U.S. patent application number 15/580674 was filed with the patent office on 2018-07-05 for radiation carrier and use thereof in an optical sensor.
This patent application is currently assigned to IMEC VZW. The applicant listed for this patent is IMEC VZW. Invention is credited to Dries Vercruysse.
Application Number | 20180188152 15/580674 |
Document ID | / |
Family ID | 53491451 |
Filed Date | 2018-07-05 |
United States Patent
Application |
20180188152 |
Kind Code |
A1 |
Vercruysse; Dries |
July 5, 2018 |
Radiation Carrier and Use Thereof in an Optical Sensor
Abstract
A radiation carrier for carrying at least a radiation beam has,
on a surface thereof, at least one excitation grating, for
directing at least an excitation radiation beam directionally out
of the radiation carrier, thereby illuminating a region of
interest; and at least one structure for redirecting emission
radiation emanating from the region of interest. Further a sensor
is provided comprising at least one such radiation carrier and at
least one detector, the structure being adapted for redirecting
radiation from the region of interest into the at least one
detector.
Inventors: |
Vercruysse; Dries;
(Sint-Andries, BE) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
IMEC VZW |
Leuven |
|
BE |
|
|
Assignee: |
IMEC VZW
Leuven
BE
|
Family ID: |
53491451 |
Appl. No.: |
15/580674 |
Filed: |
June 30, 2016 |
PCT Filed: |
June 30, 2016 |
PCT NO: |
PCT/EP2016/065394 |
371 Date: |
December 7, 2017 |
Current U.S.
Class: |
1/1 |
Current CPC
Class: |
G01N 21/645 20130101;
G02B 6/0036 20130101; G01N 2021/6463 20130101; G02B 6/30 20130101;
G02B 6/124 20130101; G01N 21/6428 20130101; G01N 15/1436 20130101;
G01N 15/1434 20130101; G01N 2021/6482 20130101; G01N 15/1484
20130101; G01N 2021/0346 20130101; G01N 21/65 20130101; G01N
2015/1006 20130101 |
International
Class: |
G01N 15/14 20060101
G01N015/14; G01N 21/64 20060101 G01N021/64; G01N 21/65 20060101
G01N021/65; F21V 8/00 20060101 F21V008/00 |
Foreign Application Data
Date |
Code |
Application Number |
Jun 30, 2015 |
EP |
15174678.1 |
Claims
1-22. (canceled)
23. A radiation carrier for a sensor, comprising: a surface; at
least one excitation grating on the surface of the radiation
carrier, positioned and adapted to couple an excitation radiation
beam directionally out of the radiation carrier, thereby
illuminating a region of interest; and at least one structure
positioned and adapted for redirecting emission radiation emanating
from the region of interest, wherein the radiation carrier is
adapted for carrying at least a radiation beam.
24. The radiation carrier according to claim 23, wherein the at
least one structure positioned and adapted for redirecting emission
radiation is at least one emission grating adapted for reflecting
emission radiation to a detector.
25. The radiation carrier according to claim 23, wherein the at
least one structure positioned and adapted for redirecting emission
radiation is at least one emission grating adapted for coupling
emission radiation into a radiation carrier.
26. The radiation carrier according to claim 23, wherein the at
least one structure positioned and adapted for redirecting emission
radiation comprises planar optics.
27. A sensor comprising: at least one radiation carrier according
to claim 23; and at least one detector, wherein the at least one
structure is positioned and adapted for redirecting emission
radiation from the region of interest into the at least one
detector.
28. The sensor according to claim 27, wherein the at least one
structure positioned and adapted for redirecting emission radiation
is adapted to further collimate the redirected emission radiation
from the region of interest to the at least one detector.
29. The sensor according to claim 27, wherein the at least one
structure positioned and adapted for redirecting emission radiation
is adapted to further focus the redirected emission radiation from
the region of interest to the at least one detector.
30. A microfluidic device comprising a sensor according to claim
27, further comprising a substrate being transparent for at least
the radiation beam wherein the region of interest is defined.
31. A microfluidic device according to claim 30, wherein the
substrate is furthermore transparent for the redirected emission
radiation.
32. The microfluidic device according to claim 30, wherein the
substrate further comprises a microfluidic channel.
33. The microfluidic device according to claim 32, wherein the at
least one detector is a detector array, and wherein the
microfluidic channel is interlayered between the radiation carrier
and the detector array.
34. A system comprising, as separate devices, a microfluidic chip
comprising at least one microfluidic channel, and at least one
radiation carrier for carrying at least a radiation beam, wherein
the radiation carrier comprises a surface with at least one
excitation grating, positioned and adapted to couple an excitation
radiation signal directionally out of the radiation carrier thereby
illuminating a pre-defined volume of the microfluidic channel, and
at least one structure, positioned and adapted to redirect emission
radiation origination from the pre-defined volume; and a readout
device, adapted to be operatively coupled with the microfluidic
chip, wherein the readout device comprises at least one detector
for detecting the redirected emission radiation originating from
the pre-defined volume, when the microfluidic chip and the readout
device are operatively coupled.
35. A system according to claim 34, wherein the readout device
comprises a slot for receiving the microfluidic chip.
36. A diagnostic device comprising a sensor according to claim 27,
and an output unit for providing an output of the sensor on which a
diagnosis can be based.
37. A method of performing particle detection comprising providing
radiation scattering centers; inserting the radiation scattering
centers within a region of interest; providing radiation from an
excitation grating in optical contact with the region of interest;
redirecting radiation scattered from radiation scattering centers
in the region of interest, by means of at least one structure, to
at least one detector; and monitoring emission of radiation
redirected from the region of interest.
38. The method according to claim 37, wherein providing radiation
scattering centers comprises attaching the radiation scattering
centers to analytes.
39. The method according to claim 38, wherein attaching radiation
scattering centers comprises attaching at least one type of
fluorophores, or chromatophores, or a mixture thereof.
40. The method according to claim 37, wherein inserting the
radiation scattering centers within a region of interest further
comprises providing a flow of scattering centers through the region
of interest.
41. The method according to claim 37, wherein inserting scattering
centers within a region of interest comprises attaching analyte
carrying scattering centers to affinity probes.
42. The method according to claim 37, wherein the radiation is a
continuous radiation.
43. The method according to claim 37, wherein the radiation is a
pulsed radiation.
44. The method according to claim 37, further comprising detecting
wavelengths in different wavelength ranges in different
predetermined detectors of a plurality of detectors.
Description
FIELD OF THE INVENTION
[0001] The present invention relates to the field of particle
detection and optionally analysis. More specifically it relates to
particle detection and optionally analysis via optical means. In
particular, it relates to luminescence based detection, e.g.
fluorescence based detection, of particles or detection of
particles based on Raman scattering, in flow.
