the positioning of the membrane unit 600 is position with respect to the membrane unit holder 700.
  • membrane unit 600 is placed on top of the spacer member 710, both in alignment with the alignment pins of the tray 300.
  • the cover frame 720 is positioned on top of the membrane unit 600 whereby the membrane 610 is exposed through the window 730 of the cover frame 720.
  • Figs 17a- 17c show a schematic cross sectional view of the membrane unit holder 700 with the membrane unit 600 placed therein.
  • Fig 17c shows a close up view of the membrane unit holder 700 arrangement.
  • the spacer member 710 and the cover frame 720 By selecting suitable material properties and surface characteristics for the spacer member 710 and the cover frame 720 it is possible to create a net flow of air (gas) through the membrane, along the upper surface of the spacer member 710 into the ambient surroundings, whereby the membrane is effectively flattened out against the top surface of the spacer member 710.
  • This net flow of air may in one embodiment be achieved by providing the top surface of the spacer member 710 with a suitable degree of surface roughness enough to provide a flow of air between the top surface and rigid blot frame 620 of the membrane unit, while providing a higher degree of sealing between the bottom surface of the cover frame 720 and the blot frame 620.
  • the net flow of air is indicated by arrow 760 in fig 17c.
  • spacer member 710 may be comprised of a porous material or be provided with an array of through holes and vent channels on the bottom surface thereof.
  • an electrophoresis cassette comprising a gel member in a housing with a front and a back face
  • an electrophoresis tray arranged to support the electrophoresis cassette for running electrophoresis experiments, wherein the tray comprises a cassette support surface for supporting at least the separation zone of the electrophoresis cassette during electrophoresis, and wherein the cassette support surface is flanked by a pair of buffer pad holders each one arranged to hold a buffer pad in a mating position with respect to buffer connection sections at the back face of the electrophoresis cassette,
  • the method may further comprise steps of providing an imaging unit for imaging the gel member when it is in position on the tray.
  • the step of imaging may be performed before, during and/or after the electrical field is applied.
  • an electrophoresis system comprising: at least one type of electrophoresis gel card,
  • a blot transfer unit for transfer of separated sample from the electrophoresis gel card to the blot membrane card
  • the electrophoresis gel card and the blot membrane card each comprises a rigid support provided with an alignment structure defining a positional reference for mutual alignment during transfer, and for alignment with respect to a complementary alignment structure in the imaging apparatus to provide mechanically aligned images of separated sample in the electrophoresis gel card and the blot membrane card.
  • the electrophoresis apparatus and the imaging apparatus may be integrated into one apparatus being arranged to run both electrophoresis experiments and to image the electrophoretic gel card wherein the step of imaging may be performed before, during and/or after the electrical field is applied to drive electrophoresis experiments.
  • the electrophoresis system may comprise a transfer holder with a complementary alignment structure for holding the
  • electrophoresis gel card and the blot membrane card in mutual aligned position in the blot transfer unit.
  • a transfer holder is shown in detail in the above referenced patent applications and in the co-pending application by the same applicant: 3467/DEL/2013 in India which is incorporated by reference.
  • the electrophoresis gel card may comprise a housing with removable members to expose both the first and second face of the gel member to allow blot transfer of separated sample while the gel member is attached to the rigid support.
  • the electrophoresis gel card may further comprise at least one removable member that has to be removed in order run at least one step in the electrophoresis workflow, and wherein the removable member is formed to at least partially block the alignment structure to prevent running said step without first removing the removable member.
  • the alignment structures of the electrophoresis gel card and the blot membrane card are formed to define a unique orientation of respective card.
  • the alignment structures of the electrophoresis gel card and the blot membrane card comprises at least one alignment hole and wherein a complementary alignment structure comprises a complementary alignment pin.
  • the electrophoresis gel card and the blot membrane card may each comprise an identification code, and the identification codes may be arranged to be simultaneously read when mutually aligned for transfer to establish a unique link between said cards, and the system may be arranged to store said link.
  • the identification code may e.g. be a machine readable code as a bar-code, matrix- code or the like, and provide the user and/or instruments with relevant information.
  • the imaging apparatus may be arranged to read the identification code of a card arranged for imaging, and in one embodiment, the imaging apparatus may be arranged to select an imaging protocol based on the registered identification code of a card arranged for imaging.
  • identification code that is arranged to be transferred to a blot membrane card at the blot transfer.
  • the identification code may be electrochemically transferred from the electrophoresis gel card to the blot membrane card.
