U.S. patent application number 10/435891 was filed with the patent office on 2003-10-30 for hydrodynamic injector.
This patent application is currently assigned to Symyx Technologies, Inc.. Invention is credited to Cong, Peijun, Doolen, Robert D., Wheeler, Tony N..
Application Number | 20030201181 10/435891 |
Document ID | / |
Family ID | 24488252 |
Filed Date | 2003-10-30 |
United States Patent
Application |
20030201181 |
Kind Code |
A1 |
Cong, Peijun ; et
al. |
October 30, 2003 |
Hydrodynamic injector
Abstract
A hydrodynamic injector for substantially concurrently loading
fluid samples to be analyzed into multiple capillary tubes of a
capillary electrophoresis system. The injector includes an
enclosure defining a pressure chamber for holding multiple
receptacles, each containing a fluid sample, and apertures in the
enclosure for passing capillary tubes into a position inside the
pressure chamber and in fluid communication with the samples in
respective receptacles. Electrodes on the enclosure extend into the
pressure chamber for reception in the receptacles. The pressure
chamber is pressurized with gas to substantially concurrently force
the fluid samples from respective receptacles into the capillary
tubes in preparation for a capillary electrophoresis operation.
Inventors: |
Cong, Peijun; (San Jose,
CA) ; Doolen, Robert D.; (Sunnyvale, CA) ;
Wheeler, Tony N.; (Santa Clara, CA) |
Correspondence
Address: |
SENNIGER POWERS LEAVITT AND ROEDEL
ONE METROPOLITAN SQUARE
16TH FLOOR
ST LOUIS
MO
63102
US
|
Assignee: |
Symyx Technologies, Inc.
|
Family ID: |
24488252 |
Appl. No.: |
10/435891 |
Filed: |
May 12, 2003 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10435891 |
May 12, 2003 |
|
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|
09620987 |
Jul 21, 2000 |
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6572750 |
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Current U.S.
Class: |
204/453 ;
204/604 |
Current CPC
Class: |
G01N 27/44743
20130101 |
Class at
Publication: |
204/453 ;
204/604 |
International
Class: |
G01N 027/403; G01N
027/453 |
Claims
What is claimed is
1. A hydrodynamic injector for substantially concurrently loading
fluid samples to be analyzed into multiple capillary tubes of a
capillary electrophoresis system, said tubes having first and
second ends, said injector comprising: an enclosure defining a
pressure chamber for holding multiple receptacles, each containing
a fluid sample therein, apertures in the enclosure for passing
capillary tubes into a position wherein first ends of the tubes are
positioned in said pressure chamber in fluid communication with the
samples in respective receptacles, electrodes on the enclosure
extending into the pressure chamber for reception in said
receptacles; said enclosure having a gas inlet for pressurizing the
pressure chamber whereby said fluid samples are substantially
concurrently forced from respective receptacles into the first ends
of respective capillary tubes in preparation for a capillary
electrophoresis operation.
2. An injector as set forth in claim 1 wherein said enclosure
includes a movable part movable between an open position away from
the enclosure to permit access to said receptacles and a closed
position in sealing relation with the enclosure so that the
pressure chamber can be pressurized.
3. An injector as set forth in claim 2 wherein said movable part is
configured for supporting a tray having said multiple receptacles
therein.
4. An injector as set forth in claim 3 further compressing a power
actuator for moving said moveable part between said open and closed
positions.
5. An injector as set forth in claim 4 wherein said movable part
has a surface with a recess therein sized and shaped to receive
said tray and to hold it in a fixed predetermined portion relative
to said movable part.
6. An injector as set forth in claim 1 further comprising a channel
in said enclosure surrounding the pressure chamber and
communicating with said gas inlet, and passages connecting the
channel and the pressure chamber at spaced intervals around the
pressure chamber.
7. An injector as set forth in claim 1 further comprising means for
fixedly and sealingly securing the capillary tubes in respective
apertures in the enclosure.
8. An injector as set forth in claim 1 further comprising a gas
supply system for supplying pressurized gas to said pressure
chamber, said gas supply system comprising a source of pressurized
gas, an accumulator having an inlet and an outlet, an accumulator
inlet line connecting the source of pressurized gas to the inlet of
the accumulator, an accumulator outlet line connecting the
accumulator to the gas inlet of the enclosure, and a valve in the
accumulator outlet line movable from a closed position to an open
position after the accumulator has been pressurized for effecting
the transfer of gas under pressure from the accumulator to the
pressure chamber of the enclosure.
9. An injector as set forth in claim 8 further comprising a
pressure controller in said accumulator inlet line for controlling
the pressure in said accumulator.
