U.S. patent application number 15/438754 was filed with the patent office on 2017-08-10 for disposable multi-channel bio-analysis cartridge and capillary electrophoresis system for conducting bio-analysis using same.
The applicant listed for this patent is BIOPTIC, INC.. Invention is credited to Varouj D. AMIRKHANIAN, Shou-Kuan TSAI.
Application Number | 20170227493 15/438754 |
Document ID | / |
Family ID | 59497578 |
Filed Date | 2017-08-10 |
United States Patent
Application |
20170227493 |
Kind Code |
A1 |
TSAI; Shou-Kuan ; et
al. |
August 10, 2017 |
DISPOSABLE MULTI-CHANNEL BIO-ANALYSIS CARTRIDGE AND CAPILLARY
ELECTROPHORESIS SYSTEM FOR CONDUCTING BIO-ANALYSIS USING SAME
Abstract
A multi-channel bio-separation system configured to utilize a
cartridge that has a individual, separate integrated reagent (i.e.,
a separation buffer) reservoir dedicated for each separation
channel. The multiple channels may have different characteristics,
such as different separation medium of different chemistries,
different separation length, different channel sizes and internal
coatings. In one embodiment, the cartridge does not include
integrated detection optics. Not all channels need to be operative.
One or more of the channels in the cartridge may be "dummy
channels" that are not operative (e.g., not provided with a
capillary tube). A capillary tube may be routed between the
reservoir/electrode (anode) of one channel to an electrode
(cathode) in another channel, thus allowing a longer length of
capillary tube to be used to define a longer separation channel to
improve resolution.
Inventors: |
TSAI; Shou-Kuan; (New Taipei
City, TW) ; AMIRKHANIAN; Varouj D.; (La Crescenta,
CA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
BIOPTIC, INC. |
New Taipei City |
|
TW |
|
|
Family ID: |
59497578 |
Appl. No.: |
15/438754 |
Filed: |
February 21, 2017 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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14720723 |
May 22, 2015 |
|
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15438754 |
|
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62297073 |
Feb 18, 2016 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
G01N 27/44726 20130101;
B01L 2400/0421 20130101; B01L 2200/0631 20130101; B01L 3/5025
20130101; G01N 27/44721 20130101; G01N 27/44791 20130101; B01L
2300/0864 20130101; B01L 2300/12 20130101; B01L 2300/0627 20130101;
B01L 2300/0838 20130101 |
International
Class: |
G01N 27/447 20060101
G01N027/447; B01L 3/00 20060101 B01L003/00 |
Claims
1. A multi-channel cartridge for bio-separation, comprising: a
plurality of cartridge elements, wherein each cartridge element
comprises: a body defining an opening as a detection window for
receiving external detection optics; a capillary column supported
in the body; and a reservoir attached to a first end of the body in
fluid flow communication with a first end of the capillary column,
wherein the reservoir is dedicated to a single cartridge element;
wherein the cartridge elements are coupled, and wherein the
reservoirs of the respective cartridge elements are not in flow
communication.
2. The multi-channel cartridge as in claim 1, wherein the cartridge
elements are coupled in a series in a plane.
3. The multi-channel cartridge as in claim 1, wherein the detection
window of at least one cartridge element exposes a section along
the capillary column, to which the external optics are aligned
through the detection window.
4. The multi-channel cartridge as in claim 3, wherein the capillary
column is supported coaxially by two ferrules that are supported in
the body, wherein each of the ferrules is cantilevered by the body
and having an end extending into the detection window, and wherein
a detection zone along the capillary column is exposed between the
extended ends of the ferrules.
5. The multi-channel cartridge as in claim 1, the body is slender
and generally longitudinal and blade shaped.
6. The multi-channel cartridge as in claim 5, wherein the capillary
column is supported along a central axis of the body.
7. The multi-channel cartridge as in claim 1, wherein in each
cartridge element, the body has a body section from which a first
end of the body extends, wherein the body section has a width, and
wherein the first end of the body is narrower than the width of the
body section.
8. The multi-channel cartridge as in claim 7, wherein the first end
of the body tapers from the body section to a narrower end.
9. The multi-channel cartridge as in claim 1, wherein at least a
first cartridge element further comprises an anode, wherein the
anode is conductively coupled to fluid contained in the reservoir,
and wherein the anodes of the respective cartridge elements are not
conductive coupled in the cartridge.
10. The multi-channel cartridge as in claim 9, wherein at least a
second capillary column further comprises a cathode disposed at the
first end of the body, and wherein the capillary column extends
between the reservoir of the first cartridge element and the
cathode of the second cartridge element.
11. The multi-channel cartridge as in claim 9, wherein each
cartridge element comprises an anode and a cathode, wherein the
cathode is disposed at the first end of the body of the cartridge
element, and wherein the capillary column extends between the
reservoir and the cathode in each cartridge element, exposing a
section along the capillary column in the detection window, to
which the external optics are aligned through the detection
window.
12. The multi-channel cartridge as in claim 11, wherein each
cartridge element defines a different separation channel.
13. A multi-channel cartridge for bio-separation, comprising: a
body defining a plurality of openings as detection windows for
receiving external detection optics, wherein each detection window
corresponds to a separate one of a plurality of separation
channels; a plurality of reservoirs coupled to the body, each
corresponding to a separate one of the separation channels; and a
plurality of capillary columns, each having a first end in fluid
flow communication with a separate reservoir, and a second end
extending away from the reservoir to a distal end of the body.
14. A bio-separation system, comprising: a chassis, a multi-channel
cartridge as in claim 1, wherein the cartridge body is supported by
the chassis; a table support at least a tray containing a sample
and a buffer with respect to an extended end of at least one
capillary column; at least one fork assembly supporting detection
optics, wherein the fork assembly is movable to extend into the
detection window defined in the cartridge element; a separation
mechanism effecting bio-separation within the capillary column; and
a controller controlling movement of the fork assembly and the
separation mechanism to effect separation.
15. The bio-separation system as in claim 14, further comprising a
first fork assembly and a second fork assembly, wherein the
detection optics comprises first optics supported by the first fork
assembly directing incident radiation to a detection zone and
second optics supported by the second fork assembly collecting
radiation from the detection zone.
16. The bio-separation system as in claim 15, wherein the first and
second fork assemblies are positioned on opposite lateral sides of
the cartridge, wherein the first and second fork assemblies move
between a first position in which the first and second fork
assemblies do not extend into the detection window defined in the
cartridge, and a second position in which the first and second fork
assemblies extend into the detection window defined in the
cartridge.
17. The bio-separation system as in claim 14, further comprising a
temperature control mechanism to control the temperature of the
sample in the tray.
18. A bio-separation system, comprising: a chassis, a multi-channel
cartridge as in claim 12, wherein the cartridge body is supported
by the chassis; a table support at least a tray containing a sample
and a buffer with respect to an extended the capillary columns; a
plurality of fork assemblies supporting detection optics, wherein
the fork assemblies are movable to extend into the detection
windows defined in the cartridge elements, and wherein each
separation channel is provided with at least one fork assembly; a
separation mechanism effecting bio-separation within the capillary
columns in each separation channel; and a controller controlling
movement of the fork assemblies and the separation mechanism to
effect separation.
Description
PRIORITY CLAIM
[0001] This application claims the priority of U.S. Provisional
Patent Application No. 62/297,073 filed on Feb. 18, 2016, and this
application is a continuation-in-part of U.S. Utility patent
application Ser. No. 14/720,723 filed on May 22, 2015. These
applications and all documents discussed below are fully
incorporated by reference as if fully set forth herein.
BACKGROUND OF THE INVENTION
[0002] 1. Field of the Invention
[0003] The present invention relates to bio-analysis, in particular
a cartridge and a capillary electrophoresis instrument using same
for performing bio-analysis.
[0004] 2. Description of Related Art
[0005] Currently, most of bio-separation tools applied in the
laboratories utilizes slab gel based electrophoresis technologies,
which have routinely been used for bio-analysis of bio-molecules
(i.e. DNA, Protein & Carbohydrate) applications since their
inception more than 20 years ago. However, slab gel electrophoresis
for bio-analysis is labor intensive and needs to be drastically
improved in terms of resolving power, throughput and cost per
sample.
[0006] Capillary electrophoresis (CE) is a micro fluidic approach
to gel-electrophoresis (micro-channel device to simplify
gel-electrophoresis), whose greatest advantage is its diverse range
of applications. CE technology is commonly accepted by the
biotechnology industry specifically in the nucleic acid-based
testing as a reliable, high resolution and highly sensitive
detection tool, and CE has been applied for, e.g., protein,
carbohydrate and DNA-related analyses such as oligonucleotides
analysis, DNA sequencing, and dsDNA fragments analysis, and glycan
profiling. CE is commonly avoided in routine analysis because it is
reputed to be a troublesome technique with high failure rates.
However, this is no longer true because instrument manufacturers
have drastically improved instrument design and overall CE
knowledge has increased. There are three key factors for reducing
failure rate and producing accurate, precise and robust CE data:
operator training, system stability, and operation ease of the
instrument with low maintenance.
[0007] Without a doubt, CE with laser-induced fluorescence (LIF) is
one of the most powerful analytical tools for rapid, high
sensitivity and high-resolution bio-analysis. However, the current
selling price for CE-based LIF systems is much more expensive than
traditional slab-gel based bio-analysis systems due to the
complicated optical detection mechanism. The expensive CE-based
systems are thus out of reach for all but a few well-funded
laboratories and seems to be a high-cost barrier for the expansion
of bio-analysis applications/business.