  • a separation and identification method comprising the steps: separating a sample, by electrophoresis, in an electrophoresis gel card comprising a rigid support provided with an alignment structure defining a positional reference, acquiring an image of the sample separated in the electrophoresis gel card using an imager with a complementary alignment structure, and wherein the alignment structure of the electrophoresis gel card is arranged in alignment with the complementary alignment structure, transferring sample constituents from the gel card to a blot membrane card comprising a rigid support provided with an alignment structure defining a positional reference, wherein the electrophoresis gel card and the blot membrane card are arranged in mutual alignment by means of the alignment structures, acquiring an image of transferred sample constituents on the blot transfer card wherein the alignment structure of the blot transfer card is arranged in alignment with the complementary alignment structure of the imager, and analyzing the images comprising the step of correlating the images based on the mutual alignment.
  • the concept of providing a support frame for the gel member and/or blot membrane thus provides a whole range of benefits for a protein analysis system based on electrophoresis and immunoblotting.
  • Figs. 18a and 18 b disclose one embodiment of a support surface 310 with good heat transfer characteristics and low fluoresensce, but which is electrically insulated in order to avoid short circuits or spikes due to the high voltages involved.
  • efficient cooling of the gel member 36 with an effect of e.g. 20 Watt, while preventing electric flashover at high voltages involved in electrophoresic separation, e.g. of more than 600 Volts.
  • it induces very Low fluorescence due to the materials properties, while it provides resistance to withstand mechanical wear as well as being chemically resistant to withstand salty substances (Buffer Gel Liquids) and the like.
  • the base 805 of the support surface 310 is comprised of a material with high thermal conductivity properties, such as aluminum, copper or the like.
  • a polymer foil 800 is arranged on the cooling plate 310 and/or combined with different surface treatments underneath in closer connection to the cooling plate 310.
  • the cooling plate 310 also needs to be protected on most of the surfaces, not only the top surface because the buffer gel units 322 (not shown) are placed underneath the electrophoresis cassette (not shown). But in other embodiments, where buffer pads units 322 are not provided as an integrated part of the tray 300, e.g. provided on top of the electrophoresis cassette, it may be sufficient to only cover the top surface.
  • FIG. 18b one embodiment of the electrically insulated cooling plate (support surface) 310 is shown in an exploded view.
  • the cooling plate 310 being comprised of a thermally (and electrically) conductive main body 805, an insulating film 800, a first insulating end piece 810 with alignment pins, and a second insulating end piece 820.
  • the end pieces 810 and 820 are made of any suitable insulating material such as plastics, ceramics or the like.
  • Fig. 18a shows the cooling plate in assembled state with the lid elevated there above for illustrative purposes.
  • the disclosed embodiment comprises an insulating foil (film) 800 on top of the main body 805 and with non-conducting end pieces 810 and 820 e.g. made of parylene and plastic.
  • This design manages to protect all important surfaces from getting exposed to any buffer liquids in combination with high voltage from the electrophoresis process.
  • the temperature of the electrophoresis gel member 36 in the cassette 10 is preserved by the cooling plate 310, and constant temperature is ensured by a peltier based cooling unit (not shown) (cooled by air ventilation) combined with an extra separate aluminum plate with very thin protective layers.
  • the cooling plate 310 is provided with a three layer protective arrangement comprised of an insulating aluminum anodization layer, an insulating parylene coating on the anodized aluminium surface and as mentioned above an insulating film of plastic material facing the cassette 10, e.g. a PET-G foil with acryl based adhesive.
  • the peltier cooling unit needs no secondary liquids, only air. It can also be provided as a separate unit. This is beneficial for the moving function of the electrophoreses unit during scanning e.g. in order to avoid attachment of liquid conduits to the cooling unit in case of a liquid based cooling unit.
  • the peltier cooling concept is selected based on having a few number of components used, simplicity and cost effectiveness. Other designs may be based on liquid cooling for the cooling plate and heat pipes. Testing has showed that peltier elements secured enough cooling effect performance.
  • the warmest part or region of the gel card is located in the sample front.
  • the cooling concept according to the present embodiment has been verified to fulfill the
  • the cooling temperature is also very even on the cooling surface in accordance with the present embodiment.
  • the divided cooling plate (upper plate and lower plate) (not shown) also improves the evenness. It has further been verified that the present cooling unit 310 is capable of cooling the cassette 10 within acceptable time. The performance of the cooling unit 310 is enough for the time to cool the cassette lOfast enough for the electrophoreses. Further, the cooling plate 805 is surrounded by plastic parts and air gaps to reduce the loss of cooling effect.
  • Fig 19 discloses one embodiment of a buffer cup 900 wherein the electrode 910 is formed as an essentially integral part of the cup, by sputtering or another suitable surface modification of at least a section of the inner surface of the cup arranged to be in contact with a buffer strip 324 when placed therein.
  • a buffer cup 900 may be provided as a disposable buffer pad unit 322.
  • the buffer cup 900 is partly used as a mold for molding a buffer pad directly therein, whereby the cost of production may be reduced.

