WO2005036160A1 - Cassette d'electrophorese - Google Patents

Cassette d'electrophorese Download PDF

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Publication number
WO2005036160A1
WO2005036160A1 PCT/CA2004/001827 CA2004001827W WO2005036160A1 WO 2005036160 A1 WO2005036160 A1 WO 2005036160A1 CA 2004001827 W CA2004001827 W CA 2004001827W WO 2005036160 A1 WO2005036160 A1 WO 2005036160A1
Authority
WO
WIPO (PCT)
Prior art keywords
cassette
gel
engagement
opening
agarose
Prior art date
Application number
PCT/CA2004/001827
Other languages
English (en)
Inventor
Pierre Sevigny
Original Assignee
Mirador Dna Design Inc.
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 Mirador Dna Design Inc. filed Critical Mirador Dna Design Inc.
Publication of WO2005036160A1 publication Critical patent/WO2005036160A1/fr

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Classifications

    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D57/00—Separation, other than separation of solids, not fully covered by a single other group or subclass, e.g. B03C
    • B01D57/02—Separation, other than separation of solids, not fully covered by a single other group or subclass, e.g. B03C by electrophoresis
    • 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

Definitions

  • This invention relates to a cassette for electrophoresis that can be precast and disposed after use. (b) Description of Prior Art
  • Agarose gel electrophoresis is a technique used for charged macromolecules separation using their electrical properties, their size and their structure.
  • the gel form a porous network serving as a stable environment for the buffer and the samples.
  • the buffer is the most active component in the sample migration-separation. This is the reason why the ionic strength and pH must be carefully chosen to insure an appropriate electrophoretic mobility.
  • the gel pore size is another factor influencing the electrophoresis.
  • the pore size is slowing down the sample mobility based on the physical structure.
  • the pore size more exactly the agar or agarose gel pore radius, is directly related to the agar or agarose concentration.
  • the agarose gels are mainly used horizontally. Such position allows a better support for the gel, most importantly for low concentration gels. It also creates less distortion in the electrophoresis pattern, it is much more regular.
  • agarose gel separation is mainly used to separate DNA molecules, using Tris-Acetate-EDTA or Tris — Borate-EDTA buffers.
  • Agarose powder is expensive.
  • the normal way to prepare a gel is to weigh the powder, add water and buffer and bring it to boil on a flame, or mostly used, in a microwave oven. During boiling, it is important to stop the over and mix the agarose to allow an equilibrated dissolution and prevent non-dissolved agarose particles to stay and contaminate the gel, which would affect the migration path and resolving quality.
  • This process is time consuming and not efficient, especially for the ones having to perform a electrophoresis assays on a regular basis.
  • a cassette for containing an electrophoresis gel comprising: a first portion; a second portion for hermetic engagement with the first portion; the first and second portion defining, when engaged, an internal volume for receiving gel; an opening for allowing filing of the internal volume with gel; one of the first and second portion having on an internal surface thereof a series of teeth forming a comb allowing the formation of channels upon filing of the cassette with the gel.
  • a cassette for containing an electrophoresis gel comprising: a first portion and second portion for hermetic engagement with the first portion, the first and second portion defining, when engaged an internal volume, an opening for allowing filing of the internal volume with the gel; and a standing surface opposed to the opening, the cassette standing on the standing surface upon filing of the internal volume with the gel.
  • a cassette for containing an electrophoresis gel comprising: a first portion; a second portion for hermetic engagement with the first portion, the first and second portion defining, when engaged, an internal volume; releasable attachment means for maintaining hermetic engagement of the first and second portion and for allowing opening of the cassette.
  • a method of filing the cassette of the present invention comprising the steps of positioning the cassette vertically standing on a face opposed to its opening; and filing the internal volume with said gel through the cassette's opening.
  • the first portion further comprises engagement means and the , second portion further receiving means complementary to the engagement means for hermetic engagement of the first and second portion together.
  • engagement means is preferably an insert portion extending from the periphery of the first portion and the receiving means is preferably a groove formed in the periphery of the second portion.
  • the hermetic engagement can also be provided by a gasket between the first and second portion to provide hermetic engagement of the first and second portion.
  • the cassette further comprises stacking means on the first and second portion for stacking a plurality of cassettes together.
  • the cassette further comprises standing means on at least one of the first and second portion to provide vertical stability to the cassette.
  • the first and second portion are secure together by means of attachments.
  • these attachment are made breakable upon opening.
  • These attachment can however also be resilient are allowing multiple opening and closing of the cassette for the cassettes used in applications were contamination is not a concern.
