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WO2004029338A1 - Appareil de dialyse - Google Patents

Appareil de dialyse Download PDF

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Publication number
WO2004029338A1
WO2004029338A1 PCT/GB2002/003717 GB0203717W WO2004029338A1 WO 2004029338 A1 WO2004029338 A1 WO 2004029338A1 GB 0203717 W GB0203717 W GB 0203717W WO 2004029338 A1 WO2004029338 A1 WO 2004029338A1
Authority
WO
WIPO (PCT)
Prior art keywords
membrane
sample
chamber
solution
protein
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/GB2002/003717
Other languages
English (en)
Inventor
Stephen Lee
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.)
RAYNER PAUL
Original Assignee
RAYNER PAUL
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
Priority claimed from GB0119528A external-priority patent/GB0119528D0/en
Priority claimed from GB0212418A external-priority patent/GB0212418D0/en
Application filed by RAYNER PAUL filed Critical RAYNER PAUL
Priority to AU2002326011A priority Critical patent/AU2002326011A1/en
Anticipated expiration legal-status Critical
Publication of WO2004029338A1 publication Critical patent/WO2004029338A1/fr
Ceased legal-status Critical Current

Links

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/40—Concentrating samples
    • G01N1/4005—Concentrating samples by transferring a selected component through a membrane
    • C—CHEMISTRY; METALLURGY
    • C30—CRYSTAL GROWTH
    • C30B—SINGLE-CRYSTAL GROWTH; UNIDIRECTIONAL SOLIDIFICATION OF EUTECTIC MATERIAL OR UNIDIRECTIONAL DEMIXING OF EUTECTOID MATERIAL; REFINING BY ZONE-MELTING OF MATERIAL; PRODUCTION OF A HOMOGENEOUS POLYCRYSTALLINE MATERIAL WITH DEFINED STRUCTURE; SINGLE CRYSTALS OR HOMOGENEOUS POLYCRYSTALLINE MATERIAL WITH DEFINED STRUCTURE; AFTER-TREATMENT OF SINGLE CRYSTALS OR A HOMOGENEOUS POLYCRYSTALLINE MATERIAL WITH DEFINED STRUCTURE; APPARATUS THEREFOR
    • C30B29/00—Single crystals or homogeneous polycrystalline material with defined structure characterised by the material or by their shape
    • C30B29/54—Organic compounds
    • C30B29/58—Macromolecular compounds
    • C—CHEMISTRY; METALLURGY
    • C30—CRYSTAL GROWTH
    • C30B—SINGLE-CRYSTAL GROWTH; UNIDIRECTIONAL SOLIDIFICATION OF EUTECTIC MATERIAL OR UNIDIRECTIONAL DEMIXING OF EUTECTOID MATERIAL; REFINING BY ZONE-MELTING OF MATERIAL; PRODUCTION OF A HOMOGENEOUS POLYCRYSTALLINE MATERIAL WITH DEFINED STRUCTURE; SINGLE CRYSTALS OR HOMOGENEOUS POLYCRYSTALLINE MATERIAL WITH DEFINED STRUCTURE; AFTER-TREATMENT OF SINGLE CRYSTALS OR A HOMOGENEOUS POLYCRYSTALLINE MATERIAL WITH DEFINED STRUCTURE; APPARATUS THEREFOR
    • C30B7/00—Single-crystal growth from solutions using solvents which are liquid at normal temperature, e.g. aqueous solutions
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/40—Concentrating samples
    • G01N1/4005—Concentrating samples by transferring a selected component through a membrane
    • G01N2001/4016—Concentrating samples by transferring a selected component through a membrane being a selective membrane, e.g. dialysis or osmosis

