U.S. patent application number 15/159942 was filed with the patent office on 2016-09-15 for particle manipulation system with out-of-plane channel and focusing element.
This patent application is currently assigned to Owl biomedical, Inc.. The applicant listed for this patent is Owl biomedical, Inc.. Invention is credited to John S. FOSTER, Mehran R. Hoonejani, Stefan MILTENYI, Kamala R. Qalandar, Kevin E. Shields, Kimberly L. Turner.
Application Number | 20160263575 15/159942 |
Document ID | / |
Family ID | 51626460 |
Filed Date | 2016-09-15 |
United States Patent
Application |
20160263575 |
Kind Code |
A1 |
FOSTER; John S. ; et
al. |
September 15, 2016 |
Particle Manipulation System with Out-of-plane Channel and Focusing
Element
Abstract
A particle manipulation system uses a MEMS-based,
microfabricated particle manipulation device which has an inlet
channel, output channels, and a movable member formed on a
substrate. The movable member moves parallel to the fabrication
plane, as does fluid flowing in the inlet channel. The movable
member separates a target particle from the rest of the particles,
diverting it into an output channel. However, at least one output
channel is not parallel to the fabrication plane. The device may be
used to separate a target particle from non-target material in a
sample stream. The target particle may be, for example, a stem
cell, zygote, a cancer cell, a T-cell, a component of blood,
bacteria or DNA sample, for example. The particle manipulation
system may also include a microfluidic structure which focuses the
target particles in a particular portion of the inlet channel.
Inventors: |
FOSTER; John S.; (Santa
Barbara, CA) ; MILTENYI; Stefan; (Bergische Gladbach,
DE) ; Qalandar; Kamala R.; (Ojai, CA) ;
Shields; Kevin E.; (Santa Barbara, CA) ; Turner;
Kimberly L.; (Santa Barbara, CA) ; Hoonejani; Mehran
R.; (Goleta, CA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Owl biomedical, Inc. |
Goleta |
CA |
US |
|
|
Assignee: |
Owl biomedical, Inc.
Goleta
CA
|
Family ID: |
51626460 |
Appl. No.: |
15/159942 |
Filed: |
May 20, 2016 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
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13998095 |
Oct 1, 2013 |
9372144 |
|
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15159942 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
G01N 2201/06113
20130101; B01L 2400/0633 20130101; G01N 15/1404 20130101; F16K
99/0028 20130101; B01L 2300/0627 20130101; G01N 2021/6439 20130101;
B01L 2300/0864 20130101; F16K 99/0011 20130101; B01L 2300/0654
20130101; G01N 15/1484 20130101; G01N 21/6428 20130101; G01N
21/6486 20130101; B01L 3/502738 20130101; B01L 2200/0652 20130101;
B01L 3/502707 20130101; F16K 99/0046 20130101; F16K 2099/0084
20130101; G01N 21/6402 20130101; B01L 3/502715 20130101; F16K
99/0013 20130101; B01L 2400/0622 20130101; B01L 3/502761 20130101;
G01N 2015/149 20130101 |
International
Class: |
B01L 3/00 20060101
B01L003/00; G01N 21/64 20060101 G01N021/64 |
Claims
1. A micromechanical particle manipulation device, formed on a
surface of a substrate, comprising: a microfabricated, movable
member formed on a surface of the substrate in a first plane, and
having a first diverting surface, wherein the movable member moves
from a first position to a second position in response to a force
applied to the movable member, wherein the motion is substantially
in the first plane parallel to the surface of the substrate; a
sample inlet channel formed in the substrate and through which a
fluid flows, the fluid including target particles and non-target
material, wherein the flow in the sample inlet channel is
substantially in the first plane parallel to the surface of the
substrate; a sort output channel into which the microfabricated
member diverts the target particles and a waste output channel into
which the non-target material flows, and wherein the waste output
channel is in a second, different plane than the movable member and
the sample inlet channel, and wherein the waste output channel is
located directly below at least a portion of the microfabricated
member over at least a portion of its motion; and a sheath fluid
inlet in fluid communication with the sample channel; and a
focusing element coupled to the sheath fluid inlet, which is
configured to urge the target particles into a particular portion
of the sample channel.