BACKGROUND OF THE INVENTION
[0002] Cytometry, in particular flow cytometry, consists in
identification of analytes (e.g. tumor cells) based on
morphological and/or chemical characteristics. In the latter case,
markers, e.g. luminescent markers such as for instance fluorescent
markers, are often used to label particular molecules, such as
proteins, that can identify the cell. This typically requires an
optical system that focuses laser light to excite the luminophores,
e.g. fluorophores, and that collects and filters the luminescence,
e.g. fluorescence, coming from the cell.
[0003] Luminescent, e.g. fluorescent, detection of particles is a
technique whereby a particle of interest in a fluid sample is
stained or labeled with one or more luminophores, e.g.
fluorophores. To detect the particle, the luminophores, e.g.
fluorophores, attached to the particle are activated by a light
signal and luminescence, e.g. fluorescence, from the luminophores,
e.g. fluorophores, is then detected by an optical detector. During
the phase of staining or labeling the particle, a number of
luminophores, e.g. fluorophores, bind to the particle, and a
remaining number of luminophores, e.g. fluorophores, do not bind.
In prior art devices, the remaining number of unbound luminophores,
e.g. fluorophores, have to be removed before activating the
luminophores, e.g. fluorophores, bound to the particle, because the
background noise reduces the sensibility of the detectors, which
typically have low resolution. These additional steps of removal of
unbound markers increase the complexity of the process and the area
needed for washing the markers off as well as controllable pumps
and other elements for flow control, typically making the devices
bulky.
[0004] Even simple cell counting tests require a lot of throughput.
To perform cell counting, cells need to flow through the system
very fast, which means that only little signals are available from
every single cell. Hence extremely sensitive systems that require
fine alignment of the laser and the optical systems are
required.
[0005] In order to manipulate the laser input, waveguides are often
used. However, the breadth of the frequency spectrum of
luminescent, e.g. fluorescent, light (broadband spectrum) hinders
or impedes an efficient input in a waveguide.
SUMMARY OF THE INVENTION
[0006] It is an object of embodiments of the present invention to
provide a compact and easy to use optical sensor and analyzer, for
performing luminescent, for instance fluorescent, or Raman
scattering detection.
[0007] In an aspect, the present invention provides a radiation
carrier for a sensor, the radiation carrier being adapted for
carrying at least a radiation beam, the radiation carrier
comprising a surface. The radiation carrier comprises at least one
excitation grating on the surface of the at least one radiation
carrier, positioned and adapted to couple an excitation radiation
beam directionally out of the radiation carrier, thereby
illuminating a region of interest (ROI), and further at least one
structure positioned and adapted for redirecting, for instance for
receiving and redirecting, such as for collecting and redirecting,
e.g. for reflecting, emission radiation emanating from the region
of interest. Emission radiation emanating from the region of
interest may be excitation radiation which is for instance simply
reflected on e.g. particles present in the region of interest, or
may be a type of radiation different from the excitation radiation,
which is generated in the region of interest, by interaction of the
excitation radiation with particles present in the region of
interest, such as for instance fluorescence or phosphorescence
radiation.
[0008] In embodiment of the present invention, the structure for
redirecting emission radiation may be a structure for reflecting
emission radiation. Alternatively, the structure for redirecting
emission radiation may be a structure for transmitting the emission
radiation. The structure for redirecting emission radiation may
include a structured or patterned surface.
[0009] In embodiments of the present invention, the at least one
structure for redirecting emission radiation may be at least one
emission grating adapted for reflecting emission radiation to a
detector. In alternative embodiments, it may be at least one
emission grating adapted for coupling emission radiation into a
radiation carrier. This radiation carrier may be the radiation
carrier for carrying the radiation beam, or it may be another,
second, radiation carrier. The second radiation carrier may be
positioned in the plane of the radiation carrier for carrying the
radiation beam, or angled, for instance substantially
perpendicular, thereto.
[0010] In yet alternative embodiments, the at least one structure
positioned and adapted for redirecting emission radiation comprises
planar optics, such as for instance a planar lens.
[0011] It is an advantage that a cheap, disposable radiation
carrier can be obtained with inexpensive materials. In some
embodiments, the radiation carrier comprises planar optics, for
producing a spread radiation beam and directing it towards the
region of interest. It is an advantage of embodiments of the
present invention that a compact device can be obtained.
[0012] In a further aspect, the present invention provides a sensor
comprising
[0013] at least one radiation carrier for carrying at least a
radiation beam, the radiation carrier comprising a surface,
[0014] at least one excitation grating on the surface of the at
least one radiation carrier, for directing at least an excitation
radiation beam into a region of interest (ROI),
[0015] at least one detector,
[0016] at least one structure, for instance but not limited
thereto, an emission grating or planar optics, for redirecting,
e.g. reflecting, radiation from the region of interest into the at
least one detector.
[0017] It is an advantage of embodiments of the present invention
that alignment of the optical system may be simplified or even
avoided in a compact device.
[0018] In a sensor according to embodiments of the present
invention, the at least one structure, e.g. emission grating of
planar optics, positioned and adapted for redirecting, e.g.
reflecting, radiation may be adapted to further collimate the
redirected, e.g. reflected, radiation from the region of interest
to the at least one detector. Collimation of the radiation allows
as much radiation as possible to hit the detector, such that a
usable amount of radiation for getting reliable results hits the
detector.
[0019] It is an advantage of embodiments of the present invention
that the whole area of a detector may be used, improving its
sensibility.
[0020] In a sensor according to embodiments of the present
invention, the at least one structure, e.g. emission grating or
planar optics, for redirecting, e.g. reflecting, radiation may be
adapted to further focus the redirected, e.g. reflected, radiation
from the region of interest to the at least one detector.
[0021] It is an advantage of embodiments of the present invention
that imaging and a good resolution can be obtained.
[0022] In a sensor according to embodiments of the present
invention, the at least one radiation carrier may comprise planar
optics for producing and directing a spread excitation radiation
beam towards a region of interest.
[0023] It is an advantage of embodiments of the present invention
that a ROI may comprise a wide length or volume of a microfluidic
channel, or a big area. A larger ROI can be created.
[0024] It is an advantage of integrating the focusing optics on the
chip that the optics can be very nicely aligned with the
microfluidics that are fabricated in the same process.
[0025] In a further aspect, the present invention provides a
microfluidic device comprising a sensor according to any of the
embodiments of the first aspect, and further comprises a substrate
being transparent for at least the radiation beam, wherein the
region of interest is defined. A microfluidic device according to
embodiments of the present invention may furthermore be transparent
for the redirected emission radiation.
[0026] It is an advantage of embodiments of the present invention
that an integrated optical sensor suitable for fluorescence
analysis can be obtained.
[0027] In a microfluidic device according to embodiments of the
present invention, the substrate may further comprise a
microfluidic channel.
[0028] It is an advantage of embodiments of the present invention
that an inexpensive miniaturized flow cytometer requiring little
maintenance can be obtained.