10. An injector as set forth in claim 9 wherein said pressure
chamber has a first volume and said accumulator has a second
volume, the ratio of said first volume to said second volume being
in the range of 5:1-1:5.
11. An injector as set forth in claim 10 wherein said ratio is in
the range of 2:1-1:2.
12. An injector as set forth in claim 11 wherein said ratio is
about 1:1.
13. An injector as set forth in claim 8 wherein said accumulator
outlet line and the gas inlet of the enclosure are sized so that
pressure equilibrium in said pressure chamber is reached at about
one second after said valve is opened.
14. An injector as set forth in claim 8 further comprising a vent
system for venting the accumulator and said pressure chamber when
said valve is open.
15. An injector system as set forth in claim 1 wherein said
enclosure is a first enclosure and said system further comprises a
second enclosure having a second chamber therein containing one or
more receptacles for receiving fluid samples transmitted through
the capillary tubes from the first enclosure, apertures in the
second enclosure for passing said capillary tubes into a position
wherein second ends of the tubes are positioned in said second
chamber for the flow of fluid into said one or more receptacles,
and electrodes on the second enclosure extending into the second
chamber for reception in said one or more receptacles.
16. An injector as set forth in claim 15 wherein said second
pressure chamber is a pressure chamber adapted for holding multiple
receptacles, each containing a fluid sample therein, said second
enclosure having a gas inlet for pressurizing the pressure chamber
whereby said fluid samples in the second enclosure are
simultaneously forced from respective receptacles into the second
ends of respective capillary tubes in preparation for a capillary
electrophoresis operation.
17. An injector as set forth in claim 16 in combination with said
capillary tubes, said tubes extending between said first and second
enclosures and having portions defining a detection window for the
passage of light therethrough, said detection window being at a
location closer to one enclosure than the other.
18. A method of capillary electrophoresis involving the
substantially concurrent transfer of fluid samples from multiple
receptacles into first ends of multiple capillary tubes, said
method comprising: positioning the first ends of the capillary
tubes and the receptacles in a single pressure chamber so that the
first ends are in fluid communication with the samples in the
receptacles, pressurizing the pressure chamber to force fluid from
the receptacles into the capillary tubes, and causing an electric
current to flow through the capillary tubes and contents thereof to
cause a first capillary electrophoresis operation.
19. A method as set forth in claim 18 wherein said pressurizing
step comprises pressurizing an accumulator to a predetermined
pressure, and then establishing gas flow communication between the
accumulator and said pressure chamber to pressurize the pressure
chamber.
20. A method as set forth in claim 19 wherein said pressurizing
step is carried out so that the pressure in said pressure chamber
reaches equilibrium in no longer than about one second after said
gas flow communication is established.
21. A method as set forth in claim 20 further comprising sizing the
volumes of the pressure chamber and accumulator so that the ratio
of the pressure chamber volume to said accumulator volume is in the
range of 5:1-1:5.
22. A method as set forth in claim 21 wherein said ratio is in the
range of 2:1-1:2.
23. A method as set forth in claim 22 wherein said ratio is about
1:1.
24. A method as set forth in claim 18 wherein said pressure chamber
is a first pressure chamber, said method further comprising
positioning second ends of the capillary tubes in a second pressure
chamber containing multiple receptacles for receiving said second
ends.
25. A method as set forth in claim 24 further comprising causing a
second electrophoresis operation by pressurizing the second
pressure chamber to a pressure greater than the pressure in said
first pressure chamber to force fluid from the receptacles in the
second pressure chamber into the second ends of the capillary
tubes, and causing an electric current to flow through the
capillary tubes and contents thereof to cause capillary
electrophoresis during which said fluid flows from the second
pressure chamber to said first pressure chamber.
26. A method as set forth in claim 24 further comprising detecting
the flow of said fluid samples as they move through the capillary
tubes at a location other than midway between said first and second
pressure chambers.
Description
BACKGROUND OF THE INVENTION
[0001] This invention is generally in the field of capillary
electrophoresis, and relates particularly to apparatus and method
for substantially concurrently loading fluid samples to be analyzed
into multiple capillary tubes of a multiplexed or "parallel"
capillary electrophoresis system.
[0002] Capillary electrophoresis (CE) is a chemical separation
technique involving the use of one or more capillary tubes.
Parallel CE, a recently developed technique using many parallel
capillary tubes, is growing in popularity since this technology
allows multiple samples to be analyzed quickly and efficiently.