[0008] U.S. Pat. No. 8,784,626, commonly assigned to the assignee
of the present invention, discloses a simplified, low cost,
efficient, highly sensitive, non-moving and stable micro-optical
detection configuration for bio-separation (e.g., capillary
electrophoresis) through a separation channel (e.g., defined by a
column) filled with a separation support medium (e.g., a liquid or
sieving gel including a running buffer). More particularly, the
disclosed invention is directed to an improved detection
configuration that includes optics for application of incident
radiation at and detection of output radiation from a detection
zone along the separation channel, for the detection of radiation
emitted by sample analytes (e.g., radiation induced fluorescence
emission). In one aspect of the disclosed invention, the direction
of incident radiation (e.g., from a laser or LED source), the axis
of the separation channel at the detection zone, and the direction
of collection of the output radiation are all substantially in the
same plane. In one embodiment, the incident radiation is provided
to the detection zone and/or the output radiation is collected from
the detection zone, using light guides in the form of optical
fibers. In an embodiment, the detection configuration of the
present invention has optical fibers positioned at opposite sides
of the detection zone along the separation channel. The optical
fibers may be positioned at less than 180 degrees (e.g., 40 to 160
degrees, such as 120 degrees) apart from each other for high
detection sensitivity. In another aspect of the disclosed
invention, the detection configuration of the present invention
incorporates ball-end optical fibers to provide incident radiation
and collection of output radiation. In a further aspect of the
disclosed invention, the detection optics configuration of the
present invention may be implemented in an improved bio-separation
instrument, in particular a capillary electrophoresis
instrument.
[0009] U.S. Pat. No. 8,778,155, commonly assigned to the assignee
of the present invention, discloses a cartridge-based
bio-separation system configured to utilize a pen shaped
bio-separation cartridge that is easy to assemble and use with no
moving parts and that has an integrated reagent (separation buffer)
reservoir. The cartridge includes a body, defining an opening as a
detection window for receiving external detection optics, at least
one capillary column supported in the body, having a first end
extending beyond a first end of the body, wherein the detection
window exposes a section along the capillary column, to which the
external optics are aligned through the detection window, and a
reservoir attached to a second end of the body in fluid flow
communication with a second end of the capillary column. The
reservoir is structured to be coupled to an air pressure pump or N2
tank that pressurizes the gel reservoir to purge and fill the
capillaries with buffer/gel as the separation support medium.
[0010] U.S. patent application Ser. No. 14/720,723, published as
US20150338347A1, commonly assigned to the assignee of the present
invention, discloses a method for glycan profiling by capillary
electrophoresis (CE), and a CE system for glycan analysis
(N-Glycan). The CE system uses integrated dual optical fibers for
both radiation excitation and emission detection. The CE system is
configured for performing a two-color detection for data analysis.
A single radiation excitation source is used to excite two emission
fluorophores or dyes in the sample solution to be analyzed. One
emission dye is to tag the sample and the other dye is used to
provide a reference marker (e.g., a Dextran Ladder) for the sample
run. Two detectors (e.g., photomultiplier tubes (PMTs)) are applied
to simultaneously detect the fluorescent emissions from the dyes.
The data collected by both detectors are correlated (e.g.,
synchronized, and/or super-positioned for analysis) for accurate
data peak identification.
[0011] Based on the above disclosed detection technology, there is
a need for a multi-channel capillary electrophoresis system that is
simple and less expensive to operate (i.e. low cost per sample
run), providing rapid analysis with high efficiency, sensitivity
and throughput.
SUMMARY OF THE INVENTION
[0012] The present invention provides a simplified, low cost, high
efficiency, highly sensitive, high throughput, multi-channel
bio-separation system (e.g., capillary electrophoresis system). The
bio-separation system includes an instrument that includes optics
for application of incident radiation at and detection of output
radiation from a detection zone along each separation channel, for
the detection of radiation emitted by sample analytes (e.g.,
radiation induced fluorescence emission), without requiring fine
alignment of the optics to each separation column (e.g., a
capillary tube or column). The instrument is configured to conduct
bio-separation in the separation channels of the bio-separation
cartridge in an automated manner.
[0013] In one aspect of the present invention, is directed to
multi-channel cartridge-based bio-separation system configured to
utilize a reliable, compact, simplified, removable, portable,
interchangeable, reusable, low cost, recyclable and/or disposable
bio-separation cartridge that is easy to assemble and use with no
moving parts and that has an individual, separate, integrated
reagent (i.e., a separation buffer) reservoir dedicated for each
separation channel. The bio-separation cartridge includes multiple
separation channels defined therein. The multiple channels may have
different characteristics, such as different separation medium of
different chemistries, different separation length, different
channel sizes and internal coatings.
[0014] In one embodiment, the overall size of the cartridge is
characterized by the separation channel having an effective
separation length (i.e., the length of the capillary along which
bio-separation is expected to take place between the cathode and
the detection zone, which would be shorter than the actual length
of the capillary column) of no longer than 50 cm, preferably in the
range of 11 to 15 cm. In one embodiment, the bio-separation system
includes an instrument that is provided with a detection
configuration that includes optics for application of incident
radiation at and detection of output radiations at multiple
wavelengths/colors from a detection zone along each separation
channel, for the detection of radiations emitted by sample analytes
(e.g., radiation induced fluorescence emission) without requiring
fine alignment of optics to the capillary column. The instrument is
configured to conduct bio-separation in each separation channel of
the bio-separation cartridge in an automated manner. Each
separation channel may be controlled to effect bio-separation under
different parameters (e.g., different applied voltages, different
incident wavelengths, different run times, etc.).
[0015] In one embodiment, a capillary column that is supported by
and within the cartridge defines each separation channel. In one
embodiment of the present invention, the bio-separation system is
for capillary electrophoresis separation and analysis, and the
instrument therein is structured to utilize the capillary cartridge
to conduct capillary electrophoresis separation, detection and
analysis in an automated manner. In one embodiment, the capillary
column is defined by a capillary tube for each channel, having a
particular length, size (internal diameter), and internal
coating.
[0016] In another aspect of the present invention, the chemistry of
the medium and the characteristics of the separation column (e.g.,
for separation channels defined by capillary tubes, the capillary
size I.D., internal coating and length) are defined for each
channel in the cartridge. Different cartridges can be easily
interchanged for use in the bio-separation system to suit the
particular sample based separations. The buffer reservoir of each
channel is structured to be coupled to an air pressure pump or N2
gas, that pressurizes the reservoir to purge and fill the
associated separation channel with buffer (e.g., a gel) as the
separation support medium. The cartridge does not require detection
optics to be integrated into the cartridge, and the separation
channel does not require fine alignment with respect to the
detection zones. In one embodiment, the cartridge does not include
integrated detection optics.
[0017] In one embodiment, not all channels need to be operative.
One or more of the channels in the cartridge may be "dummy
channels" that are not operative (e.g., not provided with a
capillary tube).
[0018] In one embodiment, a capillary tube may be routed between
the reservoir/electrode (anode) of one channel to an electrode
(cathode) in another channel, thus allowing a longer length of
capillary tube to be used to define a longer separation channel to
improve resolution.
[0019] In one embodiment, the cartridge is provided with a RFID to
identify the configuration of the cartridge, such as the number of
operative channels, the chemistry in each of the channels, and the
characteristics of the separation column (e.g., capillary tube) in
each channel. The cartridge may be used for multiple runs without
the need to replace the cartridge or refurbish the cartridge with
fresh separation medium for reuse. The RFID may be configured to
track the number of runs completed, to determine the end-of-life of
the cartridge. The spent cartridge may be disposed or refurbished
by replacing the separation medium and other parts (e.g., capillary
tubes and seals).
BRIEF DESCRIPTION OF THE DRAWINGS
[0020] For a fuller understanding of the nature and advantages of
the invention, as well as the preferred mode of use, reference
should be made to the following detailed description read in
conjunction with the accompanying drawings. In the following
drawings, like reference numerals designate like or similar parts
throughout the drawings.
[0021] FIG. 1A is a schematic view of a capillary electrophoresis
system that incorporates a multi-color detection configuration and
cartridge in accordance with one embodiment of the present
invention; FIG. 1B illustrates the region A in FIG. 1A, including
the detection region, and schematically showing the configuration
of the excitation fiber, emission fiber and the capillary
column.
[0022] FIG. 2A illustrates the external view of a capillary
electrophoresis (CE) instrument, FIGS. 2B and 2C are sectional
views showing internal components, FIGS. 2D and 2E are internal
views, in accordance with one embodiment of the present
invention.
[0023] FIGS. 3A to 3H illustrate various views of a multi-channel
cartridge, in accordance with one embodiment of the present
invention.
[0024] FIG. 4A is an exploded view of the cartridge; FIGS. 4B and
4C are sectional views of the cartridge.
[0025] FIGS. 5A to 5C are various views of the front cover of the
cartridge, in accordance with one embodiment of the present
invention.
[0026] FIGS. 6A and 6B are various views of the rear cover of the
cartridge, in accordance with one embodiment of the present
invention.
[0027] FIGS. 7A to 7C are various views of the reservoir of the
cartridge, in accordance with one embodiment of the present
invention.
[0028] FIGS. 8A and 8B illustrate internal routing of capillary
tubes within the cartridge, in accordance with one embodiment of
the present invention.
[0029] FIG. 9A shows the insertion of a cartridge into a receiver
block in the instrument, and 9B shows additional heating/cooling
module for the receiver block, in accordance with one embodiment of
the present invention; FIG. 9C shows a receiver block, and FIG. 9D
is a sectional view showing the fork assembly in the interior of
the receiver block, in accordance with one embodiment of the
present invention.