  • electrophoretic is intended to mean a technique in which charged complexes of proteins and/or nucleic acids with an appropriate gel are subjected to an electric field causing thereby migration of the charged molecule in the gel and where the relationship between the molecular weight of the charged molecule and its mobility is linear and can be used to estimate the charged molecule's molecular weight.
  • gel is intended to mean any gel suitable for performing electrophoresis, such as, but not limited to, agar, agarose, acrylamide and starch.
  • Fig. 1 is a perspective view of an embodiment of the cassette of the present invention when assembled and closed;
  • Fig. 2A is a perspective view of the first portion of the cassette of Fig. 1 ;
  • Fig. 2B is a top elevation view of the first portion of the cassette of Fig. 1 ;
  • Fig. 2C is a profile cross-sectional view of the first portion of the cassette of Fig. 1 taken along lines A-A of Fig. 2B;
  • Fig. 2D is a front cross-sectional view of the first portion of the cassette of Fig. 1 taken along lines B-B of Fig. 2B;
  • Fig. 2E illustrates the details of the closing device of the first portion of the cassette of Fig. 1 ;
  • FIG. 3A is a perspective view of the second portion of the cassette of Fig. 1 ;
  • Fig. 3B is a top elevation view of the second portion of the cassette of Fig. 1 ;
  • Fig. 3C is a profile cross-sectional view of the second portion of the cassette of Fig. 1 taken along lines A-A of Fig. 3B; [0028] Fig. 3D illustrates the stand of the second portion of the cassette of Fig. 1 ;
  • Fig. 3E is a front cross-sectional view of the second portion of the cassette of Fig. 1 taken along lines B-B of Fig. 3B;
  • Fig. 4A is a profile cross-sectional view of the cassette of Fig. 1 when assembled;
  • Fig. 4B illustrates the details of the hermetic closing of the cassette illustrated in Fig. 4A;
  • Fig. 4C is a front cross-sectional view of the cassette of Fig. 1 when assembled;
  • FIGs. 5A-5B illustrate different embodiments of combs suitable for the first portion of a preferred embodiment of the cassette of the present invention
  • Fig. 6A is an exploded perspective rear view of another embodiment of the cassette of the present invention.
  • Fig. 6B is an exploded perspective front view of another embodiment of the cassette of the present invention.
  • Fig. 6C illustrates a stack of another embodiment of the cassette of the present invention ready for gel casting
  • Fig. 7 illustrates an electrophoresis assay performed with the cassette of Fig. 1 made by stereo-lithography (SLA) technology
  • Fig. 8 illustrates an electrophoresis assay performed with the cassette of Fig. 1 made by injection molding technique. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT OF THE INVENTION
  • a cassette for electrophoresis which is suitable for being precasted. This allows mass production of precast gel, leak free container.
  • the cassette of the present invention is preferably prepared by injection molding in a UV transparent and heat resistant plastic, but one skilled in the art would be aware of alternative fabrication modes that would be suitable for preparation of the cassette of the present invention, such as, but not limited to, forming using extrusion, machining, compression molding, injection molding, reaction injection molding (RIM), blow molding, casting and dipping, rotational molding (rotomolding), thermoforming and cold molding, foaming (foamed thermoplastics or foamed thermosets), thermoforming, pressure forming, vacuum forming including compression molding technique, hot embossing, transfer molding technique, drape forming, snap-back vacuumforming, billow snap-back vacuumforming, straight vacuumforming, plug assist vacuumforming, plug assist pressure forming, billow plug assist vacuum/pressure forming and different techniques like micro molding, plastics blow molding, plastic injection molding or KoolGasTM injection-molding technique, microwave- cured acrylic resins and silicone-gypsum molding technique. It also includes stereo-lithography (SLA), rapid proto-pregne
  • the cassette (10) of the present invention comprises a first portion (12) and a second portion (14) for hermetic engagement together.
  • Figs. 2A-2E illustrate the first portion (12).
  • the first portion (12) has a front lip (16) extending from its front part and a rear lip (18) extending from its back. Moreover, it comprises male projections (20) on its top surface for stacking a plurality of cassettes (10) together, which is further explained in details later in the description of the invention.
  • the inside surface of the first portion (12) comprises a plurality of teeth aligned on the surface of the cassette (10) forming a comb (22).
  • the first portion (12) comprises a insert portion (24) along its periphery.
  • Figs. 3A-3E illustrate the second portion (14).
  • the receiving portion (14) has a second portion opening (28) which, when aligned with the first portion opening (26) upon assembly of the cassette (10), provides an opening useful for the gel casting of the cassette (10).
  • the second portion (14) also comprises front hooks (30) for engaging the front lip (16) when the cassette (10 is assembled, providing therein closing and releasing means for the cassette (10). It is understood that numerous variations of this embodiment could be readily envisioned by one skilled in the art and that such variations are intended to be part of the present invention. For example, only one hook (30) could be used instead of two as illustrated in Fig. 3A. Also, the second portion (14) comprises a rear hook (32) for engaging the rear lip (18) when the cassette (10) is assembled.