This is particularly advantageous in combinatorial chemistry where
many hundreds and even thousands of samples are analyzed over a
short period of time. Parallel CE involves the use of a "bundle" of
capillary tubes, e.g., 96 such tubes. A chemical sample to be
analyzed is loaded in each tube, and a high voltage is applied to
the tube, causing the components of the sample to migrate in the
tube at different speeds, thereby causing separation of the
components which can then be analyzed by conventional light
absorption or other techniques. Reference may be made to the
following patents and publications for a more detailed description
of CE, including parallel CE, and various analytical techniques
used in CE: U.S. Pat. Nos. 5,900,934, 5,324,401, 5,312,535,
5,303,021, 5,239,360; C. Culbertson et al., Analytical Chemistry,
70, 2629-2638 (1998); and X. Gong et al., Analytical Chemistry,
71(21); 4989-4996 (1999).
[0003] In prior multiplexed CE systems, the capillary tubes have
been loaded with liquid samples either hydrostatically (i.e., by
siphoning) or electrokinetically. However, these methods have
various drawbacks, and there is a need for an improved loading
system which is more reliable, reproducible and versatile.
SUMMARY OF THE INVENTION
[0004] Among the several objects of this invention may be noted the
provision of a hydrodynamic injector for loading liquid samples
into the inlet ends of multiple capillary tubes in preparation for
a CE operation; the provision of such an injector which loads
capillary tubes substantially concurrently and very quickly using a
standard microtiter plate; the provision of such an injector which
is easy to operate; the provision of such an injector which can be
used to flush and condition the capillary tubes prior to sample
loading; the provision of such an injector which is safe to use;
the provision of two such injectors which can be used at opposite
ends of the capillary tubes to enable loading from both ends of the
tubes; and the provision of a method of simultaneously transferring
liquid samples into the inlet ends of multiple capillary tubes to
carry out a CE operation.
[0005] In general, the present invention is directed to a
hydrodynamic injector for substantially concurrently loading fluid
samples to be analyzed into multiple capillary tubes of a capillary
electrophoresis system, the tubes having first and second ends. The
injector comprises an enclosure defining a pressure chamber for
holding multiple receptacles, each containing a fluid sample
therein, and apertures in the enclosure for passing capillary tubes
into a position wherein first ends of the tubes are positioned in
the pressure chamber in fluid communication with the samples in
respective receptacles. Electrodes on the enclosure extend into the
pressure chamber for reception in the receptacles. The enclosure
has a gas inlet for pressurizing the pressure chamber whereby the
fluid samples are substantially concurrently forced from respective
receptacles into the first ends of respective capillary tubes in
preparation for a capillary electrophoresis operation.
[0006] A method of this invention generally involves the
substantially concurrent transfer of fluid samples from multiple
receptacles into first ends of multiple capillary tubes. The method
comprises positioning the first ends of the capillary tubes and the
receptacles in a single pressure chamber so that the first ends are
in fluid communication with the samples in the receptacles,
pressurizing the pressure chamber to force fluid from the
receptacles into the capillary tubes, and causing an electric
current to flow through the capillary tubes and contents thereof to
cause a first capillary electrophoresis operation.
[0007] Other objects and features of this invention will be in part
apparent and in part pointed out hereinafter.
BRIEF DESCRIPTION OF THE DRAWINGS
[0008] FIG. 1 is a schematic view of a parallel CE system using a
pair of hydrodynamic injectors of the present invention;
[0009] FIG. 1A is a plan view of a cooling body and an overlying
array of capillary tubes;
[0010] FIG. 2 is an enlarged vertical section on line 2-2 of FIG. 1
showing a hydrodynamic injector in an open position;
[0011] FIG. 2A is a view similar to FIG. 2 showing the injector in
a closed position;
[0012] FIG. 3 is a plan view of a support block of the injector, a
microtiter plate being shown in phantom positioned in a recess in
the block;
[0013] FIG. 4 is a plan view of a power plate of the injector;
[0014] FIG. 5 is an enlarged fragmentary portion of FIG. 2
illustrating how the capillary tubes are affixed to the power
plate;
[0015] FIG. 6 is a bottom view of a sealing block of the injector,
a microtiter plate being shown in phantom received inside an
opening in the sealing block;
[0016] FIG. 7 is a perspective of the sealing block with portions
of a seal being broken away to show details; and
[0017] FIG. 8 is a section on line 8-8 of FIG. 4 showing the
injector in a closed position.
[0018] Corresponding parts are designated by corresponding
reference numbers throughout the drawings.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT
[0019] Referring now to the drawings, FIG. 1 shows a multiplexed
(parallel) capillary electrophoresis (CE) system, generally
indicated at 1, for simultaneously separating and analyzing the
components of multiple chemical samples. The system comprises a
pair of hydrodynamic injectors of the present invention, the
injector on the left as viewed in FIG. 1 being generally designated
3L and the injector on the right 3R. The system also includes a
bundle 5 of capillary tubes 7 having left end portions (as viewed
in FIG. 1) attached to the left injector 3L and right end portions
attached to the right injector 3R. As will be explained in more
detail later in this description, one or both of these injectors
3L, 3R can be used to substantially concurrently load samples to be
analyzed into the capillary tubes 7 prior to the separation phase
of an electrophoresis operation. The capillary tubes 7 have
intermediate portions between the left and right end portions
arranged in a generally planar, ribbon-like array 11 in which the
intermediate portions extend side-by-side in closely spaced
generally parallel relation (FIG. 1A). The array 11 lies in a
horizontal plane as viewed in FIG. 1.