[0030] FIGS. 10A to 10D are various views of the fork assembly, in
accordance with one embodiment of the present invention.
[0031] FIG. 11 schematically illustrates a two-color detection
scheme in accordance with one embodiment of the present
invention.
[0032] FIG. 12 illustrates determination of resolution of bases
from separation.
[0033] FIGS. 13-15 illustrate resolution of fluorescence detection
of by the inventive CE instrument and method in accordance with the
present invention.
[0034] FIGS. 16-19 illustrate results of fluorescence detection by
the inventive CE instrument and method in accordance with the
present invention.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
[0035] This invention is described below in reference to various
embodiments with reference to the figures. While this invention is
described in terms of the best mode for achieving this invention's
objectives, it will be appreciated by those skilled in the art that
variations may be accomplished in view of these teachings without
deviating from the spirit or scope of the invention.
[0036] The present invention provides a high-throughput capillary
gel-electrophoresis (4-channel) system for highly efficient, high
speed, high throughput, biomolecules analysis. The 4-Channel CGE
Analyzer (Qsep400) is a newly developed product that will
significantly increase the pace at which DNA research is performed
in the lab, saving hours of preparation time and assuring accurate,
consistent and economical results. In one aspect of the present
invention, a multi-channel high-performance capillary gel
electrophoresis analyzer system has been optimized for
DNA/RNA/Glycoprotein applications. The system uses integrated dual
fiber optic fluorescence detection technology (Excitation and
Emission detection) and a novel 4-Channel disposable gel-cartridge.
The system can hold a total of 96 samples, which can be
automatically analyzed within 1-2 hours. This high-throughput CGE
system with Multi-Color fiber optic based fluorescence detection
can be used in laboratories for high speed genotyping
applications.
[0037] Reference is made to the bioanalytical system including
detection system disclosed in U.S. Pat. Nos. 8,778,155 and
8,784,626, and U.S. Patent Application Publication No.
US20150338347A1, the entirety of which are incorporated by
reference as if fully set forth herein. These patents and patent
applications are commonly assigned to BiOptic, Inc., the applicant
and assignee of the present invention. In particular, these patents
disclosed a simplified, low cost, high efficiency, highly
sensitive, high throughput bio-separation system (e.g., capillary
electrophoresis (CE) system). The bio-separation system includes an
instrument that is configured to work with a single channel
capillary cartridge, and that is provided with a detection
configuration that includes optics for application of incident
radiation at and detection of output radiation from a detection
zone along the separation channel, for the detection of radiation
emitted by sample analytes (e.g., radiation induced fluorescence
emission), without requiring fine alignment of the optics to the
separation column. The instrument is configured to conduct
bio-separation in the separation channel of the bio-separation
cartridge in an automated manner. The CE system has a less complex
optical detection mechanism to reduce costs, which complements
simplicity in operation, rapid analysis with high efficiency,
sensitivity and throughput. US20150338347A1 further discloses
fluorescence detection at two colors. The present invention adopts
and modifies these systems to include improvements to the system
(namely, a multi-channel cartridge having individual, separate
reservoirs) in accordance with the disclosure hereinbelow.
[0038] For purpose of illustrating the principles of the present
invention and not limitation, the present invention is described by
reference to embodiments directed to capillary electrophoresis
using a capillary separation column. Further, the present invention
will be described, without limitation, in connection with radiation
induced fluorescence detection (e.g., using a laser or LED source).
Fluorescence is a spectrophotometric method of analysis where the
molecules of the analytes are excited by irradiation at a certain
wavelength and emit radiation at a different wavelength. The
emission spectrum provides information for both qualitative and
quantitative analysis. Generally, the advantage of fluorescence
detection over absorbance detection is the superior detectability
(detection sensitivity). For efficient fluorophores, single
molecule detection in small volumes has been demonstrated. This is
in part because fluorescence signal is measured against a
relatively dark background, as a result of the emitted radiation
being detected at a wavelength that is different from the
wavelength of the incident radiation (e.g., the wavelength of the
emitted fluorescence is at longer wavelengths than the excitation
radiation).
[0039] System Overview
[0040] Miniaturization and automation of analytical instrumentation
has many advantages over conventional labor intensive techniques
(i.e. manual Slab-gel Electrophoresis). These advantages include
improved data precision and reproducibility, short analysis times,
minimal sample consumption, improved automation and integration of
complex workflows.
[0041] Referring to FIG. 1A, a capillary electrophoresis (CE)
system 100 incorporates the detection configuration as
schematically illustrated. The CE system 100 generally comprises a
capillary separation column 10 (e.g., 200-500 .mu.m O.D.), which
defines internal separation channels 12 (e.g., 25-200 .mu.m I.D.),
which may be capillary columns 10 (only one separation
channel/capillary column is illustrated for simplicity). The
capillary column 10 may be made of fused silica, glass, polyimide,
or other ceramic/glassy materials. The inside walls of the
separation column 10 (i.e., the walls defining the separation
channel 12) may be coated with a material that can build up an
electrostatic charge to facilitate electrophoresis and/or
electrokinetic migration of the sample components. The separation
channel 12 may be filled with a separation support medium, which
may be simply a running buffer, or a sieving gel matrix (of a
linear or non-linear polymeric composition) known in the art.
[0042] One end of the capillary column 10 is coupled to a reservoir
14 of running buffer. The other end of the capillary column 10 is
coupled to another reservoir 16, which may alternately contain a
sample (to be injected into the separation channel 12) and running
buffer (after sample injection, to undertake separation). A power
supply 18 supplies a high voltage to the reservoirs 14 and 16 via
electrodes 20 and 22.
[0043] The mechanism of electrophoresis and radiation induced
fluorescence when considered alone are outside the scope of the
present invention. For the sake of completeness, it is sufficient
to briefly mention the operation of the CE system 100. In
operation, a prepared biological sample, tagged with at least one
known fluorophore, is introduced into the far end of the capillary
column away from the detection zone, by any of a number of ways
that is not part of the present invention (e.g., electrokinetic
injection from a sample reservoir or physical pressure injection
using a syringe pump). When a DC potential (e.g., 1-30 KV) is
applied by the power supply 18 to the electrodes 20 and 22, the
sample migrates under the applied electric potential along the
separation channel 12 in the direction 24 (e.g., sample that is
negatively charged travels toward the positive electrode 22 as
shown in FIG. 1) and separates into bands of sample components. The
extent of separation and distance moved along the separation
channel 12 depends on a number of factors, such as migration
mobility of the sample components, the mass and size or length of
the sample components, and the separation support medium. The
driving forces in the separation channel 12 for the separation of
samples could be electrophoretic, pressure, or electro-osmotic flow
(EOF) means.
[0044] When the sample reaches the detection zone 32, excitation
radiation is directed via the excitation fiber 34 in a direction 35
at the detection zone 32. The sample components would fluoresce
with intensities proportional to the concentrations of the
respective sample components (proportional to the amount of
fluorescent tag material). The detector 42 detects the intensities
of the emitted fluorescence via the emission fiber 36 in a
direction 37, at one or more wavelengths different from that of the
incident radiation. The detected emitted radiation may be analyzed
by a multi-color (e.g., two-color) detection scheme (further
discussed in reference to FIG. 11 below). For an automated system,
a controller 26 (e.g., in the form of an external notebook computer
or a desktop computer, or a computing unit integrated on-board the
instrument) having a processor, controls the operations of the
various components in the CE system 100 to effect capillary
electrophoresis separation and data collection, and controls of
other functions discussed herein below. The specific implementation
of such control is well within the knowledge of one skilled in the
art given the disclosure herein.
[0045] Detection Configuration
[0046] In the particular illustrated embodiment in FIG. 1A, the
detection optics configuration (schematically indicated in the area
30 located about a detection window/zone 32) corresponds to the
embodiment illustrated in FIG. 1B (region A in FIG. 1A). The
direction 35 of incident radiation (e.g., from a laser or LED
source), the axis of the separation channel at the detection zone,
and the direction 37 of collection of the output radiation are all
substantially in the same plane. In the illustrated embodiment, the
detection configuration of the present invention has optical fibers
positioned at opposite sides of the detection zone separation
channel. In one embodiment, the incident radiation is provided to
the detection zone and/or the output radiation is collected from
the detection zone, using light guides in the form of optical
fibers, in particular ball-ended optical fibers (i.e., optical
fibers terminating in a micro ball that is integral to the fiber
end in a unitary structure).
[0047] Referring also to FIG. 1B, a ball-ended fiber (the
excitation fiber 34) extends from a radiation source (e.g., LED or
laser source 41, schematically shown in FIG. 1A) to direct
excitation radiation in a direction 35 at the detection zone 32.
The ball end of the excitation fiber 34 is positioned at or
proximate to the exterior surface of the separation column 10 about
the detection zone 32. In the illustrated embodiment, the ball end
of the excitation fiber 34 is positioned at a distance spaced from
the exterior surface of the separation column 10 (i.e., non-contact
mode). In this illustrated embodiment, another ball-ended fiber
(the emission fiber 36) extends to a detector (e.g., a fluorescence
detector 42, schematically shown in FIG. 1) to collect emitted
radiation at a direction 37 from the detection zone 32. The ball
end of the emission fiber 36 is positioned at or approximate to the
exterior surface of the separation column 10 about the detection
zone 32. In the illustrated embodiment, the ball end of the
emission fiber 36 is positioned at a distance spaced (in a
non-contact mode) from the exterior surface of the separation
column 10. Both excitation and emission fibers 34 and 36 with ball
tips are positioned at opposite sides of the separation column 10
in a non-contact mode (spaced from the exterior of the capillary
column) to reduce background fluorescence and not cause any
physical damage to either capillary column or the micro-ball.