2. The micromechanical particle manipulation device of claim 1,
wherein focusing element comprises a z-focus channel, wherein the
z-focus channel curves in an arc of about 180 degree from the
sheath fluid inlet, and urges the target particles into
substantially a single plane.
3. The micromechanical particle manipulation device of claim 2,
wherein the z-focus channel has a radius of curvature of at least
about 100 microns and less than about 500 microns.
4. The micromechanical particle manipulation device of claim 1,
wherein the focusing element is disposed in the same plane as the
movable member, and formed in the same substrate.
5. The micromechanical particle manipulation device of claim 2,
wherein the inlet channel and z-focus channel both have
characteristic dimensions of about 50 microns.
6. The micromechanical particle manipulation device of claim 2,
further comprising a y-intersection point at which the target
particles are compressed from the plane to a stream line near the
center of the z-focus channel.
7. The micromechanical particle manipulation device of claim 6,
wherein the y-intersection point occurs where two flows join the
z-focus channel from substantially antiparallel directions, and
each substantially orthogonal to the z-focus channel at the
y-intersection point.
8. The micromechanical particle manipulation device of claim 1,
wherein the target particles are at least one of a stem cell, a
cancer cell, a zygote, a protein, a T-cell, a bacteria, a component
of blood, and a DNA fragment.
9. The micromechanical particle manipulation device of claim 1,
wherein the first diverting surface has a shape which is
substantially tangent to the direction of flow in the input channel
at one point on the shape and substantially tangent to the
direction of flow of the first output channel at a second point on
the shape, and wherein the first diverting surface diverts flow
from the input channel into a first output channel when the movable
member is in the first position, and allows the flow into the
second output channel in the second position.
10. The micromechanical particle manipulation device of claim 1,
wherein the plurality of output channels comprises a sort channel
and a waste channel, wherein flow in the sort channel is
substantially antiparallel to flow in the input channel, and
wherein flow in the waste channel is substantially orthogonal to
flow in the input channel and the sort channel.
11. The micromechanical particle manipulation device of claim 1,
further comprising: a first permeable magnetic material inlaid in
the movable member; a first stationary permeable magnetic feature
disposed on the substrate; and a first source of magnetic flux
external to the movable member and substrate on which the movable
member is formed.
12. The micromechanical particle manipulation device of claim 11,
wherein the movable member moves from the first position to the
second position when the source of magnetic flux is activated.
13. The micromechanical particle manipulation device of claim 1,
wherein the force is at least one of magnetic, electrostatic, and
piezoelectric.
14. A particle manipulation system, comprising: the micromechanical
particle sorting device of claim 1; at least one laser directed to
a laser interrogation region disposed in the input channel; and at
least one set of detection optics that detects a fluorescent signal
from a fluorescent tag affixed to the target particle in the
fluid.
15. The particle manipulation system of claim 14, further
comprising: an electromagnet; and a circuit that provides a control
waveform to the electromagnet.
16. The particle manipulation system of claim 14, further
comprising: at least one additional laser directed at a region in
at least one of the output channels and configured to confirm
results of a particle manipulation.
17. The particle manipulation system of claim 15, wherein the
control waveform includes a higher amplitude acceleration phase
which sets the movable member in motion, a constant amplitude phase
which opens the movable member, and a braking phase which slows the
movable member at closure.
18. A method of making a microfabricated particle manipulation
device on a surface of a fabrication substrate, comprising: forming
a movable member on the surface of the substrate having a first
diverting surface, wherein the movable member is configured to move
from a first position to a second position in response to a force
applied to the movable member, wherein the motion is substantially
in a first plane parallel to the surface; forming a sample inlet
channel formed in the substrate and through which a fluid flows,
the fluid including at least one target particle and non-target
material, wherein the flow in the sample inlet channel is
substantially in the first plane parallel to the surface; forming a
plurality of output channels into which the microfabricated member
diverts the target particle, and wherein the flow in at least one
of the output channels is in a second different plane than the
movable member and the sample inlet channel, and wherein at least
one output channel is located directly below at least a portion of
the microfabricated diverter over at least a portion of its motion
forming a sheath fluid inlet in fluid communication with the sample
channel; and forming a focusing element coupled to the sheath fluid
inlet, which is configured to urge the target particles into a
particular portion of the sample channel, wherein the focusing
element is formed in the same substrate as the movable member and
inlet channel.