[0020] The system 1 also includes a power source 15 for applying a
potential (voltage) difference between the ends of the capillary
tubes 7 to cause an electrical current to flow through the contents
of the tubes, a light source 19 for emitting light to pass through
the closely spaced array 11 of intermediate portions of the
capillary tubes, and a photodetector generally designated 21
comprising photodetector elements (not shown) for receiving light
passing through the planar array 11 of intermediate portions of the
capillary tubes. Light passing through the tubes is imaged on the
photodetector 21 by an imaging lens, generally designated 25.
[0021] More specifically, the capillary bundle 5 may comprise a
series of 96 capillary tubes 7, although this number may vary. For
example, the capillary bundle 5 can include 8 or more capillaries,
16 or more, 24 or more, but more preferably 48 or more and most
preferably 96 or more capillaries. Each tube is of relatively small
diameter (e.g., 150 microns OD; 75 microns ID) and of a suitable
electrically nonconductive material, such as fused silica so that
high voltages can be applied across tube without generating
excessive heat. The tubes 7 may have a polyimide coating which is
removed by a laser beam, for example, in an area extending across
the planar array of intermediate portions of the capillary tubes,
thereby forming what may be referred to as a detection window (27
in FIG. 1A) which is transparent or translucent so that light from
the light source can pass through the walls of the tubes at this
location. Alternatively, the tubes 7 may be translucent or
transparent along their entire lengths. The bundle 5 in the area
adjacent the detection window 27 may be cooled by a suitable
conductive cooling body 29 having a window 31 therein generally in
alignment with the detection window. The cooling body 29 may be a
thermoelectric device. Alternatively, the cooling body 29 may be
cooled by coolant flowing through passages in the body, as
described in copending application Ser. No. ______, filed ______,
2000. The bundle 5 is of any appropriate length (e.g., 10 cm-2 m).
As illustrated in FIG. 1, the bundle 3 may be supported by suitable
supports 35 on opposite sides of the cooling body 29. The capillary
tubes of the bundle may be held in the aforementioned planar array
11 by any suitable means, such as by strips of adhesive tape (not
shown) extending across the array on opposite sides of the
detection window 27.
[0022] The light source 19 may be of any suitable type, such as a
deuterium or tungsten lamp or a 254-nm mercury lamp, emitting light
having a certain wavelength (e.g., 200-800 nm and generalizable to
other wavelengths) corresponding to the absorption band of the
sample components of interest. The light is typically ultraviolet
or visible light. Light emitted from the source 19 is adapted to
pass through the detection window 27 and through the window of the
cooling body 29 for incidence on the imaging lens 25 and
photodetector 21 therebelow.
[0023] The photodetector 21 is of a conventional type, such as a
photodiode device, having the aforementioned photodetector
elements. These elements may be photodiodes, for example, arranged
in one or more linear rows. For example, the photodetector may be a
model C5964 multichannel detector head by Hamamatsu incorporating a
linear image sensor chip, a low-noise driver/amplifier circuit, and
a temperature controller. In this example, the linear image sensor
chip has 1024 diodes, each of which is 25 microns in width and 2500
microns height. Other types of photodetectors can be used without
departing from the scope of this invention. The photodetector
elements generate output signals which are then transmitted to a
digital processor 35 (FIG. 1) and related equipment (e.g., a
computer 37) for generating and displaying an electropherogram 39,
i.e., a plot of light intensity versus time, as will be understood
by those skilled in this field. This plot can then be evaluated to
identify components of interest in the samples being analyzed. As
shown in FIG. 1, the electropherogram can be displayed on a screen
41 of the computer 37. Optionally, to improve the quality of the
electropherogram, the photodetector 21 may be mounted for selective
rotation about an axis generally perpendicular to the plane of the
detection window 27, as described in copending U.S. patent
application Ser. No. ______, filed Jul. 20, 2000, incorporated
herein by reference.
[0024] The imaging lens 25 may also be of conventional design, such
as a quartz lens (Sodern f.1.=94 mm; F=4.1) in combination with an
interference filter 45 (Oriel) employed to define the absorption
wavelength. The lens 25 is positioned between the detection window
27 and the photodetector 25 to receive light passing through the
capillary tubes 7 and to image that light on the linear array 11 of
photodetector elements. The image of the capillary tubes 7
projected by the lens on the photodetector 25 may be an image 1.5
times actual size, for example.