  • the second portion (14) further comprises a groove (36) formed in its periphery, providing tight engagement with the insert portion (24) when the cassette (10) is assembled, which results in an hermetic closing of the cassette (10) as illustrated in Figs. 4A-4C.
  • Alternative embodiments could be readily envisioned to one skilled in the art for providing hermetic closing of the cassette (10, such as, but not limited to, providing a gasket between the first portion (12) and the second portion (14). Such alternative embodiments are intended to be part of the present invention.
  • at least one stand (34) is provided on the second portion (14) for allowing the cassette (10 ) to stand vertically and female projections (38) are provided to allow staking of a plurality of cassettes (10) when engaged with the male projections (20).
  • the stand (34) can be part of the rear lip (18) or be part of the rear portion of the first portion (12) itself.
  • Figs. 5A and 5B illustrates different embodiments of the first portion (12) having the comb (22) formed into in order to form channels in the gel upon filing of the cassette (10). Different configurations of combs (22) allow different channel formations.
  • FIGs. 6A and 6B when the cassette (10) is assembled, the front lip (16) engage with the front hook (30), the first portion opening (26) aligns with the second portion opening (28) and the rear lip (20) aligns with the rear hook (32).
  • a plurality of cassette (10) can then be stacked together as illustrated in Fig. 6C by engaging the male projections (20) into the female projections (38), assuring that the plurality of cassette (10) are attached together.
  • the stand (34) provides vertical stability to the cassette.
  • the cassette (10) For casting the cassette (10) with electrophoresis gel, gel is simply poured directly by the opening formed by the alignment of the first portion opening (26) and the second portion opening (28).
  • the comb (22) provided at the bottom of the cassette (10) allows the formation of channels and it is typically positioned at a distance of 0,2254 cm from the bottom.
  • several configurations of comb can be provided, another configuration being illustrated at Fig. 5B. However, all configurations are having in common that upon filing of the cassette, the cassette is standing on a surface opposed to the surface having the opening from which the cassette is filled.
  • the gel is cast in the cassette (10) in the same manner as filling a bottle, making the production process very simple and easy to adapt to any existing liquid handling or industrial filling device. Contrary to actual agarose gel production, all operations are done in this single step; otherwise, it should be required to install the gel tray in position, level it, when required to block both ends with tape or blocks to prevent hot agarose leakage, to pour the hot agarose, to properly position the comb, to let jellified, to remove the comb and if conservation is required, to cover and seal the gel to prevent dehydration.
  • a plug (40) can be inserted in the opening formed by the first portion opening (26) and the second portion opening (28) as illustrated in Fig. 6B.
  • This plug (40) prevents the gel to dry and also prevents contamination of the gel during transportation of the cassette (10).
  • the cassette (10) of the present invention could be designed so that the attachment of the first portion (12) and the second portion (14) is provided by the presence a living hinge between the first portion (12) and the second portion (14). With a cassette of that embodiment, one would only have to break the front lip (16) to access the gel.
  • the cassette of the present invention In applications where contamination is not a concern, it is desirable to use the cassette of the present invention for more than one use, and therefore the front lip (16) and the rear lip (18) could be resilient and engageable and disengageable from the front hook (30) and the rear hook (32) respectively. Again, the rear lip (18) and the rear hook (32) could be replaced by a living hinge between the first portion (12) and the second portion (14).
  • the cassette (10) comprises a mechanism for retaining the gel in the second portion (14) upon opening of the cassette (10).
  • This mechanism can be side walls of the second portion (14) being higher than the side walls of the first portion (12), providing therefore a greater resistance in the second portion (14) and forcing the gel to stay in the second portion (14) upon opening of the cassette (10).
  • cassette of the present invention is intended for use in electrophoresis assays, it is understood that the cassette needs to be prepared with material suitable for use in such assays.
  • This material would be typically a heat resistant and translucent plastic, being also UV transmitting and suitable for injection molding.
  • one skilled in the art of the fabrication of cassettes would understand that variations to this embodiment could be performed without affecting the functionality and/or spirit of the invention.
  • Example 1 Typical characteristic of an agarose gel cassette
  • the cassette (10) measurements are 6,4135 cm length, 5.4064 cm wide and 1.2065 cm thick.
  • the comb measurements are 0.2 cm wide, 0.3969 cm of length and 0.5747 height and is found 0.2254 cm from the base of the cassette (10).
  • Possible variations comprises cassettes of the following measurements: Table 1 Possible measurement variations of the cassette