Definitions

  • the invention relates to apparatus suitable for carrying out dialysis on small volume samples, usually of the microlitre order and more commonly referred to as microdialysis.
  • the invention also relates to methods of microdialysis using the apparatus as well as to other applications of the apparatus such as for protein crystallization either by vapour diffusion or by other means.
  • Such a device includes a sample chamber, which may have a volume of anything from about l ⁇ l to about 300 ⁇ l.
  • the chamber is usually within a circular housing which has a circumferential groove on the external surface for receiving an o-ring, which is used to secure a dialysis membrane across the mouth of the chamber once the sample has been introduced.
  • the device is then contacted with the solution against which the sample is to be dialysed.
  • Such known devices suffer from the disadvantage that they are difficult to handle, both in introduction and removal from the dialysis buffer and in the application of the membrane, where it is easy to trap air bubbles between the membrane and the sample.
  • a further disadvantage is that if one wishes to try out a number of different dialysis conditions, several different receptacles are needed, taking up a lot of bench space. In addition, it is not possible to visualize conditions in the sample chamber while the device is in use, which may be desirable for some applications.
  • the present invention relates to apparatus for microdialysis which overcomes these difficulties.
  • the invention relates to an apparatus suitable for carrying out microdialysis which comprises a chamber for holding a sample, said chamber being disposed in a housing, the external surfaces of which are shaped to secure a dialysis membrane across the mouth of said chamber, said apparatus being characterized in that said housing includes a handle to allow said membrane when secured to be brought into or out of contact with a solution of different ionic strength from said sample and retention means to allow said membrane to be maintained in contact with said solution.
  • the sample chamber may have a volume in the region of about 1 ⁇ l up to about 300 ⁇ l, although the usual range would be between about 5 ⁇ l and about 200 ⁇ l and most preferably between from about 5 ⁇ l to about 20 ⁇ l.
  • the provision of a handle allows for much easier manipulation of the apparatus so that improper application of, or damage to, the membrane is avoided.
  • the sample chamber is cylindrical and is disposed within a housing of circular horizontal cross-section. This allows a circumferential groove to be present on the external surface of the housing so that the dialysis membrane of the appropriate molecular weight cut-off (MWC) may be fitted over the mouth of the chamber and sealed with an o-ring of elastic material such, as rubber or other elastomer.
  • MWC molecular weight cut-off
  • a "snap-on" cap member of sufficiently flexible material, preferably a plastics material, can be used whereby said cap is provided with a circumferential lip member capable of engaging in the circumferential groove in the housing.
  • the cap member is provided with a hole, preferably circular, which, when snapped to the housing, is substantially in alignment with the mouth of the chamber.
  • the external surface of the housing is shaped to provide a screw thread. This allows the membrane to be secured over the mouth of the chamber by a threaded cap member.
  • the cap member is provided with a hole, preferably circular which when screwed to the housing is substantially in alignment with the mouth of the chamber.
  • the top surface of the housing surrounding the sample chamber be flat in order to ensure proper sealing of the chamber by the membrane.
  • the membrane is sealed by means of an ⁇ o'ring the top of the housing may be bevelled rather than flat.
  • a particular advantage of the present apparatus is that it may be sized and shaped so that the sample chamber and membrane fit into a well of a receptacle comprising multiple cylindrical wells and the retention means permits the membrane to be held at an optimum distance from the bottom of the well for efficient dialysis.
  • a multiwell receptacle allows multiple samples to be processed simultaneously while minimizing the amount of space required.
  • Particularly preferred are 24 well tissue culture plates such as the known VDX or Linbro plates. These can be obtained from Hampton Research, 27632, El Lazo Road, Suite 100, Website Georgia Niguel, CA 92677-3913, USA.
  • Preferred retention means to hold the chamber and housing in the correct position within the well include at least one catch member protruding from the housing, providing the apparatus with a diameter greater than the diameter of the well and positioned on the housing between the sample chamber and the handle.
  • the catch member is a circular flange capable of engaging the rim of the well.
  • the flange may include a seal of rubber or other suitable elastomer around its circumference in order to form a seal with the well rim.
  • the handle may be any shape but where the housing is cylindrical it is practical for the handle to be in the form of a rod of circular cross-section. For certain applications it is desirable for the handle to have a flat surface at the end distal to the retention means while, in other cases, it may have an alternative shape as discussed below.
  • the apparatus of the invention may be made from any suitable material, for example mild steel or a plastics material, but preferred is a material that is substantially transparent such as a clear plastics material or glass. A particularly preferred material is perspex which may be suitably moulded.
  • An advantage of the use of a transparent material is that it allows visualisation of the sample chamber through the handle during use.
  • one of the applications of the apparatus of the invention is for crystallization of proteins. By use of the transparent material it is possible to determine the presence of crystals in the sample chamber and/or their rate of growth, without removal of the apparatus from the well and hence disturbance of the crystals.
  • the handle is shaped as a lens so as to provide magnification of the sample.
  • the invention provides a method of carrying out microdialysis of a sample which comprises the steps of : a) introducing a sample into the sample chamber of the apparatus of the invention as described above, b) securing a dialysis membrane across the mouth of said chamber, c) moving the apparatus so as to bring the membrane into contact with a solution of different ionic strength to the sample, said solution being contained in a receptacle, d) creating a seal between the mouth of said receptacle and the retention means of the apparatus and e) allowing dialysis to proceed.
  • the sample is usually placed in the chamber so as to create a slight dome of liquid at the top edge of the chamber.