[0025] The left and right injectors 3L, 3R are essentially
identical in construction, so only one will be described. As shown
in FIGS. 2 and 4, each injector 3L, 3R comprises a rectangular
metal power plate 51 connected to the power source 15 by suitable
electrical cable 53. The power plate may be of copper, for example.
The power plate 51 is supported by four legs 57, one at each corner
of the plate, having lower ends secured to a base plate 59. The
legs are affixed to the power plate and base by fasteners 61 and
63, respectively. An annular sealing block 67 is fastened by
suitable means (e.g., fasteners 69) to the underside of the power
plate 51. The sealing block 67 is of a suitable dielectric
material, such as Delrin.RTM. plastic. The injector 3 also includes
a thick support block 73 of dielectric material (e.g., Delring
plastic) for supporting a series of receptacles 75 containing
samples to be analyzed. These receptacles may be the wells 75 of a
standard 96-well microtiter plate 77, for example, although it will
be understood that other receptacles may be used. The support block
73 is movable between a raised position (FIG. 2A) in which it is in
sealing engagement with the sealing block 67 and a lowered (FIG. 2)
position in which it is spaced from the sealing block. When the
support block 73 is in its raised position, an enclosure (generally
designated 81 in FIG. 2A) is formed defining a pressure chamber 83
receiving the microtiter plate 77 therein, the walls of the
pressure chamber being formed by the bottom surface of the power
plate 51, the interior walls of the annular sealing block 67, and
the top surface of the support block 73. The pressure chamber 83 is
sealed by an upper annular seal 87 between the power plate 51 and
the sealing block 67 and by a lower annular seal 89 between the
sealing block and the support block.
[0026] As shown in FIGS. 2A and 3 the support block 73 has a recess
91 in its upper surface for holding the microtiter plate 77 or
whatever other container is used for holding the samples. The
outline of the recess 91 should be sized to hold the microtiter
plate in a fixed predetermined position for reasons which will
become apparent.
[0027] Referring to FIG. 5, the power plate 51 has a series of
vertical apertures 95 (holes) therein for passage of the capillary
tubes 7 through the power plate to a position in which the tubes
extend down from the plate for reception in respective wells 75 of
the microtiter plate 77 when the support block 73 is in its raised
position forming the aforementioned pressure chamber 83. Each
capillary tube 7 is secured in position by a fitting comprising a
stub screw 99 threaded in the upper end of a respective hole 95 and
having an axial passage through it for receiving the tube 7, and a
ferrule 101 in the hole 95 below the screw 99 having a conical
surface 103 engageable with a tapered shoulder 105 in the hole. The
design is such that threading the screw 99 down in the hole 95
against the ferrule 101 drives the conical surface 103 of the
ferrule into sealing engagement against the tapered shoulder 105 of
the hole and wedges the axial opening in the ferrule closed against
the capillary tube 7 to clamp the tube securely in place without
crushing or otherwise blocking the tube. The ferrule also forms a
seal against the tube and power plate to prevent leakage through
the hole 95. Other means may be used for sealingly securing the
tube 7 in place without departing from the scope of this
invention.
[0028] A series of tubular metal electrodes 111, one for each well
75 in the microtiter plate 77, are secured (e.g., brazed) to the
lower face of the power plate 51 generally coaxially with the holes
95 in the plate for receiving the capillary tubes 7. Alternatively,
the tubes 7 could extend down outside the electrodes. In either
case, the electrodes 111 extend down from the plate 51 for
reception in the wells 75, one electrode for each well, for
electrifying the contents of the wells when the power source 15 is
activated and when the support block 73 is in its raised position
closing the pressure chamber 83. The power plate 51 and electrodes
111 are preferably of copper or other suitable metal and are
preferably gold plated to render them chemically inert or
non-reactive.
[0029] The support block 73 is movable up and down relative to the
power plate by an actuator generally designated 115. The actuator
may be a linear actuator such as a pneumatic cylinder 117 secured
to the base 59 and having its rod end attached to a pusher plate
119 affixed to the support block 73. However, it will be understood
that other types of power actuators or manually operated devices
may also be used. The support block 73 is guided as it moves up and
down by a pair of vertical guide pins 121 which extend down from
the power plate 51 through edge grooves 123 (FIG. 6) in the
periphery of the sealing block 67 for reception in clearance holes
125 in the support block (FIG. 8), each clearance hole having a
diameter only slightly larger than that of the respective guide
pin. The guide pins 121 are suitably affixed to the power plate 51,
as by a press fit.