19. The method of making the microfabricated particle manipulation
device of claim 18, further comprising: forming the diverting
surface with a smoothly curved shape which is substantially tangent
to the direction of flow in the input channel at one point on the
shape and substantially tangent to the direction of flow of a first
output channel at a second point on the shape, wherein the first
diverting surface is configured to divert flow from the input
channel into the first output channel when the movable member is in
the first position, and allows the flow into a second output
channel in the second position; and wherein forming the focusing
element comprises forming a z-focus channel with a curved arc of
about 180 degrees from the sheath fluid inlet, wherein the z-focus
channel urges the target particles into substantially a single
plane.
20. The method of making the microfabricated particle manipulation
device of claim 19, further comprising: forming a sort channel and
a waste channel, wherein flow in the sort channel is substantially
antiparallel to flow in the input channel, and wherein flow in the
waste channel is substantially orthogonal to flow in the input
channel and the sort channel, and wherein the sort channel and
input channels are formed by etching trenches into the surface of
the substrate, and bonding a flat plate over the substrate and the
trenches; and wherein the z-focus channel is formed with a radius
of curvature of at least about 100 microns and less than about 500
microns.
Description
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] Not applicable.
STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH
[0002] Not applicable.
STATEMENT REGARDING MICROFICHE APPENDIX
[0003] Not applicable.
BACKGROUND
[0004] This invention relates to a system and method for
manipulating small particles in a microfabricated fluid
channel.
[0005] Microelectromechanical systems (MEMS) are very small, often
moveable structures made on a substrate using surface or bulk
lithographic processing techniques, such as those used to
manufacture semiconductor devices. MEMS devices may be moveable
actuators, sensors, valves, pistons, or switches, for example, with
characteristic dimensions of a few microns to hundreds of microns.
A moveable MEMS switch, for example, may be used to connect one or
more input terminals to one or more output terminals, all
microfabricated on a substrate. The actuation means for the
moveable switch may be thermal, piezoelectric, electrostatic, or
magnetic, for example. MEMS devices may be fabricated on a
semiconductor substrate which may manipulate particles passing by
the MEMS device in a fluid stream.
[0006] In another example, a MEMS devices may be a movable valve,
used as a sorting mechanism for sorting various particles from a
fluid stream, such as cells from blood. The particles may be
transported to the sorting device within the fluid stream enclosed
in a microchannel, which flows under pressure. Upon reaching the
MEMS sorting device, the sorting device directs the particles of
interest such as a blood stem cell, to a separate receptacle, and
directs the remainder of the fluid stream to a waste
receptacle.
[0007] MEMS-based cell sorter systems may have substantial
advantages over existing fluorescence-activated cell sorting
systems (FACS) known as flow cytometers. Flow cytometers are
generally large and expensive systems which sort cells based on a
fluorescence signal from a tag affixed to the cell of interest. The
cells are diluted and suspended in a sheath fluid, and then
separated into individual droplets via rapid decompression through
a nozzle. After ejection from a nozzle, the droplets are separated
into different bins electrostatically, based on the fluorescence
signal from the tag. Among the issues with these systems are cell
damage or loss of functionality due to the decompression, difficult
and costly sterilization procedures between sample, inability to
re-sort sub-populations along different parameters, and substantial
training necessary to own, operate and maintain these large,
expensive pieces of equipment. For at least these reasons, use of
flow cytometers has been restricted to large hospitals and
laboratories and the technology has not been accessible to smaller
entities.
[0008] A number of patents have been granted which are directed to
such MEMS-based particle sorting devices. For example, U.S. Pat.