[0029] In a microfluidic device according to embodiments of the
present invention, the at least one detector may be a detector
array, and the microfluidic channel may be interlayered between the
radiation carrier and the detector array.
[0030] It is an advantage of embodiments of the present invention
that a compact and simple flow cytometer can be obtained.
[0031] In a further embodiment, the present invention provides a
system that comprises, as separate devices
[0032] a microfluidic chip comprising at least one microfluidic
channel, and
at least one radiation carrier for carrying at least a radiation
beam, the radiation carrier comprising a surface with at least one
excitation grating, positioned and adapted to couple an excitation
radiation signal directionally out of the radiation carrier thereby
illuminating a pre-defined volume of the microfluidic channel, and
at least one structure, e.g. emission grating or planar optics,
positioned and adapted to redirect, e.g. reflect, emission
radiation origination from the pre-defined volume; and
[0033] a readout device, adapted to be operatively coupled with the
microfluidic chip, wherein the readout device comprises at least
one detector for detecting the redirected emission radiation
originating from the pre-defined volume, when the microfluidic chip
and the readout device are operatively coupled.
[0034] It is an advantage of a system according to embodiments of
the present invention that sensitive detectors, which are more
expensive, may be used. By having such sensor present in a separate
readout device, the sensor can be reused, rather than being
disposable. The use of sensitive detectors allows to do detection
in a high throughput system.
[0035] Because of the detector not being on a disposable chip in
these embodiments, hence the distance between the source of
radiation and the detector, the emitted radiation must travel a
distance which may be several mm to cm. Therefore the radiation may
have to be collimated if a usable amount of radiation should hit
the detector.
[0036] In a system according to embodiments of the present
invention, the readout device may comprise a slot for receiving the
microfluidic chip.
[0037] In yet another embodiment, the present invention provides a
diagnostic device comprising a sensor according to embodiments of
the present invention, and an output unit for providing an output
of the sensor on which a diagnose can be based. The output unit may
be adapted for outputting a signal representative for
presence/absence or concentration of an analyte in a pre-defined
volume of the microfluidic channel.
[0038] In a further embodiment, the present invention provides a
method of performing particle detection. The method comprises
[0039] providing radiation scattering centers,
[0040] inserting the radiation scattering centers within a region
of interest,
[0041] providing radiation from an excitation grating in optical
contact with the region of interest,
[0042] redirecting radiation scattered from radiation scattering
centers in the region of interest, by means of at least one
structure, e.g. emission grating or planar optics, to at least one
detector, and
[0043] monitoring emission of radiation redirected from the region
of interest.
[0044] It is an advantage of embodiments of the present invention
that the use of flat optics such as planar waveguides, gratings and
Fresnel lenses may reduce or avoid alignment steps, and less
maintenance may be required.
[0045] In a method according to embodiments of the present
invention, providing radiation scattering centers may comprise
attaching radiation scattering centers to analytes.
[0046] It is an advantage of embodiments of the present invention
that luminescence, e.g. fluorescence, cytometry can be used with
the present method.
[0047] In a method according to embodiments of the present
invention, attaching radiation scattering centers may comprise
attaching at least one type of luminophores, e.g. fluorophores, or
chromatophores, or a mixture thereof.
[0048] In a method according to embodiments of the present
invention, inserting scattering centers within a region of interest
may further comprise providing a flow of scattering centers through
the region of interest. In alternative embodiments, inserting
scattering centers within a region of interest may comprise
attaching analyte carrying scattering centers to affinity
probes.
[0049] It is an advantage of these embodiments that scattering
centers shall not be fixed to the affinity probe directly, hence
the noise is substantially reduced as the majority of the detected
signal may mainly stem from fixed analytes.
[0050] Particular and preferred aspects of the invention are set
out in the accompanying independent and dependent claims. Features
from the dependent claims may be combined with features of the
independent claims and with features of other dependent claims as
appropriate and not merely as explicitly set out in the claims.
[0051] These and other aspects of the invention will be apparent
from and elucidated with reference to the embodiment(s) described
hereinafter.
BRIEF DESCRIPTION OF THE DRAWINGS
[0052] FIG. 1 illustrates a lateral view of a radiation carrier
according to embodiments of the present invention, comprising, as
an example only, and not intended to be limiting for the present
invention, excitation and emission gratings, a region of interest
(ROI) and at least one detector.
[0053] FIG. 2 illustrates a schematic perspective view of a planar
waveguide according to embodiments of the present invention,
comprising, as an example only, and not intended to be limiting for
the present invention, excitation and emission gratings, a ROI, two
detectors and a forward-scattering detector.
[0054] FIG. 3 shows a model of the angular distribution of the
radiation from an oscillating dipole radiator before and after
incidence on a collimating holographic detector.
[0055] FIG. 4 illustrates the front view of a planar waveguide
according to embodiments of the present invention, with spread
excitation gratings and focusing emission gratings, a system for
introducing analytes in the ROI and three diagrams showing the
results in time of the forward-scattering detector and the
detectors receiving radiation from the emission gratings.
[0056] FIG. 5 illustrates the front view of an alternative
arrangement of detectors with respect to the gratings, according to
embodiments of the present invention.
[0057] FIG. 6 illustrates a flowchart of method according to
embodiments of the present invention.
[0058] The drawings are only schematic and are non-limiting. In the
drawings, the size of some of the elements may be exaggerated and
not drawn on scale for illustrative purposes.
[0059] Any reference signs in the claims shall not be construed as
limiting the scope.
[0060] In the different drawings, the same reference signs refer to
the same or analogous elements.
DETAILED DESCRIPTION OF ILLUSTRATIVE EMBODIMENTS
[0061] The present invention will be described with respect to
particular embodiments and with reference to certain drawings but
the invention is not limited thereto but only by the claims. The
drawings described are only schematic and are non-limiting. In the
drawings, the size of some of the elements may be exaggerated and
not drawn on scale for illustrative purposes. The dimensions and
the relative dimensions do not correspond to actual reductions to
practice of the invention.
[0062] The terms first, second and the like in the description and
in the claims, are used for distinguishing between similar elements
and not necessarily for describing a sequence, either temporally,
spatially, in ranking or in any other manner. It is to be
understood that the terms so used are interchangeable under
appropriate circumstances and that the embodiments of the invention
described herein are capable of operation in other sequences than
described or illustrated herein.
[0063] Moreover, the terms top, under and the like in the
description and the claims are used for descriptive purposes and
not necessarily for describing relative positions. It is to be
understood that the terms so used are interchangeable under
appropriate circumstances and that the embodiments of the invention
described herein are capable of operation in other orientations
than described or illustrated herein.
[0064] It is to be noticed that the term "comprising", used in the
claims, should not be interpreted as being restricted to the means
listed thereafter; it does not exclude other elements or steps. It
is thus to be interpreted as specifying the presence of the stated
features, integers, steps or components as referred to, but does
not preclude the presence or addition of one or more other
features, integers, steps or components, or groups thereof. Thus,
the scope of the expression "a device comprising means A and B"
should not be limited to devices consisting only of components A
and B. It means that with respect to the present invention, the
only relevant components of the device are A and B.