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  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Molecular Biology (AREA)
  • Electrochemistry (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Physics & Mathematics (AREA)
  • Environmental & Geological Engineering (AREA)
  • Engineering & Computer Science (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
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  • Pathology (AREA)
  • Investigating Or Analysing Biological Materials (AREA)

Abstract

L'invention concerne une cassette destinée à contenir un gel d'électrophorèse comprenant une première partie, une deuxième partie destinée à s'emboîter de manière hermétique avec la première partie, une ouverture permettant le remplissage de la cassette avec du gel. Une des première ou deuxième parties possède une série de dents formant un peigne sur sa surface interne. Ce peigne permet la formation de canaux lorsque la cassette est remplie.
PCT/CA2004/001827 2003-10-17 2004-10-15 Cassette d'electrophorese WO2005036160A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US51164703P 2003-10-17 2003-10-17
US60/511,647 2003-10-17

Publications (1)

Publication Number Publication Date
WO2005036160A1 true WO2005036160A1 (fr) 2005-04-21

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PCT/CA2004/001827 WO2005036160A1 (fr) 2003-10-17 2004-10-15 Cassette d'electrophorese

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Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5164066A (en) * 1987-04-27 1992-11-17 Eg&G Inc. Electrophoresis cassette system with apparatus and method for filling same
US5186807A (en) * 1991-08-13 1993-02-16 Erie Scientific Company Electrophoresis gel slab cassettes having pull cords and methods
CA2381962A1 (fr) * 1999-09-01 2001-03-08 Mirador Dna Design Inc. Cassette d'electrophorese thermoformees a jeter
US6368481B1 (en) * 1997-10-01 2002-04-09 Amersham Pharmacia Biotech Ab Cassette for electrophoresis system
WO2002035224A1 (fr) * 2000-10-26 2002-05-02 Gradipore Limited Ensemble cassette pour gels electrophoretiques

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5164066A (en) * 1987-04-27 1992-11-17 Eg&G Inc. Electrophoresis cassette system with apparatus and method for filling same
US5186807A (en) * 1991-08-13 1993-02-16 Erie Scientific Company Electrophoresis gel slab cassettes having pull cords and methods
US6368481B1 (en) * 1997-10-01 2002-04-09 Amersham Pharmacia Biotech Ab Cassette for electrophoresis system
CA2381962A1 (fr) * 1999-09-01 2001-03-08 Mirador Dna Design Inc. Cassette d'electrophorese thermoformees a jeter
WO2002035224A1 (fr) * 2000-10-26 2002-05-02 Gradipore Limited Ensemble cassette pour gels electrophoretiques

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