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Abstract

La présente invention concerne un système d'électrophorèse comprenant un plateau d'électrophorèse servant à recevoir une cassette de gel d'électrophorèse et une unité d'imagerie servant à prendre une image du gel d'électrophorèse, le plateau d'électrophorèse comprenant une surface de support de cassette servant à supporter au moins une zone de séparation de la cassette d'électrophorèse pendant une électrophorèse, et un bouchon fermant la cassette d'électrophorèse 10 pendant le fonctionnement du système d'électrophorèse, le bouchon comprenant une fenêtre d'imagerie permettant de prendre une image de la zone de séparation.
PCT/EP2014/076019 2013-11-29 2014-11-28 Système d'électrophorèse Ceased WO2015079048A1 (fr)

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Cited By (13)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2016141495A3 (fr) * 2015-03-06 2016-11-03 4D Lifetec Ag Système d'électrophorèse sur gel conçu pour une électrophorèse sur gel à cellule unique
WO2018138383A1 (fr) 2017-01-30 2018-08-02 4D Lifetec Ag Plaque support pour appareils de laboratoire
WO2019126693A1 (fr) * 2017-12-22 2019-06-27 Quest Diagnostics Investments Llc Analyse automatisée de gels et d'empreintes de transfert analytiques
WO2020079211A1 (fr) * 2018-10-18 2020-04-23 Danmarks Tekniske Universitet Ensemble d'électrophorèse
WO2020192755A1 (fr) * 2019-03-28 2020-10-01 上海易孛特光电技术有限公司 Procédé et appareil pour l'imagerie d'un objet auto-lumineux sur un film d'échantillon biologique
CN112684161A (zh) * 2020-12-11 2021-04-20 东南大学 任意泳道的western blot孵育与检测的装置
US20220026390A1 (en) * 2020-07-27 2022-01-27 Bio-Rad Laboratories, Inc. Electrophoresis apparatus with minimal autofluorescence to enable gel processing in situ
CN114166916A (zh) * 2021-12-17 2022-03-11 武汉泰沃科技有限责任公司 一种高分辨蛋白电泳预制胶容器
WO2022015661A3 (fr) * 2020-07-13 2022-04-14 Life Technologies Corporation Dispositifs, systèmes et procédés d'électrophorèse et d'électrotransfert
EP4006536A1 (fr) * 2020-11-27 2022-06-01 Simo Abdessamad Baallal Jacobsen Ensemble d'electrophorese et methodes
WO2022115076A1 (fr) * 2020-11-24 2022-06-02 T.C. Trakya Universitesi Cytométrie de flux à électrophorèse sur gel à une seule cellule
USD975873S1 (en) 2020-07-13 2023-01-17 Life Technologies Corporation Electrophoresis and electrotransfer device
WO2024122250A1 (fr) * 2022-12-07 2024-06-13 日本航空電子工業株式会社 Dispositif d'électrophorèse sur gel