[0065] Reference throughout this specification to "one embodiment"
or "an embodiment" means that a particular feature, structure or
characteristic described in connection with the embodiment is
included in at least one embodiment of the present invention. Thus,
appearances of the phrases "in one embodiment" or "in an
embodiment" in various places throughout this specification are not
necessarily all referring to the same embodiment, but may.
Furthermore, the particular features, structures or characteristics
may be combined in any suitable manner, as would be apparent to one
of ordinary skill in the art from this disclosure, in one or more
embodiments.
[0066] Similarly it should be appreciated that in the description
of exemplary embodiments of the invention, various features of the
invention are sometimes grouped together in a single embodiment,
figure, or description thereof for the purpose of streamlining the
disclosure and aiding in the understanding of one or more of the
various inventive aspects. This method of disclosure, however, is
not to be interpreted as reflecting an intention that the claimed
invention requires more features than are expressly recited in each
claim. Rather, as the following claims reflect, inventive aspects
lie in less than all features of a single foregoing disclosed
embodiment. Thus, the claims following the detailed description are
hereby expressly incorporated into this detailed description, with
each claim standing on its own as a separate embodiment of this
invention.
[0067] Furthermore, while some embodiments described herein include
some but not other features included in other embodiments,
combinations of features of different embodiments are meant to be
within the scope of the invention, and form different embodiments,
as would be understood by those in the art. For example, in the
following claims, any of the claimed embodiments can be used in any
combination.
[0068] In the description provided herein, numerous specific
details are set forth. However, it is understood that embodiments
of the invention may be practiced without these specific details.
In other instances, well-known methods, structures and techniques
have not been shown in detail in order not to obscure an
understanding of this description.
[0069] Where in embodiments of the present invention reference is
made to "a particle" or "particles", this may refer to biological
material such as, but not limited thereto, cells, exosomes,
viruses.
[0070] Where in embodiments of the present invention, reference is
made to "a fluid sample", this may refer to a fluid of a biological
nature, e.g. a body fluid such as, but not limited to, blood,
saliva, urine. The fluid sample may also refer to a fluid of a
non-biological nature but suitable for transporting a particle as
defined above, e.g. a saline solution.
[0071] Where in embodiments of the present invention reference is
made to "planar laser beam", reference is made to a laser sheet,
for example a laser beam spread and formed into a thin sheet by a
long focal length spherical lens and a cylindrical lens. Any
suitable system may be used. A "planar waveguide" is understood as
a slab waveguide with substantially parallel flat surfaces, so the
radiation travels inside via total internal reflection.
[0072] In embodiments of the present invention comprising planar
waveguides, a grating (e.g. an out-coupling grating) may be
provided on the walls of the planar waveguide, disrupting the
internal reflection and producing a beam of radiation leaving the
waveguide. This beam may be used as an excitation beam for
analyzing samples or particles, and it is referred to as
"excitation grating". Likewise, a grating that receives the beam
after interaction with a sample or particle, and redirects it to a
detector, e.g. reflects the radiation or couples the radiation into
a waveguide, will be referred to as a "emission grating". However,
the present invention is not limited to said waveguides, nor to
optical lasers, nor to the presence of emission gratings. In the
most general form, the radiation carrier comprises a structure for
redirecting emission radiation emanating from the region of
interest.
[0073] Where in embodiments of the present invention reference is
made to "region of interest" or "ROI", reference is made to a
predetermined region or volume of the space which can be occupied
by a detectable specimen, such as a particle or a cell. In some
embodiments of the present invention, the ROI comprises a portion
of a microchannel, for example in a microfluidic device.
[0074] Where in embodiments of the present invention reference is
made to "optical sensor", reference is made to a device suitable
for sensing photons, for example using IR radiation, visible
radiation, UV, etc.
[0075] Where in embodiments of the present invention reference is
made to "luminescence of a target", reference is made to emission
of radiation by the target, not resulting from thermal emission.
Typically, in the context of the present invention, luminescence
will be photoluminescence, generated by absorption of photons; such
as fluorescence or phosphorescence. The present invention, however,
is not limited to this type of luminescence, and can also be
applied in case of, for instance, bioluminescence or
chemiluminescence (emission as a result of a (bio)chemical reaction
by an organism) or electroluminescence (a result of an electric
current passed through the target).
[0076] Where in embodiments of the present invention reference is
made to "Raman scattering on the target", reference is made to
photons being scattered from the target when the latter is
illuminated. Reference is made more particularly to inelastic
scattering, where photons are scattered by an excitation, with the
scattered photons having a frequency different from that of the
incident photons. The Raman effect differs from the process of
photoluminescence in that for the latter, the incident radiation is
absorbed and the system is transferred to an excited state from
which it can go to various lower states. The result of both
processes is in essence the same: a photon with a frequency
different from that of the incident photon is produced, and the
molecule is brought to a different energy level. The major
difference is that the Raman effect can take place for any
frequency of incident radiation, while photoluminescence occurs
only at a particular frequency of incident radiation.
[0077] Where in embodiments of the present invention, reference is
made to "affinity probes", this refers to the substance having a
certain affinity, e.g. a natural attraction, to the analyte, the
substance having or not having a biological origin. By the
expression "substance having a biological origin", we intend to
mean a substance that is present or produced in a living organism,
or has similar properties and/or structure and/or composition. For
instance, the affinity probe may be an antibody, an antigen, an
enzyme, a receptor, an aptamer, a nucleic acid aptamer, a peptide
aptamer, or a molecularly imprinted polymer (MIP). In one aspect,
the present invention relates to an optical sensor suitable for
particle analysis, such as analysis via flow cytometry, the present
invention not being limited thereto. The optical sensor comprises a
radiation source, advantageously a substantially coherent radiation
source (e.g. laser). Radiation from the radiation source may be
guided or transported by a waveguide. At least one excitation
grating may be provided on the waveguide to direct the radiation
beam towards a ROI, which may comprise a particle, a plurality of
analytes in a flow of particles, etc. The radiation beam is made to
interact with the at least one particle, which may be fluorescent
in itself, or may be labeled with a fluorescent label. One or more
structures, for instance emission gratings, may collect the
radiation scattered from the ROI and redirect, e.g. reflect, it
into at least one detector. The structures, e.g. emission gratings,
may for example collimate the radiation from the ROI (upon
redirecting, e.g. reflecting, it into one or more detectors), but
the present invention is not limited to collimation, and
alternatively the structures, e.g. emission gratings, may focus the
redirected, e.g. reflected, radiation into one or more
detectors.
[0078] In embodiments of the present invention, the radiation
source couples radiation into a radiation carrier comprising at
least one excitation grating, for outcoupling radiation from the
radiation source. In some embodiments, the radiation carrier is
optimized for carrying laser beams. In some embodiments, the
radiation carrier may be waveguide, for example a strip or planar
waveguide or a slab waveguide.