No. 6,838,056 (the '056 patent) is directed to a MEMS-based cell
sorting device, U.S. Pat. No. 7,264,972 b1 (the '972 patent) is
directed to a micromechanical actuator for a MEMS-based cell
sorting device. U.S. Pat. No. 7,220,594 (the '594 patent) is
directed to optical structures fabricated with a MEMS cell sorting
apparatus, and U.S. Pat. No. 7,229,838 (the '838 patent) is
directed to an actuation mechanism for operating a MEMS-based
particle sorting system. Additionally, U.S. patent application Ser.
No. 13/374,899 (the '899 application) and Ser. No. 13/374,898 (the
'898 application) provide further details of other MEMS designs.
Each of these patents ('056, '972, '594 and '838) and patent
applications ('898 and '899) is hereby incorporated by
reference.
SUMMARY
[0009] One feature of the MEMS-based microfabricated particle
sorting system is that the fluid may be confined to small,
microfabricated channels formed in a semiconductor substrate
throughout the sorting process. The MEMS device may be a valve
which separates one or more target particles from other components
of a sample stream. The MEMS device may redirect the particle flow
from one channel into another channel, when a signal indicates that
a target particle is present. This signal may be photons from a
fluorescent tag which is affixed to the target particles and
excited by laser illumination in an interrogation region upstream
of the MEMS device. Thus, the MEMS device may be a particle or cell
sorter operating on a fluid sample confined to a microfabricated
fluidic channel, but using detection means similar to a FACS flow
cytometer. In particular, the '898 application discloses a
microfabricated fluidic valve wherein the inlet channel, sort
channel and waste channel all flow in a plane parallel to the
fabrication plane of the microfabricated fluidic valve.
[0010] A substantial improvement may be made over the prior art
devices by having at least one of the microfabricated fluidic
channels route the flow out of the plane of fabrication of the
microfabricated valve. A valve with such an architecture has the
advantage that the pressure resisting the valve movement is
minimized when the valve opens or closes, because the movable
member is not required to move a column of fluid out of the way.
Instead, the fluid containing the non-target particles may move
over and under the movable member to reach the waste channel.
Furthermore, the force-generating apparatus may be disposed closer
to the movable valve, resulting in higher forces and faster
actuation speeds. As a result, the time required to open or close
the valve may be much shorter than the prior art valve, improving
sorting speed and accuracy. The systems and methods disclosed here
may describe such a microfabricated particle sorting device with at
least one out-of-plane channel.
[0011] In the systems and methods disclosed here, a micromechanical
particle manipulation device may be formed on a surface of a
fabrication substrate, wherein the micromechanical particle
manipulation device may include a microfabricated, movable member
having a first diverting surface, wherein the movable member moves
from a first position to a second position in response to a force
applied to the movable member, wherein the motion is substantially
in a plane parallel to the surface, a sample inlet channel formed
in the substrate and through which a fluid flows, the fluid
including at least one target particle and non-target material,
wherein the flow in the sample inlet channel is substantially
parallel to the surface, and a plurality of output channels into
which the microfabricated member diverts the fluid, and wherein the
flow in at least one of the output channels is not parallel to the
plane, wherein at least one output channel is located directly
below at least a portion of the microfabricated diverter over at
least a portion of its motion. In one embodiment, The
micromechanical particle manipulation device of claim 1, wherein
the first diverting surface has a smoothly curved shape which is
substantially tangent to the direction of flow in the inlet channel
at one point on the shape and substantially tangent to the
direction of flow of a first output channel at a second point on
the shape, wherein the first diverting surface diverts flow from
the inlet channel into the first output channel when the movable
member is in the first position, and allows the flow into a second
output channel in the second position.
[0012] Finally, the systems and methods disclosed herein, because
they include microfabricated channels as well as the novel valve
design, may allow additional useful features to be implemented. For
example, the techniques may form a particle manipulation system
with cytometric capability, as described in co-pending U.S. patent
application Ser. No. 13/507,830 (Owl-Cytometer) filed Aug. 1, 2012
and assigned to the same assignee as the present application. This
patent application is incorporated by reference in its entirety.
The MEMS device describe here may be used to manipulate the
particles in the fluid stream enclosed in the microfabricated
channel, while a plurality of interrogation regions also exist
which may provide feedback on the manipulation. For example, in the
case of cell sorting, one laser interrogation region may exist
upstream of the MEMS device, and at least one additional laser
interrogation region may exist downstream of the MEMS device, to
confirm the results of the particle manipulation, that the correct
cell has been sorted.