[0030] As shown in FIGS. 6 and 8, the sealing block 67 has a
generally horizontal bore 131 therein forming a gas inlet for the
introduction of pressure gas into the pressure chamber 83 when the
support block 73 is raised to form the aforementioned enclosure 81.
The bore 131 has a fitting 133 at its upstream end for connection
to a gas supply system, generally designated 135 in FIG. 1, for
supplying gas under pressure to the pressure chamber 83. An annular
channel 139 extending down from the upper face of the sealing block
67 surrounds the pressure chamber 83 and communicates with the
inlet bore 131. The annular channel 139 communicates with the
pressure chamber 83 via a plurality of passages formed, for
example, by notches 145 in the upper face of the sealing block 67
spaced at intervals around the pressure chamber. Five such notches
145 are shown in FIG. 6, two relatively small notches closely
adjacent the inlet 131 on opposite sides thereof at one side of the
pressure chamber 83, a larger notch 145 on each of the two sides of
the pressure chamber adjacent the side with the inlet, and a single
very large notch 145 on the side of the pressure chamber opposite
the inlet. This arrangement insures a uniform distribution of
pressure air throughout the channel 139 for uniform and
substantially instantaneous pressurization of the pressure chamber
83, as will be described in more detail later. The number of
passages 145 and their configuration may vary.
[0031] The gas supply system 135 comprises a source of pressurized
gas (e.g., cylinder 151), an accumulator 153 having an inlet 155
and an outlet 157, an accumulator inlet line 161 connecting the gas
source 151 and the accumulator 153, and an accumulator outlet line
165 connecting the accumulator outlet and the inlet passage 131 in
the sealing block 67. A conventional regulator 169 in the
accumulator inlet line 161 reduces the pressure of the gas supplied
from the cylinder 151 to an acceptable level (e.g., from about 2000
psi to about 40 psi). A pressure controller, generally designated
171, is also provided in the inlet line 161 for controlling the
pressure in the accumulator. The pressure controller may be a
closed-loop electronic control system including a proportioning
control valve 175, PID (proportional, integrated and differential)
control electronics 177 and a pressure transducer 179. The
regulator 169 should reduce the pressure in line 161 to a level at
or below the maximum input pressure of the pressure controller. The
control system 171 may purchased as an off-the-shelf integrated
package, such as a 640 Series pressure controller commercially
available from MKS Instruments, Inc. of Andover, Mass. Other
pressure control systems may also be used, as will be understood by
those skilled in this field. A shut-off valve 181 (e.g., a
pneumatically driven shut-off valve) is provided in the accumulator
outlet line 165. A valve suitable for this purpose is a diaphragm
valve DA Series, Model 316L VAR, available from Nupro Company of
Willoughby, Ohio.
[0032] The accumulator 153 is a hollow vessel of metal, for
example, having a pressure chamber 185 therein, the volume of which
is related to the volume of the pressure chamber 83 defined by the
power plate 51, sealing block 67 and support block 73 when the
latter is in its raised position. The ratio of the volume of this
latter chamber 83 to the accumulator chamber 185 is in the range of
about 5:1--about 1:5, more preferably in the range of about
2:1--about 1:2, and most preferably about 1:1. The accumulator 153
may be of two-part construction, comprising upper and lower parts
capable of being releasably secured together in sealing relation.
The precise construction is not important to the present invention.
A vent line 187 is connected to the accumulator 153 for venting the
interior of the accumulator. The vent line 187 includes a fine leak
valve 191 and a shut-off valve 193. The shut-off valve 193 may be a
valve identical to the shut-off valve 181 in the accumulator outlet
line 165. A suitable fine leak valve 191 is a BM Series bellows
sealed metering valve available from Nupro Company of Willoughby,
Ohio.
[0033] As noted previously, the CE system shown in FIG. 1 includes
two injectors 3L, 3R for receiving opposite ends of the capillary
tubes 7 of the bundle 5. The left ends of the tubes of the bundle
extend down into the pressure chamber 83 defined by the power plate
51, sealing block 67 and support block 73 of the left injector 3L,
and the right ends of the tubes extend down into the pressure
chamber 83 defined by the power plate 51, sealing block 67 and
support block 73 of the right injector 3R. However, as will be
described later, the present invention can be practiced using only
one injector at either end of the bundle of capillary tubes.
[0034] The bundle 5 of capillary tubes, injectors 3L, 3R, light
source 19, cooling body 29, imaging lens 25 and photodetector 21
are preferably enclosed in a thermally insulated enclosure 201
having one or more doors 203. One or more convective coolers 207
are provided in the enclosure 201 for cooling it and maintaining
the interior of the enclosure at a desired temperature.