[0048] In the illustrated embodiment in FIG. 1B, the components at
the detection zone 32 as shown lie in substantially the same plane.
Specifically, the longitudinal axis of the excitation fiber 34, the
longitudinal axis of the emission fiber 36 and the longitudinal
axis of the capillary channel 12, are substantially aligned in the
same plane (i.e., substantially coplanar), at least at the region
of the detection zone 32. That is, while the lengths of the
excitation fiber 34, the emission fiber 36 and the capillary column
10 may be bent overall, however at least near the detection zone
region, the axis of the excitation fiber 34, the axis of the
emission fiber 36 and the axis of the capillary channel 12 are
substantially aligned in the same plane, such that the direction 35
of incident radiation from the excitation fiber 34 towards the
detection zone 32, the axis of the separation channel 12 at the
detection zone 32, and the direction 37 of collection of the output
radiation away from the detection zone along the emission fiber 36
are all substantially in the same plane.
[0049] Further, at the detection zone 32, the angle between the
axis of the excitation fiber 34 and the axis of the emission fiber
36 are not aligned in a straight line. At least one of the axis of
the excitation fiber 34 and the axis of the emission fiber 36 is
not perpendicular to the axis of the separation channel 12 at the
detection zone 32. In the illustrated embodiment shown in FIG. 2,
both the axis of the excitation fiber 34 and the axis of the
emission fiber 36 are not perpendicular to the axis of the
separation channel, and are at angles 39 and 40, respectively, to
the axis of the separation channel 12 at the detection zone 32. The
angle 39 and the angle 40 may be substantially the same or
different, and may be less than or greater than 90 degrees measured
with respect to a reference direction of the axis of the separation
channel 12 or a reference section of the capillary column 10 (e.g.,
the section of capillary column 10 between the fibers 34 and 36 as
shown in FIG. 1B). For example, the angle 39 may be less than 90
degrees and the angle 40 may be greater than 90 degrees, measured
from the same reference section. In the illustrated embodiment in
FIG. 1B, the angles 39 and 40 are same and substantially in the
same plane.
[0050] In the embodiment illustrated in FIG. 1B, both the
excitation fiber 34 and the emission fiber 36 each has a 200 micron
diameter core as light guide within an external cladding, and a 350
micron diameter ball shaped tip (i.e., the ratio of the fiber core
diameter to the ball diameter is 1:1.75), which comprises fused the
core and cladding material. The ball shaped tip has a substantially
spherical profile. The ball-end fibers may be formed by using a
fusion splicer, or are available from a number of available
suppliers. The capillary column 10 has an outside diameter of 200
to 370 micron (e.g., 360 micron) and an internal diameter of 20 to
150 micron (e.g., 75 micron). The tip of the ball end of the
excitation fiber 34 is spaced at approximately 50-500 micron from
the external surface of the capillary column, and the tip of the
ball end of the emission fiber 36 is spaced at approximately 10 to
500 microns (e.g., 50-200 micron) from the external surface of the
capillary column. Alternatively, the emission fiber 36 may have a
300 micron diameter core with a 500 micron diameter ball shaped tip
at its distal end (i.e., the ratio of the fiber core diameter to
the ball diameter is 1:2.5). The angles 39 and 40 each may range
from greater than 0 to less than 90 degrees, preferably between 20
to 70 degrees, and more preferably at 30 to 45 degrees. In the
illustrated embodiment of FIG. 1B, both angles 39 and 40 are about
70 degrees. The ball ends of the fibers 34 and 36 are not touching
the capillary column 10.
[0051] In one embodiment, the optical detection system is
structured with a super-bright UV LED (e.g., LG Innotek/IRTronix or
Dowa) as excitation radiation source for the fluorescent labeled
(FITC) antibody fragment detection. The modular design and fiber
optic coupling provides flexibility for exchanging the excitation
radiation to a laser module (for LIF applications) or other type of
inexpensive light sources.
[0052] It has been found that compared with flat-end fibers (bare
fiber, with no micro ball lens), the ball-ended fibers provide good
focusing of incident radiation (light concentration/power density)
for the excitation fiber 34 and high collection efficiency (high
Numerical Aperture; NA) for the emission fiber 36 as a high angle
fluorescence collector for increased fluorescence signal collection
capability and improved detection sensitivity. Using large core
(e.g., 100-1000 micron) and high NA (0.15-0.5) multi-mode fibers,
it allows high power light coupling from LED or laser into the
excitation fiber 34. By producing an integrated micro ball lens at
the distal output end of the excitation fiber 34, it allows good
coupling efficiency inside the separation channel 12 (e.g., 20-200
micron micro-fluidic channel) for high fluorescence detection
sensitivity.
[0053] A smaller diameter excitation fiber 34 having 200 micron
core diameter with a 330-350 micron diameter ball (see FIG. 1B)
directed at the capillary separation channel 12 results in a
smaller focal spot with higher power density, thereby optimizing
the fluorescence excitation signal. If an emission fiber 36 having
a 300 micron core diameter and a 500 micron diameter ball lens is
used for emission collection, the emission collection efficiency is
increased. The outside diameter of the capillary column is 360
micron, and the inside diameter is 75 micron.
[0054] The 2-fiber detection configuration with ball-end fibers has
been applied to a disposable single-channel, single capillary
cartridge concept with an integrated buffer reservoir (see, U.S.
Pat. Nos. 8,778,155 and 8,784,626, and U.S. Patent Application
Publication No. US20150338347A1). The present invention provides a
higher throughput instrument utilizing multiple gel-cartridges of
similar design to speed up the separation time (cycle) by a factor
of 4.times.-8.times. (1 hour for full 96-well sample plate
run).
[0055] The test samples are introduced to the separation capillary
column 10 by electro kinetic injection. The high voltage power
supply (e.g., EMCO, Sutter Creek, Calif.) is used to deliver, e.g.,
500V to 20 KV of electrical field to the capillary for the electro
kinetic injection and separations of bio-molecules in each channel.
For each channel, an excitation LED having broad band light energy
(e.g., FWHM=20-50 nm) and 20-100 degrees of viewing angle is
coupled to the large core excitation fiber (e.g., 100-1000 micron)
at the flat end (polished or cleaved end). A line filter (e.g.,
FWHM=2-50 nm Band Pass line filter) is placed in front of the LED
before coupling the light into the 200 micron diameter core with
350 diameter micron ball-ended excitation fiber to reduce
background noise. The micro-ball lens end of the fiber is produced
by fusion splicing (high voltage heat melting) with a well
controlled ball diameter to create a well defined exit NA and spot
size for coupling the excitation radiation energy into the inner
diameter (the separation channel) of the capillary column. For each
channel, the fluorescence emission signal produced by the separated
analytes are then collected at the detection zone of the capillary
channel using a similar ball-ended fiber (larger core fiber with
500 micron diameter ball) and is relayed to an external detector
module (e.g., fluorescence detector 42 schematically shown in FIG.
1), which may include one or more photomultiplier tubes (PMTs) or
SiPMTs or CCDs, and may also include beam-splitters, built-in
emission filters (e.g., Band Pass Filters) for multi-color
detection, in accordance with further disclosure below.
[0056] Further details of the detection optics and detection scheme
may be referenced to U.S. Pat. Nos. 8,778,155 and 8,784,626, and
U.S. Patent Application Publication No. US20150338347A1, the
entirety of which have been incorporated by reference as if fully
set forth herein.
[0057] The CE instrument may include a temperature control
mechanism, such as a Peltier heating/cooling module 207 interfaced
with (e.g., below) the table 221 (see, FIG. 2D), or an external
Peltier heating/cooling module 207' as shown in FIG. 9B.
Temperature control allows for both PCR Amplification and
Electrophoresis/Detection in one instrument,
[0058] CE Instrument
[0059] FIG. 2A illustrates the external view of a CE instrument 200
that is comprised in the CE system 100. The CE instrument 200
incorporates some of the components of the CE system 100 discussed
above within the instrument, in accordance with one embodiment of
the present invention (some of the components of the CE system may
be external, e.g., a power supply, a pressurized gas source, a
computing unit, etc.). The CE instrument 200 includes components
including the detection configuration schematically shown in FIG.
1A. FIGS. 2B and 2C are sectional views that illustrate the
internal views of the CE instrument 200, in accordance with one
embodiment of the present invention. The CE instrument 200 includes
various components, including a receiver block 205 for receiving a
cartridge 60 in accordance with the present invention. The receiver
block supports a cartridge interface mechanism 204, which comprises
multiple fork assemblies that support the detection optics
discussed above and that interface/engage the detection optics to
the capillary columns in the cartridge. FIGS. 2D and 2E are
schematic views further illustrating the components of the CE
instrument 200 which reside within the instrument housing. The CE
instrument 200 comprises a system board 201, operatively coupled to
a sample transport mechanism 202 (e.g., a three-axis X-Y-Z
transport mechanism), a cartridge interface mechanism 204, an
optical signal detector such as comprising photomultiplier (PMT)
modules 206, a high voltage power supply 208 (which may
alternatively reside outside of the CE instrument 200), detection
optics (e.g., as shown in FIG. 1B), and a pressurized gas source
(which may reside outside the CE instrument 200, but connected to a
port in the instrument housing).