[0013] The systems and methods disclosed here also enable the
construction of a single-input/double output sorting device,
wherein the flow from a single input channel can be diverted into
either of two sort output channels, or allowed to flow through to
the waste channel.
[0014] In another embodiment, the novel valve architecture may make
use of hydrodynamic particle focusing techniques, as taught by, for
example, "Single-layer planar on-chip flow cytometer using
microfluidic drifting based three-dimensional (3D) hydrodynamic
focusing," by Xaiole Mao, et al. (hereinafter "Mao," Journal of
Royal Society of Chemistry, Lab Chip, 2009, 9, 1583-1589). The
microfabricated architecture of the systems and methods disclosed
herein make them especially suitable for the techniques disclosed
in Mao, as described further below.
[0015] These and other features and advantages are described in, or
are apparent from, the following detailed description.
BRIEF DESCRIPTION OF THE DRAWINGS
[0016] Various exemplary details are described with reference to
the following figures, wherein:
[0017] FIG. 1 is a simplified plan view of a microfabricated
particle sorting system in the quiescent (no sort) position;
[0018] FIG. 2 is a simplified plan view of a microfabricated
particle sorting system in the actuated (sort) position;
[0019] FIG. 3a is a simplified plan view of a microfabricated
particle sorting system showing the field of view of the detector,
with the microfluidic valve in the quiescent (no sort) position;
FIG. 3b is a simplified illustration of a microfabricated particle
sorting system showing the field of view of the detector, with the
microfluidic valve in the actuated (sort) position;
[0020] FIG. 4a is a simplified cross sectional view of a
microfabricated particle sorting system in the actuated (sort)
position, showing the flow of the sample stream into the sort
channel which is in the same plane as the inlet channel; FIG. 4b is
a simplified cross sectional view of a microfabricated particle
sorting system in the quiescent (no sort) position, showing the
flow of the sample stream into the waste channel which is not in
the same plane as the inlet channel; FIG. 4c is a simplified cross
sectional view of a microfabricated particle sorting system in the
quiescent (no sort) position, showing the flow of the sample stream
into the waste channel which is not in the same plane as the inlet
channel, wherein the sample stream flows around the top and the
bottom of the diverter;
[0021] FIG. 5 is a simplified plan view of a microfabricated
particle sorting system in the quiescent (no sort) position,
showing the stationary magnetically permeable feature;
[0022] FIG. 6 is a plan view of the actuation mechanism for the
microfabricated particle sorting system, showing the functioning of
the external magnetic field in combination with the stationary
magnetically permeable feature;
[0023] FIG. 7 is a plan view of the actuation mechanism for the
microfabricated particle sorting system, showing the functioning of
the external magnetic field in combination with the stationary
magnetically permeable feature, in the actuated (sort)
position;
[0024] FIG. 8 is a simplified view of the microfabricated particle
sorting system, wherein multiple microfabricated particle sorters
are arranged to provide a serial sorting capability;
[0025] FIG. 9 is a plan view of a two-way microfabricated particle
sorting system, wherein the system has more than one sort
output;
[0026] FIG. 10 is a plan view of the a two-way microfabricated
particle sorting system, with more than one sort output, with the
two-way microfabricated particle sorting device in the actuated
position;
[0027] FIG. 11 is a plan view of the microfabricated particle
sorting system in combination with a hydrodynamic focusing
manifold;
[0028] FIG. 12 is a system-level illustration of a microfabricated
particle sorting system according to the present invention, showing
the placement of the various detection and control components;
and
[0029] FIG. 13 is a representation of a signal waveform from the
control system to the microfabricated particle sorting device,
showing the different in pulses used to control the motion of the
device.
DETAILED DESCRIPTION
[0030] The system described herein is a particle sorting system
which may make use of the microchannel architecture of a MEMS
particle manipulation system. More generally, the systems and
methods describe a particle manipulation system with an inlet
channel and a plurality of output channels, wherein at least one of
the plurality of output channels is disposed in a different plane
than the inlet channel. This architecture has some significant
advantages relative to the prior art.