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1996034276A1 (fr) * 1995-04-26 1996-10-31 Brun, Heidi, M. Appareil et procede d'electrophorese
US20040050699A1 (en) * 2002-09-11 2004-03-18 Goncalves Antonio M. Automated system for high-throughput electrophoretic separations
WO2006119941A1 (fr) * 2005-05-11 2006-11-16 Ge Healthcare Bio-Sciences Ab Procede et dispositifs servant a prendre l'image d'un specimen

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1996034276A1 (fr) * 1995-04-26 1996-10-31 Brun, Heidi, M. Appareil et procede d'electrophorese
US20040050699A1 (en) * 2002-09-11 2004-03-18 Goncalves Antonio M. Automated system for high-throughput electrophoretic separations
WO2006119941A1 (fr) * 2005-05-11 2006-11-16 Ge Healthcare Bio-Sciences Ab Procede et dispositifs servant a prendre l'image d'un specimen

Cited By (22)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2016141495A3 (fr) * 2015-03-06 2016-11-03 4D Lifetec Ag Système d'électrophorèse sur gel conçu pour une électrophorèse sur gel à cellule unique
US10591439B2 (en) 2015-03-06 2020-03-17 4D Lifetec Ag Gel electrophoresis system for single cell gel electrophoresis
WO2018138383A1 (fr) 2017-01-30 2018-08-02 4D Lifetec Ag Plaque support pour appareils de laboratoire
US11351554B2 (en) 2017-01-30 2022-06-07 4D Lifetec Ag Carrier plate for laboratory devices
WO2019126693A1 (fr) * 2017-12-22 2019-06-27 Quest Diagnostics Investments Llc Analyse automatisée de gels et d'empreintes de transfert analytiques
US12174142B2 (en) 2017-12-22 2024-12-24 Quest Diagnostics Investments Llc Automated analysis of analytical gels and blots
US11686703B2 (en) 2017-12-22 2023-06-27 Quest Diagnostics Investments Llc Automated analysis of analytical gels and blots
WO2020079211A1 (fr) * 2018-10-18 2020-04-23 Danmarks Tekniske Universitet Ensemble d'électrophorèse
WO2020192755A1 (fr) * 2019-03-28 2020-10-01 上海易孛特光电技术有限公司 Procédé et appareil pour l'imagerie d'un objet auto-lumineux sur un film d'échantillon biologique
US12032149B2 (en) 2019-03-28 2024-07-09 Shanghai E-Blot Photoelectric Technology Co., Ltd. Imaging method and apparatus for self-luminous object on biological sample film
USD1065585S1 (en) 2020-07-13 2025-03-04 Life Technologies Corporation Electrophoresis and electrotransfer device
WO2022015661A3 (fr) * 2020-07-13 2022-04-14 Life Technologies Corporation Dispositifs, systèmes et procédés d'électrophorèse et d'électrotransfert
US12292408B2 (en) 2020-07-13 2025-05-06 Life Technologies Corporation Electrophoresis and electrotransfer devices, systems, and methods
USD975873S1 (en) 2020-07-13 2023-01-17 Life Technologies Corporation Electrophoresis and electrotransfer device
US20220026390A1 (en) * 2020-07-27 2022-01-27 Bio-Rad Laboratories, Inc. Electrophoresis apparatus with minimal autofluorescence to enable gel processing in situ
WO2022115076A1 (fr) * 2020-11-24 2022-06-02 T.C. Trakya Universitesi Cytométrie de flux à électrophorèse sur gel à une seule cellule
EP4006536A1 (fr) * 2020-11-27 2022-06-01 Simo Abdessamad Baallal Jacobsen Ensemble d'electrophorese et methodes
CN112684161B (zh) * 2020-12-11 2024-03-12 东南大学 任意泳道的western blot孵育与检测的装置
CN112684161A (zh) * 2020-12-11 2021-04-20 东南大学 任意泳道的western blot孵育与检测的装置
CN114166916A (zh) * 2021-12-17 2022-03-11 武汉泰沃科技有限责任公司 一种高分辨蛋白电泳预制胶容器
WO2024122250A1 (fr) * 2022-12-07 2024-06-13 日本航空電子工業株式会社 Dispositif d'électrophorèse sur gel
JP7529221B2 (ja) 2022-12-07 2024-08-06 日本航空電子工業株式会社 ゲル電気泳動装置

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