[0060] A controller 26 is provided for user interface and
programming of experiment/test settings and parameters. The
controller includes the necessary application software routines,
which may also include data reduction applications. The controller
26 may be an integral part of the instrument 200 (e.g., as part of
the system board 201, with application routines coded in ASICs), or
it may be a separate unit coupled/interfaced to the CE instrument
200. In the illustrated embodiment, the controller is external to
the housing of the CE instrument 200, in the form of a desktop
computer or notebook computer, which is coupled to the CE
instrument 200 via the system board 201 via a USB interface. The
external controller 26 may include mass storage devices, display,
keyboard, etc., or some of these user interface components may be
configured integral to the CE instrument (e.g., a display and a
keyboard on the front housing). Alternatively, the system board 201
may be incorporated as part of the external controller 26, without
departing from the scope and spirit of the present invention.
[0061] The system board 201 includes the necessary electronics to
drive the various components in the CE instrument, e.g., the
movements of the transport mechanism 202, the output of the power
supply 208, the PMT modules 206, the valve release of the
pressurized gas, the movements of the cartridge interface 204, an
RFID transmitter/reader, etc. It is noted that the system board 201
is schematically represented in the figures. It may include other
electronic boards for controlling specific components (e.g.,
electronic board for controlling motors in the sample transport
mechanism 202), or these other boards may be separate from and in
communication with the system board 201 to perform the intended
function. The exact electronic board configuration is not critical
to the present invention, and it is well within the knowledge of
one skill in the art to configure the boards to achieve the desired
functions and features disclosed herein. The controller 26 and/or
the system board 201 may be built into a front panel 203 of the
instrument housing to allow user access to place and remove a
sample and/or reagent tray 220. The front panel 203 includes a
touch screen user interface panel 25. The touch screen panel 25 can
thus be used as a control panel for setting operation of the
instrument 200. As illustrated, the front panel 203 can be driven
by stepper motors to slide up to provide access to place/remove a
tray 220 (e.g., supporting buffer solutions, reagents and/or
samples) on/from the transport mechanism 202, and to slide down to
prevent access.
[0062] As illustrated in FIG. 2D, the sample transport mechanism
202 includes a table 221 supporting the tray 220 having multiple
wells (e.g., a standard 96-well titer plate), and a tray 220a
having larger wells for a buffer, cleaning solutions and waste
collection, to move with three degrees of freedom. The multiple
wells may include wells containing cleaning solutions and samples
and also for waste collection. It is noted that in the figures, X,
Y and Z are orthogonal axes. Y is the vertical axis; X is in a
horizontal direction across the instrument (parallel to the rear of
the instrument); and Z is in a horizontal direction into and out of
the instrument. The table 221 is controlled by the transport
mechanism 202 to move up and down, and to move within a plane in a
straight line and rotate within the plane. That is, the table 221
moves in a single horizontal direction (Z-direction), and in a
vertical (Y-direction), and rotation about the vertical axis
(Y-axis). The combination of rotation and translation motions would
be able to place any of the multiple wells in the tray 220 and tray
220a for access by the tip of the depending capillary column
60.
[0063] The pressurized gas source (e.g., pressurized air or N2) may
be a gas cartridge installed within the housing of the CE
instrument, or may be an external source (e.g., air-pump) providing
pressurized gas to the CE instrument via a gas connection port at
the instrument housing (in which case, the pressurized gas source
would be the gas connection port to the external gas source). The
pressured gas is fed to the reservoir 62 in the cartridge 60 via
appropriate gas tubing and valves (which is operatively coupled to
the system board 201).
[0064] The power supply 208 includes a system DC power supply
(e.g., 12-24 VDC from external AC power) coupled to the system
board 201, and a variable high voltage power supply providing the
necessary high voltage to electrode contacts/probes for electrical
contact with electrodes 66 and 67 in the cartridge 60 for carrying
out electrophoresis therein. Alternatively, instead of using an
internal 12-24 VDC power supply with external AC power, the CE
instrument 200 may use an external 12-24 VDC power supply, which
makes the instrument simpler and safer to use without the internal
AC to DC conversion. This would also allow for battery operation
for field portability and operations. The contact probes may be
actuated pneumatically (e.g., by regulating pressurized gas from
the gas source, or electromechanically, to contact against the
exposed surfaces of the electrodes 66 and 67, or the contact probes
may be simply spring loaded to bias against the exposed surfaces of
the electrodes 66 and 67. Further details of the contact probes may
be referenced to U.S. Pat. Nos. 8,778,155 and 8,784,626, and U.S.
Patent Application Publication No. US20150338347A1, the entirety of
which have been incorporated by reference as if fully set forth
herein.
[0065] The excitation fiber 34 is optically coupled to a light
source in the form of a 4-channel LED modules 226, which may be
part of the system board 201. The emission fiber 36 is optically
coupled to the PMT modules 206 via appropriate optical filters. The
electrical output of the PMT modules 206 is coupled to the system
board 201.
[0066] Referring also to FIGS. 9A and 9B, the cartridge interface
mechanism 204 is supported in the receiver block 205 of the
instrument, and is configured to receive the cartridge 60, and
support its location positively and accurately with respect to the
detection optics. A cartridge-door 261 (FIGS. 2A to 2C) is provided
at the top side of the instrument housing to allow insertion and
removal of the cartridge 60.
[0067] As shown in FIGS. 9A to 9C, a hole puncher 119 is provided
to facilitate punching a seal that may have been provided on a tray
220 to prevent evaporation and/or contamination. The hole puncher
119 may be configured to be actuated pneumatically or
electromechanically for vertically up/down and/or lateral
movements.
[0068] Multi-Channel CE Cartridge
[0069] In one aspect of the present invention, the system 100 is a
multi-channel cartridge-based bio-separation system that comprises
a CE instrument 200 (e.g., shown in FIGS. 2A to 2E) that is
configured to utilize a reliable, compact, simplified, removable,
portable, interchangeable, reusable, low cost, recyclable and/or
disposable bio-separation cartridge that is easy to assemble and
use with no moving parts and that has a separate integrated reagent
(separation buffer) reservoir for each channel. Each of the
multi-channel bio-separation cartridge could be structured to have
an overall size and slender shape generally conforming to the
general shape of a pen. The bio-separation system 100 is provided
with the above-described detection configuration that includes
optics for application of incident radiation at and detection of
output radiation from a detection zone along each of the separation
channels, for the detection of radiation emitted by sample analytes
(e.g., radiation induced fluorescence emission) without requiring
fine alignment of optics to the capillary column. The system 100 is
configured to conduct bio-separation in each of the separation
channel of the multi-channel bio-separation cartridge in an
automated manner.
[0070] In the disclosed embodiments, the present invention provides
reusable 4-channel gel-cartridge, which permits easy plug-and-play
use in a robust injection molded body with integrated 4-independent
gel-reservoir design that incorporates 4-micro-fluidic glass
capillary (20-100 .mu.m ID) with an effective separation length of,
e.g., 11 cm. The shortened capillary length allows for reduced
operating voltages (1-15 KV) and the elimination of expensive
cooling systems such as Peltier or recirculating chillers. The
design includes top and bottom electrodes (Anode & Cathode), an
exposed detection zone and an imbedded RFID chip/label to provide
ID for the gel-cartridge type and track the number of runs per
cartridge. Each cartridge contains linear gel-matrix and is capable
of analyzing 200 samples in as few as 2 minutes per sample,
consuming as little as 1 pl from the 1 .mu.l-20 .mu.l sample
volume.
[0071] The present gel-cartridge is a simple yet very robust design
approach for large volume type manufacturing for an easy to operate
CGE instrument that provides significant background noise
reduction, which results in improved S/N for high detection
sensitivity of biomolecules at very low-cost per sample run. The
4-channel independent gel-reservoir design provides flexibility to
use different gel-matrix (buffer) combinations for different
separation resolutions. The individual channels could be assembled
with different glass capillary inner diameters and lengths to
improve the performance for different biomolecules (see, FIGS. 8A
and 8B). Longer capillary tube could be looped from one reservoir
to the next section of the cartridge for longer separation runs to
improve resolution.
[0072] FIGS. 3A to 3H illustrate a multi-channel cartridge 60 in
accordance with one embodiment of the present invention. As
illustrated, the cartridge 60 includes four separate cartridge
elements 60' each representing/corresponding to a single channel.
Each cartridge element is blade shaped. More or less channels may
be implemented without departing from the scope and spirit of the
present invention (e.g., 2, 3, 6, 8, etc. channels). A capillary
column 10 is supported by and within each cartridge element
60'/channel of the cartridge 60. In the illustrated embodiment,
each of the cartridge element 60' has a slender and generally
longitudinal body 80. While the illustrated body 80 of the
cartridge 60 is generally flat cylindrical or blade shaped, it may
have other sectional cylindrical profile, such as round, square,
rectangular, hexagonal, elliptical, or other regular and irregular
profiles. As illustrated, the body 80 has a body section that has a
generally uniform or constant width, with the bottom end of the
body being narrower than the uniform width of the body section. The
bottom end of the body 80 may be tapered to a narrower section,
e.g., terminating in a generally conical portion 97. The capillary
column 10 is held by or within the cartridge body 80 generally in
line with the longitudinal central axis of the cartridge body 80.
In one embodiment, the overall size of the cartridge is
characterized by the separation channel being no longer than 30 cm,
preferably in the range of 10-20 cm.
[0073] Referring also to FIGS. 4A to 4C, 5A to 5C, 6A to 6B and 7A
to 7C, an outlet buffer reservoir 62 is attached to the top end of
the body 80. The buffer reservoir 62 includes a cap 85 (e.g.,
screw-on or plug) that seals the top opening of the reservoir 62,
to retain separation support medium (e.g., a gel buffer) therein.