[0079] The excitation grating on the radiation carrier, e.g.
waveguide, may be a focusing grating, or a grating providing e.g. a
planar excitation beam, and it may be patterned as a grating
coupler, the present invention not limited thereto. For example the
focusing grating may comprise planar dielectric grating reflectors
with focusing abilities, a Fresnel lens, etc. The excitation
grating may be patterned, oriented or adapted to direct or focus
radiation on a ROI, for example it may comprise gratings and
patterns on the surface of a waveguide, so upon passage of a laser
beam travelling in a radiation carrier, e.g. waveguide, the beam
may exit the radiation carrier and be directed to, e.g. focused
into, a ROI. The structure may be adapted to focus the radiation in
a volume of substantially similar size as the expected cells. If
the radiation is focused in a volume much smaller than a cell, the
reliability of the detection system is reduced due to the strong
variation of the signal. In embodiments of the present invention,
on the other hand, the radiation is not necessarily focused on a
volume smaller than a cell. A consistent illumination is obtained,
which increases the reliability. Focusing radiation in a volume of
size similar to the size of a cell may be done for instance by a
dotted lens, which is a metalens formed by a structure of pillar
elements in a close grid. A phase change is caused by passing the
radiation through the pillar elements. The phase change can be very
accurately tuned. Furthermore, the pillar design can be a good
basis to create lenses with additional functionality, such as a
strong spectral change. The excitation grating may also, instead of
focusing, spread the radiation on a ROI (e.g. providing a planar
laser beam), for example on a line or an area of a transparent
conduit such as a microfluidic channel. It may comprise material
suitable for transmission of the radiation, such as silicon
nitride. The control of illumination in the ROI advantageously
reduces noise, because the ultimately detected signal may stem
solely from the ROI and not from neighboring regions.
[0080] The radiation carrier, e.g. waveguide, comprises at least
one structure, for instance an emission grating or planar optics,
for redirecting, e.g. reflecting, any radiation emanating from the
ROI to one or more detectors. In some embodiments, the redirected,
e.g. reflected, radiation is collimated to the one or more
detectors. Alternatively, the structure, e.g. gratings, may focus
the radiation towards the one or more detectors, rather than
collimating the radiation. In some embodiments of the present
invention, radiation from the ROI may be laser radiation scattered
for example by fluorescence, and it may be redirected, e.g.
reflected, by the structure, e.g. emission grating, into one or
more detectors. The one or more structures may comprise a Fresnel
lens, or any suitable optical element.
[0081] The structure, e.g. emission grating, can be a dielectric
reflector. In embodiments of the present invention, the structure,
e.g. emission grating, may comprise reflective material such as
metal, for example it may comprise a layer of reflective metal, or
it can be formed by a combination of a dielectric grating and a
reflective metal surface. For example, one or more structures, e.g.
emission gratings, may be coplanar and may be located next to the
excitation grating.
[0082] In accordance with embodiments of the present invention, a
structure, e.g. an emission grating, can be used either in
reflection mode or in transmission mode. When used in reflection
mode, the detector will be located at a same side of the radiation
carrier as the structure, e.g. emission grating. When used in
transmission mode, the detector will be located at an opposite side
of the radiation carrier compared to the structure, e.g. emission
grating, and the detected radiation is sent substantially
transversally through the radiation carrier to a detector. In the
latter case, the structure, e.g. emission grating, may for instance
be formed by a Fresnel lens that directs radiation to the other
side of the radiation carrier.
[0083] In embodiments of the present invention comprising a planar
waveguide, the at least one structure, e.g. emission grating, and
the excitation grating may extend on a same surface of the
waveguide. In some embodiments of the present invention, the
structure may be implemented as an emission grating, and the
emission grating and the excitation grating may be combined into a
single grating region. Thus one continuous grating surface,
comprising an emission grating and an excitation grating, may be
formed, with optimized patterns in different zones for one or
another behavior (obtaining excitation beam or reflecting
radiation). Additionally, part of the continuous surface may
comprise solely dielectric grating, while part of the surface (the
emission gratings) may comprise an additional reflective layer such
as a metal layer. The grating surface may have a homogeneous or
preferably an inhomogeneous pattern in the whole surface. In
embodiments of the present invention the same type of gratings can
be used for the emission grating and the excitation grating. In
such case, in particular Fresnel lenses can be used. Dielectric
gratings can also be used, but since these are designed for
particular wavelengths, they might cause more aberrations.
[0084] For example, a laser beam traveling through the radiation
carrier may be transmitted through a grating acting as an
excitation grating and may be focused on a ROI, in a point or on a
line. Any radiation scattered by particles in the ROI may be
reflected by the same grating surface, but by the section acting as
an emission grating, into a detector. This may be possible by fine
tuning of the patterning, for example.
[0085] In embodiments of the present invention, different zones of
the structure positioned and adapted for redirecting emission
radiation emanating from a region of interest, e.g. different zones
of an emission grating, may be adapted to redirect, e.g. reflect,
different parts of the spectrum, e.g. by adapting its grating,
properties of a reflective layer, etc. Hence it is possible to
include particle discrimination by redirecting, e.g. reflecting,
signals from a first predetermined wavelength range to a first
detector and from a further predetermined wavelength range to a
different further detector. For example, a first type of scattering
centers (e.g. a first type of fluorophores) may attach to a first
type of analyte, while a further type of scattering centers (e.g.
one or more different types of fluorophores) may attach to a
further type of analyte. In this case, the first analyte may be
detected by a first detector while the further analyte may be
detected by a further detector. The signals from first and second
analyte may e.g. be signals from different fluorescent markers,
e.g. different fluorophores labelling correspondingly different
types of cells, viruses, exosomes, etc.
[0086] Particle discrimination may be obtained alternatively or
additionally by including a filter on the structure, e.g. emission
grating, or on the detection system. With a diffractive grating,
part of the filtering may be done by the grating itself. The at
least one detector may be a plurality of detectors, such as
specialized detectors, which may be only sensitive to a part of the
spectrum, e.g. infrared or ultraviolet detectors, or detectors of
radiation within a particular region of the visible range.
According to some embodiments of the present invention, a plurality
of different spectral filters, each filter having a different
central wavelength, may filter the signal before it reaches a
corresponding detector in a system with a plurality of detectors.
In advantageous embodiments, the plurality of detectors may be used
to each detect a signal corresponding to a different part of the
spectrum.
[0087] It is an advantage of embodiments of the present invention
that both excitation and collection systems may be aligned or at
least roughly aligned, reducing or even avoiding calibration and
alignment steps. Additionally, the system so built may be compact
and low cost, as it does not require multiple pieces or complex
assembling. It may be advantageously implemented in medical
devices, such as flow cytometry systems, and it may be easily
applied in portable devices. For example, the sensor may be used in
flow cytometry and an additional detector, collinear with the
source of radiation and the ROI, may be used for counting particles
and obtaining data regarding characteristic compound on a cell.