[0035] In use, the CE system 1 of the present invention may be set
up as shown in FIG. 1, where the array 11 of the intermediate
portions of the parallel capillary tubes 7 drape over the two
supports 35 and rest flat on the cooling body 29 in a position in
which the detection window 27 of the tubes is in registry with the
window 31 in the cooling body so that light from the light source
19 will pass through the detection window and the window in the
cooling body for incidence on the imaging lens 25 and photodetector
21.
[0036] After the capillary tubes 7 are flushed and loaded with a
suitable buffer solution ("conditioned"), samples to be analyzed
are loaded into the tubes in accordance with the present invention.
Assuming the samples are to be loaded into the right ends of the
capillary tubes as shown in FIG. 1, a microtiter plate 77 (or other
multi-receptacle container) carrying the samples to be analyzed is
placed in the recess 91 in the top surface of the support block 73
of the right injector 3R when the support block 73 is in its
lowered position. The cylinder 117 is then actuated to move the
block 73 to its raised position in which the support block is
sealed against the sealing block 67 to form the stated pressure
chamber 83 containing the samples. In this position, the right ends
of the capillary tubes extend down into the wells 75 of the
microtiter plate 77 and contact the samples. The electrodes 111 on
the power plate 51 also extend down into the wells and contact the
samples. (The microtiter plate is positioned by the recess 91 to
insure proper alignment between the electrodes 111 and wells 75.) A
second microtiter plate 77 (or other container) is placed in the
recess 91 in the upper surface of the support block 73 of the left
injector 3L, below the left ends of the capillary tubes, for
receiving waste from the tubes. The support block 73 of the second
injector 3L can either be maintained in its lowered position or
moved to its raised position during the loading process.
[0037] Loading of samples into the right ends of the capillary
tubes is effected by introducing gas under pressure into the
pressure chamber 83 of the right injector 3R. This process is
initiated by closing the shut-off valve 181 in the accumulator
outlet line 165 and the valves 191, 193 in the accumulator vent
line 187, and by opening the proportioning valve 175 to permit
entry of pressurized gas into the accumulator chamber 185 until the
pressure in the accumulator reaches a predetermined pressure P1, as
indicated by the pressure transducer 179. The shut-off valve 181 in
the accumulator outlet line is then opened, which permits gas to
enter the pressure chamber 83 via inlet passage 131, channel 139
and notches 145. The pressure in the chamber 83 should rapidly
reach equilibrium at the desired injection pressure P2, which is
preferably about 1-100 millibars, more preferably about 10-40
millibars and most preferably about 20 millibars, preferably in
less than about one second. It will be noted in this regard that if
the gaseous volume of the pressure chamber 83 is V1, the initial
pressure in the chamber 83 is P0 (e.g., ambient), and the gaseous
volume of the accumulator chamber 185 is V2, then the final
injection pressure P2, under ideal gas laws, can be determined to
be: P2=P0(V1/(V1+V2))+P1(V2/(V1+V2)). Assuming the desired
injection pressure P2 is 20 millibars, for example, and the ratio
of V1 to V2 is 1:1, then the accumulator pressure P1 should be 40
millibars before the shut-off valve is opened; if the ratio of V1
to V2 is 5:1, then P1 should be 120 millibars; if the ratio is 1:5,
then P1 should be 24 millibars. (It will be noted in this regard
that volumes V1 and V2 are "gaseous" volumes, meaning the volume of
space occupied by gas. Therefore, when the chamber 83 holds a
microtiter plate 77, V1 is the volume of the chamber 83 when
unoccupied less the volume of space occupied by the plate 77 and
samples therein. The volume V2 of the accumulator chamber 185 is
the volume of the unoccupied chamber. The volume of the accumulator
chamber may vary, with one suitable volume being three cubic
inches.)
[0038] Upon pressurization of the pressure chamber 83, the samples
in the wells 75 of the microtiter plate 77 are forced substantially
concurrently into the right ends of the capillary tubes 7. This is
allowed to continue for a predetermined amount of time (e.g., ten
seconds) sufficient to inject a plug of sample into each tube,
following which the proportioning valve 175 is closed and the vent
valves 191, 193 are opened to vent the accumulator 153 and the
pressure chamber 83 to atmosphere.
[0039] Following sample loading, and prior to the start of an
electrophoresis operation, the cooling system is actuated to cool
the interior of the enclosure 201 and the capillary tubes therein.
This involves actuating the one or more convective cooling units
207 and also the conduction cooling device 29 for a time sufficient
to bring the interior air temperature of the enclosure down to a
temperature sufficient to prevent overheating of the capillary
tubes and the contents thereof, which is particularly important
during a chiral separation process involving the generation of
substantial heat. A temperature in the range of about 0-99.degree.