  • a dialysis membrane chosen with the desired molecular weight cut-off, may then be secured across the mouth of the chamber.
  • a "Golf Tee Applicator” or “CRE Applicator” may be used.
  • a Golf Tee Applicator is available from- Hampton Research, 27632 El Lazo Road, Suite 100, Website Georgia Niguel, CA 92677-3913, USA.
  • the receptacle is a well of a multiwell plate, for example a 24 well tissue culture plate of the VDX or Linbro type, into which the sample loaded apparatus is introduced.
  • the seal is substantially airtight to prevent evaporation of the solutions to the atmosphere.
  • this may be secured by using a grease between the rim of the well and the retention means of the apparatus, in particular between the circular flange and the well rim.
  • an ⁇ o'ring of rubber or other elastomer may be disposed around the circular flange to engage the rim of the well or, as aforesaid the apparatus itself may be provided with a rubber lip seal around the circular flange.
  • the ionic strength of the dialysis buffer may be higher or lower than that of the sample but compounds of smaller size than the pore size of the membrane will diffuse across down a concentration gradient until the system reaches equilibrium.
  • the apparatus of the invention is equally useful for protein crystallization.
  • the invention provides a method of producing a crystallized protein comprising the steps of: a) introducing a solution comprising the protein to be crystallized into the sample chamber of any of the embodiments of the apparatus described above, b) securing a dialysis membrane over the mouth of the chamber, c) bringing the membrane into contact with a " protein precipitating solution, said solution being contained in a receptacle, d) creating a seal between the mouth of the receptacle and the retention means of the apparatus and e) allowing protein crystallization to take place.
  • the protein solution will be introduced into the sample chamber and the membrane applied as described above for the dialysis applications. Similarly it is preferred that the protein precipitating solution will be placed in the well of a 24 well tissue culture plate of the type described above.
  • the apparatus be made of a substantially transparent material as described above so that the formation of crystals in the sample chamber may be viewed through the handle.
  • the handle is shaped so as to provide magnification of the sample.
  • the crystals may be recovered from the sample chamber for further processing, such as for example for X-ray diffraction experiments or the apparatus may be used as a suitable means of storage of the crystals, and for heavy atom/ligand soak experiments.
  • the apparatus of the invention may be used in an alternative method of protein crystallisation using vapour diffusion. Such use requires an embodiment of the apparatus in which the end of the handle distal to the retention means has a flat surface.
  • the apparatus is used in an ⁇ inverted' form in which the handle is introduced into the well of a receptacle with the sample chamber uppermost and exposed to the air.
  • a "hanging drop” comprising a solution of the protein to be crystallised is first applied to the flat surface of the handle and it is then introduced into a well containing a protein precipitant solution in a manner such that there is no contact between the hanging drop and the precipitant solution. An airtight seal is created between the mouth of the well and the retention means. Water vapour from the hanging drop will, by diffusion, equilibrate with the protein precipitant solution and when protein supersaturation occurs, protein crystals will form on the flat end of the handle. Again the crystals may be removed and subject to further processing or may be maintained on the end of the handle as a form of storage device.
  • the flat surface of the handle is siliconized to provide a hydrophobic , water repellant surface in case glass material is used.
  • Figure 1 shows a cross-sectional view of an o- ring
  • Figure 2 shows a cross-sectional view of a dialysis membrane disk
  • FIG. 3 shows a cross-sectional view of one embodiment of the microdialysis apparatus in accordance with the invention
  • Figure 4 shows a cross-sectional view of the whole microdialysis apparatus of Figure 3, with the dialysis membrane disk secured on top of the chamber by an o-ring.
  • FIG. 5 shows a top view of the whole microdialysis apparatus of Figure 3
  • Figure 6 shows a bottom view of the whole microdialysis apparatus of Figure 3
  • Figure 7 shows a cross-sectional view of the grease ring applied on the well rim of a, 24-well tissue culture plate
  • Figure 8 shows a cross-sectional view of an alternative embodiment of the apparatus in which the membrane is secured across the mouth of the chamber by a snap-on cap
  • Figure 9 shows a cross-sectional view of an alternative embodiment of the apparatus in which the external surface of the housing is screw-thread for receipt of the threaded cap member
  • FIG. 10 shows crystals of glucose isomerase prepared using the apparatus in accordance with the invention.
  • Figure 11 shows crystals of lyzozyme prepared using apparatus in accordance with the invention.
  • the microdialysis apparatus of the invention comprises a chamber 1 for holding the sample, a circumferential groove 2 shaped for retaining an elastic* o-ring, a retention means in the form of a circumferential flange 3 and a handle portion 4.
  • a chamber 1 for holding the sample
  • a circumferential groove 2 shaped for retaining an elastic* o-ring
  • a retention means in the form of a circumferential flange 3
  • a handle portion 4 As shown in Figure 4, to use the apparatus the dialysis membrane 5 is applied across the top of the apparatus to cover the mouth of the chamber 1 and is secured to the housing by an o-ring 6.
  • Figure 7 the apparatus is shown in use, inserted into the well 7 of a 24-well tissue culture plate 9.
  • Grease 8 is applied between the circumferential flange 3 and the rim of the well 7 to create an airtight seal.
  • FIG. 8 An alternative embodiment of the apparatus is shown in Figure 8 in which the o-ring 6 is replaced by a "snap-on" cap 12, said cap having a circumferential lip 14 which engages with the groove 2 in the housing.
  • the cap 12 has a hole 14 which aligns with the mouth of the chamber 1 when the cap is snapped on to the apparatus housing.
  • FIG. 9 A further alternative embodiment of the apparatus is shown in Figure 9 in which the circumferential groove 2 is replaced by screw thread 15.
  • the dialysis membrane 5 is then secured to the apparatus by means of a screw threaded cap 10 having a hole 11 which aligns with the mouth of the chamber 1 when the cap is screwed to the apparatus housing.
  • the embodiment is used in exactly the same manner as the previously described embodiment.