Scattering date may be used to determine size. For biological
applications, inorganic and organic dyes may be used. For example,
one or more types of target cells, or viruses, or any other
analyte, may be labelled with tagging antibodies comprising
chromatophores, fluorophores, etc.
[0088] Some embodiments of the present invention may be applied to
detect and analyze quantum dots, for example using UV laser. This
may be useful in biological analysis (quantum dots as tagging
particles), but it may also be used in semiconductor technology. In
general, the present invention may suitable be for fields of
technology involving optical analysis of particles.
[0089] Some embodiments of the sensor of the present invention are
described with reference to FIG. 1, FIG. 2 and FIG. 4, FIG. 5.
[0090] In what follows, particular embodiments of the present
invention are described. These often refer to "waveguides" rather
than the broader "radiation carrier", to "emission gratings" rather
than the broader "structure", and to "reflecting" rather than the
broader "redirecting". This way of describing is done for the
purpose of understandability and fixing the mind, and is in no way
intended to be limiting for the present invention.
[0091] FIG. 1 shows a lateral view of a radiation carrier 100, such
as a waveguide, comprising a grating 101 on its top surface 102.
The radiation carrier 100 may be an optical fiber, or more
preferably a rectangular waveguide. The radiation carrier 100 may
be made from any suitable material, such as for instance glass,
polymer or suitable semiconductor material. Radiation 103,
emanating from a radiation source (not illustrated) is coupled into
the radiation carrier 100, and travels there through, for instance
by total internal reflection, until it exits through an excitation
grating 101. The thus exciting excitation beam 104 may be adapted
for being focused into a ROI 105, which in the figure illustrated
contains a particle 106, for example a cell. The grating 101 may be
adapted to focus the beam 104 into the whole ROI, or more than half
of the ROI, or advantageously into a volume of the same order of
magnitude as the particles to be analyzed, for example a volume of
one or more cells. The radiation 107 scattered from the particle
106 present in the ROI 105, for example scattered by fluorescence,
falls into an emission grating 108, where it is reflected and
collimated, such that the reflected and collimated beam 109 is
suited for entering a detector 110. In the embodiment illustrated
in FIG. 1, only one emission grating is shown, but the invention is
not limited thereto, and also encompasses embodiments with more
than one emission grating.
[0092] FIG. 2 shows a perspective view of a planar waveguide 200
comprising an excitation grating 201 on the waveguide surface 202,
with a radiation signal 203 travelling within the waveguide 200,
for example via total internal reflection. The excitation grating
201 disturbs the surface 202 of the planar waveguide 200, and
radiation escapes from the waveguide 200, thus forming an
excitation beam 204. The excitation grating 201 has a pattern that
allows focusing the excitation beam 204 on a ROI 205. When the
radiation in the ROI 205 encounters a fluorescent analyte 206, e.g.
an analyte such as a cell showing fluorescence (e.g. by attachment
of fluorescent markers), it scatters radiation which is reflected
and collimated in one or more emission gratings 208. If there are a
plurality of emission gratings 208, they may be located at either
side of the excitation grating 201, for instance the plurality of
emission gratings 208 may surround the excitation grating 201. The
plurality of emission gratings 208 may be evenly or unevenly
distributed around the excitation grating 201. The radiation 207
reflected and optionally collimated by the emission gratings 208,
enters a couple of detectors 211, 212. In particular embodiments of
the present invention, rather than being collimated, the radiation
reflected by the at least one emission grating 208 may be focused
onto a detector surface.
[0093] In some embodiments of the present invention, a further
(optional) forward-detector 213 may be placed so as to detect a
shadow as a hologram of the cells moving through the ROI. The
forward detector 213 detects along the same axis as the incoupled
excitation beam 204. In embodiments of the present invention, the
signal from the forward detector 213 can be used as comparative
signal (e.g. to detect whether a particle 206 or analyte passes
through the ROI 205), or as an indication of the size of the
particle 206 or analyte in the ROI 205. This may be advantageous
for distinguishing different bodies passing through the ROI 205,
for example for distinguishing bodies with attached luminophores,
e.g. fluorophores, from unattached luminophores, e.g.
fluorophores.
[0094] The patterns of the gratings in accordance with embodiments
of the present invention, e.g. the excitation and/or emission
gratings, may be made so as to spread, collimate or focus the
radiation. Different types of gratings (excitation grating,
emission grating) may have a different characteristic. For example,
the excitation gratings 101, 201 of FIG. 1 and FIG. 2 may be
adapted (e.g. patterned, by adding a lens system, by forming a
Fresnel lens, etc.) for focusing the excitation beam 104, 204 which
exits the radiation carrier 100, 200, into the ROI 105, 205. In
embodiments of the present invention, no extra patterning, like an
extra Fresnel pattern, is needed to focus the excitation beam 104,
204 coupled out of the waveguide, as the excitation grating per se
can do the job.
[0095] At the same time, the one or more emission gratings 108, 208
of FIG. 1 and FIG. 2 may be patterned for collimating the reflected
radiation 107, 207 into the detectors 211, 212. Collimation using
emission gratings 108, 208 can be highly accurate.
[0096] The diagrams of FIG. 3 show a model of the angular
distribution of radiation from an oscillating dipole radiator,
which can be used for modelling a point source radiator, e.g. a
Raman scattering molecule or a luminescent molecule such as e.g. a
fluorescent molecule. The left diagram 300 of FIG. 3 shows the
angular distribution of the radiation reaching the emission
grating. It is spread over an angle between approximately
35.degree. (on the 45.degree. direction) and 25.degree. (on the
90.degree. direction). On the other hand, the right hand diagram
310 of FIG. 3 shows the angular distribution of the radiation after
being reflected and collimated by the emission grating. The angular
distribution of the radiation is concentrated in a single
direction, at 10.degree..
[0097] The present invention also encompasses types of excitation
and emission gratings other than the ones shown in FIG. 1 and FIG.
2. For example, FIG. 4 shows a front view of a planar waveguide 400
comprising an excitation grating 401 on a surface 402, surrounded
by a first and second emission gratings 403a, 403b. FIG. 4 also
schematically illustrates a radiation signal 203 traveling within
the waveguide 400. In the embodiment illustrated, the excitation
grating 401 spreads the exiting excitation beam 404 on the ROI 205.
For example, the spread excitation beam 404 may be planar and may
extend over a length defining the ROI 205, for instance it may span
the width of the microfluidic channel. In the embodiment
illustrated in FIG. 4, emission gratings 403a, 403b (for collecting
radiation reflected from the ROI and sending it to detectors) are
designed to focus the radiation in a point on a detection surface.
