Classifying – separating – and assorting solids – Precedent preparation of items or materials to facilitate... – Sorting special items or sorting by methods and apparatus...
Reexamination Certificate
1999-12-02
2001-12-25
Ellis, Christopher P. (Department: 3653)
Classifying, separating, and assorting solids
Precedent preparation of items or materials to facilitate...
Sorting special items or sorting by methods and apparatus...
C209S552000
Reexamination Certificate
active
06332540
ABSTRACT:
FIELD OF THE INVENTION
This invention relates to a novel device suitable for separating polymeric beads from a suspension of a plurality of such beads in a fluid.
BACKGROUND OF THE INVENTION
In chemistry and biochemistry chemical reactions are frequently performed on the surface of small polymeric resin beads of a diameter range 50-600 microns, typically 250-300 microns in diameter. For example an active substance may be chemically absorbed onto such a bead, e.g. on its surface and/or into the body of the bead, the treated bead may then be exposed to further active substances or labels such as fluorescence labels, and the occurrence of an interaction between the various active substances and/or labels can be detected by monitoring the bead. This technique is particularly useful in biological sciences where only minute amounts of such active substances may be available, and in the investigation of combinatorial chemistry libraries.
In such techniques it is usually necessary to transfer individual beads from a source containing many such beads, e.g. a suspension (which term as used herein includes inter alia a slurry) of the beads in a liquid, into defined locations in a receptacle, for example individual vials, or individual wells in a microtitre plate.
A number of problems hinder such transfer. The small size of the beads makes them difficult to manipulate. It is difficult to transfer beads using present techniques without at the same time transferring a relatively large quantity of liquid. Often large numbers of beads are involved, and present methods of transferring beads suffer from the problems inherent in handling large numbers of small beads. At present either manual methods, e.g. picking out single beads by hand, or excessively complex devices are used.
One device for automatically separating beads from a suspension of a plurality of beads in suspension in a fluid is disclosed in WO 94/28119, which is a large scale device, primarily intended for use with beads of diameter ca. 3 mm, and for separating beads containing encapsulated biomass from empty beads. Devices for automatically separating smaller particles, e.g. cells suspended in a fluid, are disclosed in U.S. Pat. No. 4,756,427, U.S. Pat. No. 5,030,002 and U.S. Pat. No. 4,175,662. It is considered unlikely that the bead sorting devices disclosed in these last mentioned US patents could be adapted directly for use in separating the above mentioned polymeric beads used in chemical reactions. Moreover they all suffer from the disadvantage that the suspended particles remain in a substantial bulk of suspension fluid even after separation. This means that if the particles are to be delivered to a particular location, e.g. a specified well in a microtitre plate, this excess of fluid accompanies the particle to the location.
It is an object of this invention to provide a device which is more suitable for operation with smaller volumes of suspension and for the above-mentioned smaller polymeric beads. It is also an object of this invention to provide a device which can separate beads and can be used with a range of sizes of beads, i.e. not being limited to a particular size of bead.
SUMMARY OF THE INVENTION
According to this invention, a device suitable for separating polymeric beads from a suspension of a plurality of such beads in a fluid, comprises:
a flow cell through which a suspension of the beads in the fluid can flow;
the flow cell having an inlet end through which the suspension of beads may be introduced into the flow cell, such that the beads are arranged in a successive serial flow stream as they flow through the flow cell toward an exit end of the cell;
a radiation source capable of directing incident radiation into the flow cell in an incident direction such that as beads flow through the flow cell they intercept the incident radiation;
a radiation detector positioned relative to the radiation source and to the flow of beads such that a change in the radiation detected by the detector occurs as the bead intercepts the incident beam;
a bead outlet channel and a fluid outlet channel located downstream from the point where the beads intercept the incident radiation, and being in communication with the exit end;
valve means which in a first configuration directs fluid flow preferentially through the bead outlet channel rather than through the fluid outlet channel, and which in a second configuration directs fluid flow preferentially through the fluid outlet channel rather than through the bead outlet channel, so that in the first valve configuration beads are caused to flow through the bead outlet channel;
control means responsive to a signal from the radiation detector resulting from a bead intercepting the incident beam, and which directs the valve means into the first valve configuration so that a bead flows through the bead outlet channel, and after the bead is downstream of the valve means then directs the valve means into the second valve configuration.
DETAILED DESCRIPTION OF THE INVENTION
The device is preferably used for separating polymeric beads of the type used in chemical and biochemical reactions, as mentioned above, e.g. of a diameter range 50-600 microns, typically 250-300 microns, suspended in a suspension fluid which is a mobile liquid such as water or preferably an alcohol such as methanol or ethanol.
The flow cell is suitably a cell having at least part if not all of its walls transparent to the incident radiation so that the radiation source may be located outside the flow cell. If for example the incident radiation is light, then such transparent parts or the entire cell may be made of transparent materials such as plastics materials, glass, quartz etc. In some modes of operation the detector may be responsive to secondary radiation which is of a different wavelength to the incident radiation, e.g. if as discussed below the detector is responsive to a fluorescence emission, and in this case it may be necessary for at least part if not all of the walls of the cell to be transparent to the secondary radiation. The flow cell is suitably in the form of a tube of narrow internal bore, of dimensions across the direction of flow of the beads about 1.5-4 times the diameter of the beads, for example in the case of 250-300 microns diameter beads, preferably around 600-1500 microns across. Best results appear to be achieved with a narrow bore, but not so narrow that blockage is likely.
Suitably the flow cell is a transparent walled cell having the part of its wall which faces the incoming direction of the incident radiation, and/or which face the radiation detector, substantially flat and normal to this direction. Suitably therefore the flow cell may be polygonal in cross section having opposed flat faces, particularly rectangular (which term includes square) in cross section, with the flow direction along the longitudinal tube axis. Alternatively the flow cell may have the part of its wall which faces the incoming direction of the incident radiation, and/or which face the radiation detector, of a lens shape, so that such part(s) of the wall may act as a lens and may assist in guiding the incident radiation towards the flow of beads and/or toward the radiation detector.
The suspension of beads may be introduced into the inlet end from a source by any convenient means, e.g. by injection from a reservoir such as a storage vessel or a syringe, or by means of a feeder tube etc. In a reservoir it may be necessary to provide some means such as a stirrer to keep the beads in suspension, i.e. to prevent them settling. The rate at which the beads are introduced into the inlet end may be controlled by for example a controlled pump, e.g. a metering pump or syringe pump or peristaltic pump, or by means of air pressure. The device may be constructed such that multiple samples of suspensions of beads, each sample respectively initially contained in a reservoir which is connectable with the device, may be introduced into the device, either sequentially or otherwise. A carousel of such reservoirs may be provided, and
James Philip John
Paul Frank
Payne Richard C
Ellis Christopher P.
Kinzig Charles M.
Rodriguez Joseph
SmithKline Beecham p.l.c.
Stein-Fernandez Nora
LandOfFree
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