An additional plug cover 85' may be provided under the cap 85. The
bottom of the reservoir 62 has a rim 88 defining a groove 93, and a
center stub 90 having a through hole 91 for receiving the capillary
column 10. The cap 85 of the reservoir 62 has a port 64 (e.g., a
small drilled hole) that is for coupling to an external pressurized
gas (e.g., nitrogen) supply (e.g., a gas tank or pump that is part
of the CE instrument to be discussed below). (When the cartridge 60
is not used for a while, the port 64 can be sealed by applying a
short strip of tape.) The pressurized gas provides the required air
pressure to purge and fill the capillary separation channel 12 in
the capillary column 10 with the separation support medium (buffer)
contained in the reservoir 62. Depending on the viscosity of the
separation buffer, pressures of up to 60 PSI can be applied to fill
the capillary column 10 through the top buffer reservoir 62. The
reservoir 62 is provided with an electrode 66 (anode), which
provides electrical contact to the buffer. The electrode 66 has
contact surface exposed to external through opening 63.
[0074] Referring also to FIGS. 5A to 5C and 6A to 6B that
illustrate the internal structures, the body 80 includes two half
covers or shells 82 and 83 that comprises complementary structures
on facing interior surfaces (i.e., surfaces not exposed to external
when the shells 82 and 83 are assembled), which may be generally
mirror of each other. The half shells 82 and 83 define a through
opening or window 86 for access to the detection zone 32 by
external optics (as will be explained further in connection with
the CE instrument discussed below.) The inside of the half shells
82 and 83 are generally hollow. The half shells 82 and 83 are each
provided with a flange 92, which mates with the groove 93 on the
reservoir 62 when the parts are assembled, to securely attached the
reservoir 62 to the body 80 of the cartridge 60 in an open cavity
at the top end of the cartridge body 80. Grooves and recesses (on
tabs 101) are provided at appropriate locations along the inside of
the half shells 82 and 83 to thread the capillary column 10. At the
outside surface of the half shell 82 is an alignment slot 50 and
indexing recesses 51 to provide guides to facilitate positive and
accurate positioning and alignment of the detection window 86 in
the cartridge 60 with respect to the CE instrument when the
cartridge 60 is inserted into the CE instrument. Similarly, at the
outside of the half shell 83, indexing recesses 52 are provided for
alignment and positioning the detection window 86 within the CE
instrument. Further, an alignment/indexing recess 53 are provided
to facilitate alignment and positioning of the electrode 67 to the
external power source provided in the CE instrument.
[0075] Referring also to FIG. 3H that is a sectional view of the
two half shells attached together, cylindrical sleeves or ferrules
87 upstream and downstream of the detection zone 32 support the
capillary column 10 within the body 80 of the cartridge 60 (see
also FIG. 4). The capillary column 10 is threaded through the
ferrules 87, and one end of the capillary column 10 extends into
the reservoir 62 in fluid communication with the buffer contained
in the reservoir 62, and the other end extends to depend beyond the
lower end of the cartridge body 80. In the illustrated embodiment
(see FIGS. 4B and 4C), to secure the upper end of the capillary
column 10 in the reservoir 62, a threaded nipple 96 having a
through-hole is threaded into the base of the reservoir 62 and
compress against an O-ring seal 96' (see also FIG. 6C). The upper
end of the capillary column 10 is inserted through the nipple 96.
The threaded nipple 96 provides flexibility in removing the
capillary column or accommodating capillary columns of different
lengths. Alternatively, instead of using the nipple, the end of the
capillary column 10 can be secured to the reservoir 62 by glue or
epoxy. The ferrules 87 extend from the recesses 94 into the window
86, but expose the detection zone 32 of the capillary column 10.
The two half shells 82 and 83 are assembled together to form the
body 80, e.g., by screws 89, or epoxy, or clips.
[0076] The capillary column 10 is supported coaxially by the
ferrules 87, which are supported in the cartridge body 80, wherein
each of the ferrules 87 is cantilevered by the cartridge body and
having an end extending into the detection window 86, and wherein
the detection zone along the capillary column is exposed between
the extended ends of the ferrules 87. At the lower end of the
cartridge 60 is another electrode 67 (cathode). The electrode 67
has contact surface exposed to external through opening 65 at the
conical portion 97 of the cartridge body 60 half shells 82 and 83,
for coupling to an external high voltage power supply in the CE
instrument for electrophoresis when installed inside a CE
instrument, such as the embodiment described herein below (see,
FIGS. 2A to 2E), which is designed to receive the cartridge 60. The
lower electrode 67 is configured in the form of metal/conductive
sleeve, extending from the lower end of the cartridge 60, and
surrounding the side of the depending end of the capillary column
10 completely (e.g., in the form of a co-axial metal tube) or
partially (e.g., in the form of a wire mesh, gauze or net, or an
open channel having a C-shaped cross-section), with the tip of the
capillary column 10 exposed for fluid communication with an
external buffer reservoir. The tip of the capillary column 10 may
extend beyond the end of the electrode 67, for better access to
samples.
[0077] To assembly the various components shown, the bottom rim 88
of each reservoir 62 is placed at the end of the half shells 83,
with the flange 92 inserted in the groove 93 on the reservoir 62.
For each cartridge element 60', a capillary column 10 is threaded
through the ferrules 87, and one end is threaded into the bottom
electrode 67. The other end of the capillary column 10 is inserted
into the bottom opening 91 on the reservoir 62, through the nipple
96. The nipple 96 is tightened onto the base of the reservoir 62,
compressing the O-ring 96' to provide a seal against the body of
the capillary column 10. The far ends of the ferrules 87 are
inserted in recesses 94 on the half shell 83. The lower electrode
67 is positioned in the groove 95 provided on the inside the
conical portion 97 of the half shell 83, with the end extending
beyond the conical portion 97. A drop of glue may be provided to
secure the electrode 67 in the groove 95. The other half shell 82
is placed over the half shell 83 and attached by suitable
fasteners, such as rivets or screws 89 as shown. The reservoir 62
is filled with the desired separation support medium (buffer) and
capped. The fully assembled cartridge 60 may be tested and labeled.
An electronic label, such as an RFID label 150 may be imbedded or
attached to the cartridge 60 (e.g., at the outside surface of the
reservoir 62), to provide a means of identification of the
particular configuration of the cartridge (e.g., buffer medium,
capillary size, coating and length). The RFID label may also
include the pre-set limit on the number of runs and type of
cartridge with expiration date. After assembling the cartridge 60,
the RFID label is provided with the initial configuration
parameters. The RFID may be re-recordable and updated with
information to track usage of the cartridge (e.g., the number of
runs and the conditions and/or parameters of the runs (e.g.,
applied voltage, duration, sample), the number of time the
cartridge has been reconditioned, etc.), so that the history of the
cartridge can be easy determined (e.g., by the CE instrument
discussed below or by a separate reader). The end of the useful
life of each cartridge can also be determined from the RFID label.
Alternatively, a static label, such as a bar code label may be
provided.
[0078] As will be explained in greater detail below, in
electrophoresis operation as installed in the CE instrument 200,
the end of lower electrode 67 along with the open end of the
capillary column 10 are dipped into an external buffer reservoir.
To conduct electrophoresis, high voltage is supplied to the
electrode 66 in the buffer reservoir 62 and the electrode 67 dipped
in the external reservoir, to provide a high voltage circuit across
the buffer to complete the electrophoresis path in the capillary
column 10. The electrode 67 also provides protection to prevent
breakage of the depending end of the capillary column 10.
[0079] The cartridge does not require detection optics to be
integrated into the cartridge, and the separation channel does not
require fine alignment with respect to the detection zones.
Specifically, in the illustrated embodiment, the cartridge does not
include integrated detection optics. In the detection window 86
surrounding the region of the detection zone 32, sleeves or
ferrules 87 upstream and downstream of the detection zone 32
support the capillary column 10 in the body of the cartridge 60.
External excitation fiber 34 and emission fiber 36 supported in the
CE instrument are aligned with the detection zone 32 through the
detection window 86 defined in the separation channel/column 10. In
the illustrated embodiment that will be further discuss below, the
excitation fiber 34 and emission fiber 36 are supported by the fork
assembly in the CE instrument (see FIG. 10, for example). The axes
of the fibers 34 and 36 and the capillary column 10 are coplanar.
The ball ends of the fibers 34 and 36 are in proximity to but not
touching the capillary column 10. In other words, the optical fiber
has a terminating integral ball-end structure that is spaced apart
from exterior of the separation channel, wherein the ball-end
structures do not touch exterior of the separation channel. This
conforms to the detection optic configuration shown in FIG. 1B.
[0080] As illustrated, the cartridge 60 has separate channels
defined by the separate cartridge elements 60'. The flow within
each cartridge element 60'/channel is separate from another
cartridge element/channel, as each channel is provided with its
separate reservoir. This allows the chemistry of the buffer medium,
and the characteristics of the capillaries (e.g., capillary size,
coating and effective separation length), to be separately defined
for each cartridge element 60', which may be different from one
another. Further, different cartridges 60 can be easily
interchanged for use in the CE instrument discussed below to suit
the particular sample based separation. The cartridges may be
replaced, reconditioned for reuse (e.g., with fresh buffer, seals,
new capillary column and/or electrodes, etc.), recycled, or
disposed.
[0081] The cartridge in accordance with the present invention can
be manufactured with relatively low cost. The body of the cartridge
can be made of injection molded plastic (e.g., PVC, polyurethane,
polycarbonate, acetal, etc. The electrodes can be made of stainless
steel. The ferrules could be made of injected molded plastic
material or aluminum or glass machined parts.