When the radiation in the ROI 205 encounters a scattering center
(e.g. a fluorescent analyte), the scattered radiation is collected
in the first and second emission gratings 403a, 403b. The emission
gratings 403a, 403b may each reflect radiation of the complete
spectrum, or they may reflect radiation within a first range of
wavelengths and a second range of wavelengths, respectively. The
radiation falling on the first and second emission gratings 403a,
403b may be reflected and focused (instead of being collimated) and
sent into detectors such as the ones of FIG. 2 or into a detector
array 405 like the one shown in FIG. 4. The detector array 405 may
comprise a plurality of detecting regions 406, 407, 408, for
example a plurality of regions 406, 408 for detecting radiation
scattered from the ROI and a region 407 for detecting shadows as a
hologram of the particles moving in the ROI 205. Hence, the lateral
position of cells in a ROI can be obtained as an image, for
example. The shadow of the excitation line that falls on detecting
region 407 can be used to verify the particles and possibly to do a
size measurement.
[0098] FIG. 1 and FIG. 2 illustrate embodiments with a focusing
excitation grating and one or more collimating emission gratings.
FIG. 4 illustrates an embodiment with a spreading excitation
grating and a plurality of focusing emission gratings. However,
this is not intended to be limiting for the present invention, and
also other combinations of types of excitation gratings and
emission gratings are envisioned to be part of the present
invention. For instance, in accordance with embodiments of the
present invention, any suitable type of excitation grating (e.g.
focusing, collimating, spreading) can be combined with any suitable
type of emission grating (e.g. focusing, collimating,
spreading).
[0099] Embodiments of the first aspect of the present invention may
further comprise microfluidic channels, for example a microfluidic
chip in combination with a radiation carrier, excitation and
emission gratings and optionally any planar or strip optics, and
one or more detectors. The one or more detectors can be integrated
in the microfluidic chip. For example, the imager can be on top of
the microfluidic chip.
[0100] An example of embodiments comprising a microfluidic channel
is shown in FIG. 4, in which a transparent substrate 409 comprises
a microfluidic channel 410. The region of the channel 410
illuminated by the excitation radiation 404, comprises the ROI 205.
In the case illustrated, the excitation grating 401 spreads the
radiation over substantially the whole of the width of the channel
410, optimizing the ROI 205 within the channel 410. The radiation
may be spread e.g. in a planar sheet, for example a planar laser
beam, although the present invention is not limited thereto. If a
particle, e.g. a fluorescent marker attached to an analyte, crosses
the ROI 205, excitation radiation is scattered on or by the
fluorescent marker. Back-scattered radiation from the ROI 205 is in
this case reflected and focused by the emission gratings 403a, 403b
into a point just above the microfluidic chip, e.g. into the zones
406, 408 of a detector array 405, which may be a line array, a
camera, etc. The transparent substrate 409 may further focus the
radiation reflected by the emission gratings. This configuration
may advantageously simplify the microfluidics as well as increase
the throughput. The detectors may be placed a few millimeters away
from the emission grating, or at least a distance enough to allow
the definition of a ROI (e.g. allow the placement of microfluidic
channels for defining a ROI).
[0101] The three diagrams 420, 421 and 422 at the top of FIG. 4
show the signal measured by the detector array region 406 on the
left, the region 407 on the center and the region 408 on the right,
respectively, as a function of time. The central diagram 421, the
shadow or hologram region, would detect the passing of particles
(e.g. via forward-scattering). The left diagram 420 may detect one
type of scattering centers, e.g. red fluorophores attached to a
first type of analytes, while the right diagram 422 may detect a
second type of scattering centers, e.g. green fluorophores attached
to a second type of analytes. The analysis of the graphs produces a
reconstruction of the particles flowing through the channel, in
addition to the fluorescent signal that matches these
particles.
[0102] The present invention is not limited to the distribution of
optical elements as illustrated in FIG. 4. For example, rather than
being place above (in a direction of excitation of radiation) a
radiation carrier, as illustrated in FIG. 4, detector arrays may be
placed aside the radiation carrier, e.g. waveguide, as illustrated
in FIG. 5. In such embodiments, rather than having a unique array
of detectors in a single plane, a plurality of detectors planes may
be available.
[0103] FIG. 5 shows two detector arrays 501, 502 substantially
perpendicular to the surface of the radiation carrier 400
containing the excitation grating 401 (e.g. a grating for spreading
the beam as in FIG. 4) and the emission gratings 503, 504 (e.g.
collimating gratings). The radiation scattered from the ROI 205 may
be collected in the detector arrays 501, 502 after reflection by
the emission gratings 503, 504 (focused or, as shown in the image,
collimated). This geometry may be advantageous for avoiding
circuitry or other elements in the top part of the circuit. For
instance, analytes may be introduced through the top of the device
into the ROI 205, instead of through the zone between the
excitation grating 401 and the detector array. For example, the
embodiment of FIG. 5 may comprise affinity probes in the ROI
205.
[0104] In embodiments of the present invention, the microfluidics
are provided, e.g. patterned, in or on top of a chip (e.g. a CMOS
chip) and are closed by a transparent cover, so that the radiation
(e.g. light) reflected by the emission gratings can reach at least
one detector. The microfluidics may force a fluid comprising
particles through a channel comprising the ROI, for instance by
capillary action or driven e.g. by using pumps or similar, so that
the particles interact with the focus spot of the excitation
grating and emission gratings. Embodiments of the present invention
may further comprise a first and a second microfluidic compartment
fluidically interconnected via at least one micro-fluidic channel
comprising a ROI. The second micro-fluidic compartment may comprise
or may be connected to a capillary pump for pumping a fluid sample
from the first to the second micro-fluidic compartment via the at
least one microfluidic channel. The chip may comprise also an
on-chip radiation source such as a light source, optically coupled
to the radiation carrier (e.g. a waveguide).
[0105] Embodiments of the first aspect of the present invention
have been described where particles to be detected or analyzed are
in flow. Alternative embodiments of the present invention may
comprise a substrate comprising affinity probes suitable for
binding the particles under interest, for being investigated in a
static situation. In these embodiments, analytes are fixed to at
least a portion of a substrate provided with relevant affinity
probes, e.g. antibodies, antigens, enzymes, receptors, aptamers,
nucleic acid aptamers, peptide aptamers or molecularly imprint
polymers (MIP). At least a portion of the substrate comprising thus
fixed analytes is placed in the ROI. The analytes, as before, may
further comprise one or more types of attached scattering centers.
It is an advantage of these embodiments that scattering centers not
bound to analyte to be investigated shall not be fixed to the
affinity probes, hence the noise is substantially reduced as the
majority of the detected signal may mainly stem from fixed
analytes.
[0106] Further embodiments of the present invention may comprise a
waveguide and multiple excitation gratings, for irradiating, e.g.
illuminating, multiple ROIs, e.g. in a single or a plurality of
microfluidic channels (e.g. comprising particles attached to
different types of luminophores, e.g. fluorophores), or in a
plurality of affinity probes. This allows reducing power
consumption because only one radiation source may be needed.