C., preferably in the range of about 0-40.degree. C., and most
preferably about 20.degree. C., is believed to be suitable for this
purpose.
[0040] After the enclosure 201 and capillary tubes 7 are suitably
cooled, the power source 15 is activated to apply a voltage to the
tubes 7, causing the various components of the samples to migrate
at different speeds to effect separation, as will be understood by
those skilled in this field. To separate chiral molecules, a
relatively large current is required (e.g., a sum total of 1-20
milliamps for a bundle of 96 capillary tubes), which results in the
generation of a substantial amount of heat in the tubes and
contents thereof. The conduction heat transfer device 29 removes
this heat in the area of the bundle generally adjacent the
detection window 27, where the capillary tubes 7 are relatively
closely spaced. The convective heat transfer units 207 remove this
heat from other portions of the bundle 5. As a result, overheating
of the capillary tubes 7 and contents thereof is prevented, thus
ensuring a more accurate analysis of the samples.
[0041] Light from the light source 19 passes through the planar
array 11 of the capillary tubes 7 and is projected by the lens 25
as an image of the tubes onto the photodiodes of the photodetector
21. These diodes generate signals which are processed in
conventional fashion to generate and display an electropherogram 39
plotting light intensity (indicative of absorption levels) versus
time.
[0042] As the electrophoresis operation proceeds, sample solution
flows through the capillary tubes 7 and into one or more
receptacles 75 on the support block 73 of the left injector 3L. A
96-well microtiter plate 77 may be used if there is a need or
desire to maintain the collected solution in each tube separate
from the solutions in the other tubes, as when different buffer
solutions are used in different tubes during the same test run.
[0043] It will be apparent from the foregoing that samples can be
loaded into either end of the bundle 5 and that sample flow through
the tubes can be in either direction. Also, the position of the
detection window 27 can be varied relative to the injectors so that
D1 in FIG. 1 is not equal to D2. In other words, the detection
window 27 can be positioned at a location other than midway between
the two injectors 3L, 3R. As a result, two different capillary
"separation lengths" are achieved using the same bundle.
("Separation length" is the distance between the sample loading end
of the bundle and the detection window 27.) The provision of two
different separation lengths may be useful, since different types
of samples may require different separation lengths, and since a
shorter separation length may be sufficient for sample analysis not
requiring precise results.
[0044] As noted previously, the capillary tubes 7 are "conditioned"
(i.e., flushed and filled with buffer solution) prior to each run.
Conditioning is readily accomplished using the setup shown in FIG.
1. A container containing a suitable cleaning solution is simply
placed on the support block 73 of the right injector 3R, for
example, and the block is then moved to its raised position in
which the pressure chamber 83 is closed and the ends of the
capillary tubes are received in the container. The accumulator is
then pressurized, leaving the shut-off valve 181 in the accumulator
outlet line 165 open, so that cleaning solution is forced through
the capillary tubes and collected in a suitable waste receptacle on
the support block of the other (e.g., left) injector 3L. The
procedure is repeated to fill the tubes with buffer solution.
[0045] A CE operation can be carried out using only one injector at
one end of the bundle, instead of two injectors. If only one
injector is used at one end of the bundle, the opposite end of the
bundle should be placed in an electrically grounded receptacle
containing buffer solution prior to the beginning of a run.
[0046] It will be understood that the construction of the injector
3L, 3R may vary without departing from the scope of this invention.
For example, while the pressure chamber 83 is described as being
formed by the power plate 51, sealing block 67 and support block
73, the chamber may be formed by other parts of other
configurations, any one of which parts may be movable to provide
access to the interior of the chamber for placement of one or more
sample receptacles therein. Similarly, the accumulator 153 of the
gas supply system 135 can take many forms. It is important,
however, that the accumulator 153 be sized relative to the pressure
chamber 83 so that pressure equilibrium is reached very quickly
after the shut-off valve 181 in the accumulator outlet line 165 is
opened.
[0047] For additional detail regarding the cooling system 29, 207,
reference may be made to copending U.S. patent application Ser. No.
______, filed Jul. 20, 2000, incorporated herein by reference.
[0048] When introducing elements of the present invention or the
preferred embodiment(s) thereof, the articles "a", "an", "the" and
"said" are intended to mean that there are one or more of the
elements. The terms "comprising", "including" and "having" are
intended to be inclusive and mean that there may be additional
elements other than the listed elements.
[0049] In view of the above, it will be seen that the several
objects of the invention are achieved and other advantageous
results attained.
[0050] As various changes could be made in the above constructions
and methods without departing from the scope of the invention, it
is intended that all matter contained in the above description or
shown in the accompanying drawings shall be interpreted as
illustrative and not in a limiting sense.
* * * * *