[0082] In the illustrated embodiment, the overall size of the
cartridge 60 is less than 20 cm in length (e.g., about 11 to 15
cm), and less than 3 cm in thickness (e.g., 2 to 3 cm). The length
of capillary column 10 that can be accommodated in the cartridge 60
is less than 50 cm (e.g., about 11 to 15 cm), with an effective
separation length of 11.5 cm. The capacity of the reservoir 62 is
less than 20 cc (e.g., about 10 to 20 cc).
[0083] Referring to FIG. 8A, the capillary column 10 may be routed
from the reservoir 62 (having the anode 66) to the corresponding
cathode 67 within the same cartridge element 60'. In another
embodiment, a capillary column may be routed between the
reservoir/electrode (anode) of one channel to an electrode
(cathode) in a another channel, thus allowing a longer length of
capillary tube to be used to define a longer separation channel. As
illustrated in FIG. 8A, the capillary column 10' is routed between
the reservoir 62 (anode 66) of cartridge element 60a' to the
cathode 67 of the adjacent cartridge element 60b'. This provides a
longer effective separation length, which can significantly improve
resolution of separated bases (as will be discussed in connection
with FIG. 12 below). FIG. 8B shows that a length of capillary
column of 38.5 cm having an effective separation length of 34.5 cm
may be looped within the cartridge 60, via two or more cartridge
elements 60', without having to use a cartridge having a
longitudinal length of greater than the effective separation length
in order to accommodate the capillary column.
[0084] Cartridge Interface Mechanism
[0085] The cartridge interface mechanism 204 is supported in the
receiver block 205 having an opening sized and configured to
receive the cartridge 60 as shown (e.g., FIGS. 9A to 9D). The
receiver block 205 is supported on the chassis of the instrument,
and is configured to receive the cartridge 60, and index its
location positively and accurately with respect to the detection
optics 210. As shown in FIG. 3E, for example, the right most
cartridge element 60' is wider than the left most cartridge element
60'. In addition, the left side of the top portion of the left most
cartridge element 60' protrudes slightly as compared to the right
side of the top portion of the right most cartridge element 60'.
These features provide guides to facilitate positive and accurate
positioning and alignment of the detection window 86 in the
cartridge 60 with respect to the fork assemblies 230 discussed
below, when the cartridge 60 is properly inserted into the CE
instrument.
[0086] In this illustrated embodiment, the cartridge 60 is support
by the receiver block 205 in a vertical orientation, with the
longitudinal axis of the capillary column 10 substantially vertical
with respect to the horizontal plane of the tray 220. It is within
the scope of the present invention to have the cartridge supported
with its longitudinal axis horizontal with respect to
reagent/sample containers. A safety interlocking feature may be
provided to engage to prevent the cartridge 60 from being
accidentally removed from the receiver block 205 during
electrophoresis operations. The safety interlock feature could also
include the front door (sample-door) 260 for tray 220 and top door
(cartridge-door) 261 for insertion of the cartridge 60, to prevent
user accidentally opening these doors during electrophoresis
operations. The safety interlock (not shown) will only be released
upon execution of termination sequence for an electrophoresis run
(e.g., shutting down high voltage supply, and outward movement of
the fork assemblies 230 described below). The receiver block 205
also includes an RFID reader/transmitter (e.g., on the outside of
the receiver block 205) for communicating with an RFID label on the
capillary cartridge 60.
[0087] The cartridge interface mechanism 204 comprises a pair of
opposing fork assemblies 230 for each channel. For each pair of
fork assemblies 230, track 229 is provided to allow actuated
movement of opposing fork assemblies to engage the respective fork
assemblies to the ferrules 87 in the cartridge element 60'.
Referring also to FIGS. 10A to 10C, the fork assemblies 230 are
attached to slides 231, which are actuated to be slidable along the
tracks 229 towards and away from each other. In other words, in the
illustrated embodiment, the fork assemblies 230 are supported to
slide along a same axis. The excitation fiber 34 and emission fiber
36 are each supported on a separate fork assembly 230. The fork
assemblies 230 are configured to position the ball shaped ends of
the excitation fiber 34 and the emission fiber 36 in proximity to
the detection zone 68 of the capillary column 10 for detection of
separated sample analytes. Movements of the fork assemblies 230 may
be implemented by pneumatic or electromagnetic actuation. In the
illustrated embodiment, the fork assemblies 230 are moved by
pneumatic pistons 233, which may make use of the supply of
pressurized gas 212 regulated by appropriate valve(s) (not shown)
controlled by the system board 201.
[0088] The cartridge 60 is positioned with respect to the fork
assemblies 230 in a manner such that for each channel, the opposing
fork assemblies 230 are positioned on opposite lateral sides of the
cartridge 60, wherein the fork assemblies move between a first
position in which the first and second fork assemblies do not
extend into the detection window defined in the cartridge element
60', and a second position in which the first and second fork
assemblies extend into the detection window defined in the
cartridge element 60'. The fork assemblies 230 essentially move
between a first position at which the fork assemblies are apart to
allow the cartridge element 60' to be inserted between the fork
assemblies 230, to a second position at which the fork assemblies
are pressing against the ferrules 87 (interlocked) in the detection
window 86 in the cartridge 60.
[0089] In the illustrated embodiment, there are four pairs of fork
assemblies 230 arranged in parallel, each in the orientation shown
in FIG. 10A (i.e., with the plane of the fork assemblies 230 being
vertical. The track 229 of some of the pairs of fork assemblies
would in the space between adjacent cartridge elements 60'.
[0090] The extended portion of the fork assemblies 230 are provided
with a complementary surface that facilitates alignment of the
extended surface against the ferrule 87, e.g., a V-groove or
concave surface 236 to complement the cylindrical body of the
ferrules 87. FIG. 10D is a simplified view that illustrates the
left fork assembly 230 (the one having the excitation fiber 34)
pressed against the ferrules 87 (with a cartridge element 60'
schematically illustrated). At this position, the concave surfaces
236 extend into the detection window 86 in the cartridge element
60'. In this position, the optical fiber supported on the left fork
assembly 230 delivers radiation to the capillary column 10. With
the right fork assembly 230 also engaging the ferrules 87 in the
cartridge element 60', the detection optical fiber supported on the
right fork assembly 230 collects radiation from the detection zone
32. In the particular illustrated embodiment, radiation induced
fluorescence detection scheme is implemented, but other types of
optical detection schemes may be implemented instead without
departing from the scope and spirit of the present invention. Both
fork assemblies 230 may be controlled to move together to press
against the ferrules 87 at about the same time, or move separately
to press against the ferrules 87 in sequence. In the illustrated
embodiment, the ferrules 87 provide a stop against the extended
surfaces of the fork assemblies 230, so that the terminating
integral ball-ends of the optical fibers do not touch the exterior
surface of the capillary column, but are spaced apart from the
exterior surface of the capillary cartridge at a predetermined
distance, which can be repetitively maintained when the fork
assemblies are actuated between the first and second positions
described above.
[0091] While the illustrated embodiment in FIG. 1B shows the
optical fibers oriented in a V-configuration, the optical fibers
may be configured in a straight or in-line fashion (e.g., for
absorbance type detection scheme), or with one or both optical
fibers configured with axis perpendicular to the axis of the
capillary column. Further, only one fork assembly may be used, with
both radiation delivering fiber and radiation collection fiber on
the same fork assembly.
[0092] The system board 201 controls various functions of the CE
instrument 200, including positioning the sample and buffer tray
220 with respect to the cartridge 60 held in the receiver block
205, and above described functions of the cartridge interface
mechanism 205, and other functions, such as detecting end of a run
and release of safety lock to release the cartridge 60 from the
receiver block 204.
[0093] System Operation for Electrophoresis:
[0094] To conduct a desired electrophoresis run, a user presets the
appropriate parameters using the controller 26. A cartridge 60,
having the appropriate separation support medium (buffer) and a
capillary column 10 having the desired size and coating in each
cartridge element 60', is inserted into the receiver block 205. The
controller 26 in association with the system board 201 takes over
control of the CE instrument 200, to undertake the tasks described
below.
[0095] The cartridge is "locked" in the receiver block 205 upon
proper insertion, with the detection window 86 of each cartridge
element 60' appropriately positioned with respect to the fork
assemblies 230. For each cartridge element 60', pressurized gas is
readied from its source, when the top door 261 is closed to press
the O-ring of the air outlet against the top of the cap 85 of the
cartridge reservoir 62 to access the port 64 on the cartridge
reservoir 62. The electrical contact probes 224 and 225 are pressed
against the electrodes 66 and 67. The fork assemblies 230 are moved
to mate against the ferrules 87 in the detection window 86.
[0096] By a combination of X, Y and Z-directions, the sample
transport mechanism positions the appropriate wells in the sample
and buffer tray 220 with respect to the depending tip of the
capillary column 10 supported in each cartridge element 60'. If
necessary, the separation buffer that is present in the capillary
column 10 is initially purged by application of pressurized gas
into the cartridge reservoir 62 (the tray 220 may be moved to
position a specific well for collecting waste from the capillary
column), and/or fresh separation buffer from the reservoir is
caused to fill the separation channel.
[0097] One or more test sample that are placed in one or more wells
on the tray 220, and the tray 220 is positioned to submerge the
depending tip of the capillary column 10 and the end of electrode
67 of each cartridge element 60'. The sample is introduced into the
separation capillary column 10 by electro kinetic injection
(appropriate high voltage applied for a defined period of time,
e.g., less than 60 seconds, e.g., 5 to 10 seconds), a procedure
well known to one skill in the art.