[0107] Other features may be included, such as coupling gratings,
tapers, lenses like Fresnel lenses, microlens arrays, etc.
[0108] In a second aspect, the present invention relates to a
method of performing particle detection. The method is suitable for
detecting analytes, e.g. Raman scattering particles or luminophore
labeled particles, such as fluorophore labelled particles, although
the present invention is not limited thereto. The method includes
irradiating particles or cells in a ROI (e.g. a volume of the same
order of magnitude of the particles or cells to be analyzed), for
example using an excitation grating for producing emission of
radiation characteristic of the particle or cell, collecting that
emitted radiation in at least one grating, and sending (e.g. by
reflection) said radiation to a detector (e.g. optical detector,
fluorescence detector, etc.). The method will be described with
respect to the flowchart of FIG. 6.
[0109] In a first step, providing 600 scattering centers may
comprise providing particles that scatter radiation within a
particular wavelength range, for example laser radiation. The
scattering centers may for instance be fluorescent labels. The
scattering centers may be attached 601 to analytes. For example
different types of scattering centers, having the feature of
radiation scattering at different wavelengths, may be attached to
different types of analytes. For example, one type of scattering
center may be attached to tumor cells while others may be attached
to healthy cells.
[0110] The scattering centers may be present in a fluid, such as
blood, urine, saliva, buffer, a solution, etc., and providing
scattering centers may comprise binding scattering centers to
analyte while the analyte is present in the bulk of a liquid,
optionally in flow. Alternatively, the analyte may be bound to
affinity probes, and providing scattering centers may comprise
binding scattering centers to analyte bound to affinity probes.
[0111] A further step comprises outcoupling 610 radiation from a
radiation carrier, via an excitation grating. Providing radiation
may comprise providing 611 laser radiation, with a wavelength for
example between IR and UV wavelengths. The type of radiation and
its characteristics can be selected to obtain a suitable scattering
of the scattering centers, for example via fluorescence. The
radiation may be provided continuously or discontinuously.
[0112] For example, in embodiments comprising providing 602 a flow
of analytes, it may be preferable to provide 612 continuous
radiation, while in embodiments comprising attaching 603 analytes
to affinity probes, it may be feasible or preferable to provide 613
discontinuous, e.g. pulsed, radiation.
[0113] A further step comprises inserting 620 the scattering
centers within a ROI. For example, they may be introduced 621 in a
fluid through a microfluidic channel (in flow), or affinity probes
may be placed 622 in the ROI, to which analyte under interest has
bound or may bind.
[0114] The interaction of the radiation beam from the excitation
grating with the scattering centers will produce scattered
radiation, e.g. fluorescence, which shall be collected and directed
630 from the ROI, via emission gratings, to at least one detector.
Directing 630 radiation from the ROI to a detector may comprise
directing 631 radiation within a predetermined wavelength range to
a predetermined detector, and radiation within a further
predetermined wavelength range to another predetermined detector.
Additionally, the reflection in the emission grating may comprise
either focusing 632 or collimating 633 the radiation to the at
least one detector.
[0115] A further step comprises monitoring 640 emission of
radiation from the ROI. This step comprises monitoring emissions
reflected by the one or more emission gratings, and it may further
comprise monitoring forward scattered radiation. For the step of
monitoring 640 any suitable technique may be used, such as
photoelectric cells, analog to digital converters, outputs, etc.
Additionally, a step of filtering 641 radiation within a
predetermined range of wavelengths may be included, for example a
threshold filter, chromatic filter, polarization filter, etc.
Further steps such as performing 642 peak detection or labelling
643 scattering centers may be applied.
[0116] Different particles may present different response to the
same radiation. It is possible to discern between different
responses in embodiments of the present invention, e.g. by use of
filters for filtering the radiation impinging on the detectors. For
example, some embodiments of the present invention may comprise
laser-induced fluorescence. In such embodiments, different types of
luminophores, e.g. fluorophores, may be used wherein each type has
a different wavelength range. To differentiate different radiation
signals having different wavelength ranges, different spectral
filters may be used in the detector or detectors to filter the
emission radiation signal. As an advantage, detection of emission
radiation may be performed more efficiently. For example, a peak in
the emission radiation signal may be detected more efficiently.
[0117] For example, when at least two types of luminophores, e.g.
fluorophores, are used to label a particle, a single emission
waveguide may be optically connected to an optical detector having
at least two spectral filters. The optical detector may comprise at
least two photodiodes, each photodiode being covered with a
different spectral filter. Luminescence, e.g. fluorescence, falling
onto the optical detector is filtered by each spectral filter
before being detected. This gives rise to at least two
luminescence, e.g. fluorescence, signals which may be correlated to
improve peak detection.
[0118] The present invention may be used for cytometry, like flow
cytometry. It may be applied to immunophenotyping, ploidy analysis,
cell counting or GFP expression analysis. The method and device are
advantageous for luminescent flow cytometry, e.g. fluorescent flow
cytometry, as a compact and low cost device is obtained, which may
be integrated in a chip. It may be easy to use, as it requires
little alignment and it is easy to implement in medical devices,
either as a microfluidic device or with affinity probes.
[0119] In embodiments of the present invention, the sensor
comprising both the radiation carrier with the at least one
excitation grating, the at least one structure positioned and
adapted for redirecting emission radiation emanating from a
pre-defined volume, and the at least one detector may be integrally
built, i.e. may be a single device. Alternative embodiments of the
present invention, however, also cover a system comprising
different separable parts, e.g. a sample analyzing device and a
readout device. The sample analyzing device may be a microfluidic
chip comprising at least one microfluidic channel for transporting
a fluid sample through the system, and at least one radiation
carrier for carrying a radiation beam. The radiation carrier
comprises, for instance on a surface thereof, at least one
excitation grating, positioned and adapted to couple the excitation
radiation signal carried by the radiation carrier directionally out
of the radiation carrier thereby illuminating a pre-defined volume
of the microfluidic channel, and at least one structures such as
for instance an emission grating, positioned and adapted to
redirect, e.g. reflect, emission radiation origination from the
pre-defined volume. The readout device may comprise at least one
detector, for instance a detector array, for detecting the
redirected, e.g. reflected, emission radiation originating from the
pre-defined volume. The readout device may be adapted, for instance
may be provided with a slot, for receiving the sample analyzing
device. The sample analyzing device may have the shape and size of
an SD card, for instance the shape and size of a micro-SD card or
similar. The sample analyzing device may include a radiation
source, e.g. a light source, for coupling radiation into the
radiation carrier. Alternatively, the radiation source may be
provided on or in the readout device, such that radiation may be
coupled into the radiation carrier of the sample analyzing device,
when the sample analyzing device and the readout device are
operatively coupled to one another. Device features of the system
comprising different separable parts are as explained in the
embodiments of the integrally built device, and are not repeated
here for sake of conciseness.
* * * * *