Landscapes

  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Crystallography & Structural Chemistry (AREA)
  • Materials Engineering (AREA)
  • Metallurgy (AREA)
  • Organic Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Pathology (AREA)
  • Sampling And Sample Adjustment (AREA)

Abstract

L'invention concerne un appareil capable de réaliser une microdialyse et qui comprend une chambre (1) permettant de maintenir un échantillon, disposé dans un logement, les surfaces externes de ce dernier étant conçues pour fixer une membrane (5) de dialyse à travers l'ouverture de la chambre (1). Ledit logement comprend une poignée (4) permettant à la membrane (5), lorsqu'elle est fixée, d'être placée en contact ou hors contact avec une solution à force ionique, différente de l'échantillon, et des moyens de retenue (3), de sorte que la membrane (5) est maintenue en contact avec la solution. L'appareil est utilisé pour réaliser des expériences de cristallisation des protéines.
PCT/GB2002/003717 2001-08-10 2002-08-08 Appareil de dialyse Ceased WO2004029338A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
AU2002326011A AU2002326011A1 (en) 2001-08-10 2002-08-08 Dialysis apparatus

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
GB0119528A GB0119528D0 (en) 2001-08-10 2001-08-10 Microdialysis rod
GB0119528.8 2001-08-10
GB0212418A GB0212418D0 (en) 2002-05-29 2002-05-29 Dialysis Apparatus
GB0212418.8 2002-05-29

Publications (1)

Publication Number Publication Date
WO2004029338A1 true WO2004029338A1 (fr) 2004-04-08

Family

ID=32044446

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/GB2002/003717 Ceased WO2004029338A1 (fr) 2001-08-10 2002-08-08 Appareil de dialyse

Country Status (2)

Country Link
AU (1) AU2002326011A1 (fr)
WO (1) WO2004029338A1 (fr)