[0098] A buffer reservoir in the tray 220 is then positioned to
submerge the tip of the capillary column 10 and the end of
electrode 67. Electrophoresis is carried out by application of high
voltage at an appropriate level for a defined period of time for
the particular sample and separation buffer medium. During the run,
data corresponding to radiation-induced fluorescence is collected
via the PMT 206. The data is stored in an electronic file. At the
end of the run, the tray 220 is lowered.
[0099] If no further runs, the cartridge 60 may be removed by
executing a preset release procedure, including releasing the
pressurized gas supply, moving the fork assemblies 230 away from
the cartridge elements 60' (as described above), disengaging the
electrodes 66 and 67 by the contact probes 224 and 225 (if they are
actuable), and releasing the lock on the cartridge 60. The
cartridge 60 can thus be removed, and replaced with another
cartridge for a next run at a desired time.
[0100] If further runs are desired for same or additional samples,
the old buffer (e.g., gel buffer) from the previous run is purged
into the waste well from the capillary column 60 by pressuring the
reservoir to refill the capillaries with fresh buffer. The tray 220
is positioned so that the tip of the capillary column 60 in each
cartridge element 60' is cleaned with cleaning solution (in a
well), before another sample is loaded into the capillary column 60
and electrophoresis run conducted as described earlier.
[0101] It is noted that because the sample analytes that flowed to
the buffer reservoir 62 at the exit of the capillary column are in
such small amount and volume concentration compared to the volume
of the reservoir, and that the analytes are expected to be mixed
within the gel reservoir, there will only be a negligible trace of
analytes from past runs in the reservoir, and that will be evenly
distributed in the gel that refills the capillary column for
subsequent runs. Any noise from this negligible trace would be
relatively small background noise that can be easily removed from
the detected signal in the data analysis.
[0102] If no further runs, the cartridge 60 may be removed by
executing a preset release procedure, including releasing the
pressurized gas supply, moving the fork assemblies 230 away from
the cartridge 60 (as described above), disengaging the electrodes
66 and 67 by the contact probes 225 and 225 (if they are actuable),
and releasing the lock on the cartridge 60. The cartridge 60 can
thus be removed, and replaced with another cartridge for a next run
at a desired time.
[0103] The above-mentioned sequence of process may be programmed as
one of the automated functions of the controller 26.
[0104] The collected data is analyzed by using appropriate
application software routines. Referring to FIG. 12, the resolution
of the peak separation interval (time and # bases) is determined
using a Full Width at Half Maximum (FWHM) approach.
Resolution (bases)=Peak Separation Interval (# bases)/(Peak
Separation Interval (time)/FWHM (time)).
[0105] Typically a 15 cm long capillary (with effective length of
11.5 cm) with an I.D. of 20-100 .mu.m (typically 70 .mu.m) is used
for each cartridge/channel for DNA Fragment analysis. The
resolution achieved for 155 bp (base pairs) is 1.95 using POP-7
gel. With the new design of Qsep400 one could use longer length of
capillary (see FIGS. 8A and 8B) within the 4-channel cartridge
module. For a total length of 38.5 cm (with effective length of
34.5 cm) filled with POP-7 Polymer separation matrix/denaturing-gel
(Applied Biosystems, Life Technology/ThermoFisher Scientific,
Carlsbad Calif.), one can achieve a resolution of 0.675 for the 150
bp and 160 bp (Peak Separation Interval of 29.64 seconds with
FWHM=2.0 seconds), when testing with the GX500-ROX DNA ladder
(end-labeled ssDNA from Applied Biosystems; Life
Technologies/ThermoFisher Scientific, Carlsbad Calif.). [Applying
the above formula for resolution, the resolution
achieved=(160-150)/(29.64 sec/2.0 sec)=0.675.]
[0106] From a mechanical packaging perspective, the longer
capillary could be looped between two electrodes of either the
single channel or could be routed/looped from the electrode of one
channel to the 2.sup.nd, 3.sup.rd, or 4.sup.th channels/electrode
(see, FIGS. 8A and 8B).
FIGS. 13 to 15 illustrate the comparison of resolution of 150-160
bp of GX500-ROX with longer capillary (FIG. 13), and resolution of
150-160 bp of GX500-ROX with longer capillary (Zoom) with
Resolution of 0.675 (FIGS. 14 and 15). FIGS. 14 and 15 is the
zoomed in section of FIG. 15 for the 150-160 bases and the FIG. 12
elaborates on how we achieved the 0.675 by using the formula
Example: Resolution=(160-150)/(29.64 sec/2.0 sec)=0.675 to achieve
it.
[0107] FIG. 11 illustrates one embodiment of two-color detection,
including a fiber combiner/splitter 61 for splitting an emission
signal 37 into two signals for fluorescence detection at two
different wavelengths. The 1.times.2 fiber combiner/splitter 61
couples the output signal 37 of the emission fiber 36 to the inputs
of a first emission fiber 36a and a second emission fiber 36b. The
emission fiber 36 includes fluorescence at at least two wavelengths
.lamda.1 and .lamda.2. In this embodiment, .lamda.1 corresponds to
the wavelength of the fluorescence of detected Dextran Ladder and
.lamda.2 corresponds to the wavelength of the fluorescence of
detected glycan profile. The first emission fiber 36a routes
emissions from the emission fiber 36 to a first PMT1 that detects
fluorescence at .lamda.1, and the second emission fiber 36b routes
emissions from the emission fiber 36 to a second PMT2 that detects
fluorescence at .lamda.2. The fiber combiner may be of the type
that splits orthogonal polarizations at at least two wavelengths
(e.g., at .lamda.1 and .lamda.2) or two ranges of wavelengths
(e.g., Thorlabs 1.times.2 Coupler or Gould 1.times.2 Fiber
Splitter). In addition, while not shown in FIG. 6, one or more
band-pass filters may be provided between the PMT1 and/or PMT2 and
the corresponding outputs of the first and/or second emission
fibers 36a and 36b. Another embodiment of two-color detection is to
utilize a dichroic filter/beam-splitter to split an emission signal
37 into two emission signals for fluorescence detection at two
different wavelengths. An example of an appropriate beam-splitter
may be a model no. DMLP P425 longpass type Dichroic Mirror
available from ThorLabs, Inc., which has a 45.degree. angle of
incident, a cutoff wavelength of 425 nm, with a transmission band
of 440-700 nm, and a reflectance band of 380-410 nm. In addition,
while not shown in FIG. 11, one or more band-pass filters may be
provided between the PMT1 and/or PMT2 and the corresponding outputs
of the first and/or second emission fibers 36a and 36b. Further
details of dual color detection approach can be found in U.S.
Patent Application Publication No. US20150338347A1, commonly
assigned to the assignee of the present invention, which had been
incorporated by reference herein.
[0108] In accordance with the present invention, fluorescence
detection is improved by being able to conduct 2-Color Detection
for accurate peak identification. A single excitation (LED or
Laser) is used to excite 2 different fluorophores for each
separation channel. One emission dye (fluorophore) is used for the
sample and the second dye (fluorophore) is used for the reference
ladder (DNA Ladder or Glycan Ladder: i.e Dextran) run with two
detectors (PMTs). The results from both detectors are synchronized
and displayed on top of each other (transposed electropherograms)
using the post data collection analysis software (post CE
separation analysis) for accurate peak identification.
[0109] FIGS. 16 to 19 illustrate the result of various runs using
the CE instrument disclosed above. Specifically, FIG. 16 shows the
result of 4-Channel runs of DNA Ladder (range of 20-5000 bp) in
less than 1-minute; FIG. 17 shows the result of 2-Color detection
(Detector #1+Detector #2) in the range of 20-1000 bp; FIG. 18 shows
the result of Glycan Ladder (Detector 1) transposed on top of the
glycan sample (Detector 2); FIG. 19 shows the result of Glycan
Ladder (Detector 1) and the glycan sample (Detector 2) displayed
individually and aligned by the arrows.
[0110] Using the CE system described above to perform glycan
profiling, it has been found that the present invention provides a
cost-effective capillary gel-electrophoresis system for highly
efficient, high speed, high throughput, glycan analysis (N-Glycan).
The novel method and system significantly increase the pace at
which glycoprotein research is performed in the laboratory, saving
hours of preparation time and assuring accurate, consistent and
economical results.
[0111] The two-color detection simplifies and shortens sample
separation and detection into a single run and assures accurate
data analysis for peak identification. The dual dye detection
(i.e., two dye labeling) is a very robust and accurate way to
provide reproducible peak identification and sizing for glycan
profiling.
[0112] The simplicity of the micro-optical detection also provides
flexibility in designing higher throughput (i.e., multi-channel)
type gel-cartridge without the use of optics (excitation or
emission optics) inside the cartridge assembly, hence reducing
costs for the cartridge.
[0113] Accordingly, the new fluorescence fiber-based detection for
the CE system in accordance with the present invention provides
simplicity in design, ease of operation and lower cost consumable
(e.g., for glycan profiling. It provides a good solution
particularly for the research and clinical diagnostic
laboratories/industry that demands sustained and stable recurring
revenue streams from both an installed base of instruments and
recurring need for consumables such as testing reagents and buffer
containing capillary cartridge.
[0114] While the invention has been particularly shown and
described with reference to the preferred embodiments, it will be
understood by those skilled in the art that various changes in form
and detail may be made without departing from the spirit, scope,
and teaching of the invention.
* * * * *