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2011042627A1 (fr) * 2009-10-09 2011-04-14 Universite De Franche-Comte Insert de diffusion pour l'analyse de membrane, kit cellule et procede de diffusion
FR2964671A1 (fr) * 2010-09-15 2012-03-16 Univ Grenoble 1 Dispositif et procede de cristallisation de substances inorganiques ou organiques
WO2013057401A1 (fr) * 2011-10-10 2013-04-25 Universite Claude Bernard Lyon 1 Dispositif d'etude de la permeabilite de membranes biologiques, artificielles ou synthetiques
US20190201846A1 (en) * 2017-12-30 2019-07-04 University Of Kansas Apparatus, systems, and methods for isotope exchange and/or dialysis
WO2024130038A1 (fr) * 2022-12-14 2024-06-20 The Regents Of The University Of California Dispositif de croissance d'échantillon suspendu pour applications d'imagerie

Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0265319A1 (fr) * 1986-10-09 1988-04-27 AEROSPATIALE Société Nationale Industrielle Procédé, cellule et dispositif de cristallogénèse, notamment par vaisseau spatial
US5106592A (en) * 1989-12-13 1992-04-21 Dornier Gmbh Manipulating liquids for growing crystals in outer space
US5400741A (en) * 1993-05-21 1995-03-28 Medical Foundation Of Buffalo, Inc. Device for growing crystals
US5552127A (en) * 1994-07-25 1996-09-03 Fujitsu, Ltd. Biological macromolecular substances crystallization apparatus
US5641681A (en) * 1995-04-17 1997-06-24 The United States Of America As Represented By The Administrator Of The National Aeronautics And Space Administration Device and method for screening crystallization conditions in solution crystal growth
US5643540A (en) * 1995-02-27 1997-07-01 The United States Of America As Represented By The Administrator Of The National Aeronautics And Space Administration Protein crystal growth apparatus for microgravitiy

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0265319A1 (fr) * 1986-10-09 1988-04-27 AEROSPATIALE Société Nationale Industrielle Procédé, cellule et dispositif de cristallogénèse, notamment par vaisseau spatial
US5106592A (en) * 1989-12-13 1992-04-21 Dornier Gmbh Manipulating liquids for growing crystals in outer space
US5400741A (en) * 1993-05-21 1995-03-28 Medical Foundation Of Buffalo, Inc. Device for growing crystals
US5552127A (en) * 1994-07-25 1996-09-03 Fujitsu, Ltd. Biological macromolecular substances crystallization apparatus
US5643540A (en) * 1995-02-27 1997-07-01 The United States Of America As Represented By The Administrator Of The National Aeronautics And Space Administration Protein crystal growth apparatus for microgravitiy
US5641681A (en) * 1995-04-17 1997-06-24 The United States Of America As Represented By The Administrator Of The National Aeronautics And Space Administration Device and method for screening crystallization conditions in solution crystal growth

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
AIBARA S: "Crystallization of wheat gamma-gliadin under a microgravity environment using space station MIR", JOURNAL OF CRYSTAL GROWTH, NORTH-HOLLAND PUBLISHING CO. AMSTERDAM, NL, vol. 155, no. 3, 1 October 1995 (1995-10-01), pages 247 - 253, XP004000775, ISSN: 0022-0248 *

Cited By (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2011042627A1 (fr) * 2009-10-09 2011-04-14 Universite De Franche-Comte Insert de diffusion pour l'analyse de membrane, kit cellule et procede de diffusion
FR2964671A1 (fr) * 2010-09-15 2012-03-16 Univ Grenoble 1 Dispositif et procede de cristallisation de substances inorganiques ou organiques
WO2012035268A1 (fr) 2010-09-15 2012-03-22 Universite Joseph Fourier (Grenoble 1) Dispositif et procédé de cristallisation de substances inorganiques ou organiques
US9382639B2 (en) 2010-09-15 2016-07-05 Universite Joseph Fourier Device and method for crystallizing inorganic or organic substances
WO2013057401A1 (fr) * 2011-10-10 2013-04-25 Universite Claude Bernard Lyon 1 Dispositif d'etude de la permeabilite de membranes biologiques, artificielles ou synthetiques
US20190201846A1 (en) * 2017-12-30 2019-07-04 University Of Kansas Apparatus, systems, and methods for isotope exchange and/or dialysis
WO2024130038A1 (fr) * 2022-12-14 2024-06-20 The Regents Of The University Of California Dispositif de croissance d'échantillon suspendu pour applications d'imagerie

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