[0031] In the figures discussed below, similar reference numbers
are intended to refer to similar structures, and the structures are
illustrated at various levels of detail to give a clear view of the
important features of this novel device. It should be understood
that these drawings do not necessarily depict the structures to
scale, and that directional designations such as "top," "bottom,"
"upper," "lower," "left" and "right" are arbitrary, as the device
may be constructed and operated in any particular orientation. In
particular, it should be understood that the designations "sort"
and "waste" are interchangeable, as they only refer to different
populations of particles, and which population is called the
"target" or "sort" population is arbitrary.
[0032] FIG. 1 is an plan view illustration of the novel
microfabricated fluidic device 10 in the quiescent (un-actuated)
position. The device 10 may include a microfabricated fluidic valve
or movable member 110 and a number of microfabricated fluidic
channels 120, 122 and 140. The fluidic valve 110 and
microfabricated fluidic channels 120, 122 and 140 may be formed in
a suitable substrate, such as a silicon substrate, using MEMS
lithographic fabrication techniques as described in greater detail
below. The fabrication substrate may have a fabrication plane in
which the device is formed and in which the movable member 110
moves.
[0033] A sample stream may be introduced to the microfabricated
fluidic valve 110 by a sample inlet channel 120. The sample stream
may contain a mixture of particles, including at least one desired,
target particle and a number of other undesired, nontarget
particles. The particles may be suspended in a fluid. For example,
the target particle may be a biological material such as a stem
cell, a cancer cell, a zygote, a protein, a T-cell, a bacteria, a
component of blood, a DNA fragment, for example, suspended in a
buffer fluid such as saline. The inlet channel 120 may be formed in
the same fabrication plane as the valve 110, such that the flow of
the fluid is substantially in that plane. The motion of the valve
110 is also within this fabrication plane. The decision to
sort/save or dispose/waste a given particle may be based on any
number of distinguishing signals. In one exemplary embodiment, the
decision is based on a fluorescence signal emitted by the particle,
based on a fluorescent tag affixed to the particle and excited by
an illuminating laser. Details as to this detection mechanism are
well known in the literature, and further discussed below with
respect to FIG. 12. However, other sorts of distinguishing signals
may be anticipated, including scattered light or side scattered
light which may be based on the morphology of a particle, or any
number of mechanical, chemical, electric or magnetic effects that
can identify a particle as being either a target particle, and thus
sorted or saved, or an nontarget particle and thus rejected or
otherwise disposed of.
[0034] With the valve 110 in the position shown, the input stream
passes unimpeded to an output orifice and channel 140 which is out
of the plane of the inlet channel 120, and thus out of the
fabrication plane of the device 10. That is, the flow is from the
inlet channel 120 to the output orifice 140, from which it flows
substantially vertically, and thus orthogonally to the inlet
channel 120. This output orifice 140 leads to an out-of-plane
channel that may be perpendicular to the plane of the paper showing
FIG. 1, and depicted in the cross sectional views of FIGS. 4a-4c.
More generally, the output channel 140 is not parallel to the plane
of the inlet channel 120 or sort channel 122, or the fabrication
plane of the movable member 110.
[0035] The output orifice 140 may be a hole formed in the
fabrication substrate, or in a covering substrate that is bonded to
the fabrication substrate. A relieved area above and below the
sorting valve or movable member 110 allows fluid to flow above and
below the movable member 110 to output orifice 140, and shown in
more detail in FIGS. 4a-4c. Further, the valve 110 may have a
curved diverting surface 112 which can redirect the flow of the
input stream into a sort output stream, as described next with
respect to FIG. 2. The contour of the orifice 140 may be such that
it overlaps some, but not all, of the inlet channel 120 and sort
channel 122. By having the contour 140 overlap the inlet channel,
and with relieved areas described above, a route exists for the
input stream to flow directly into the waste orifice 140 when the
movable member or valve 110 is in the un-actuated waste
position.
[0036] FIG. 2 is a plan view of the microfabricated device 10 in
the actuated position. In this position, the movable member or
valve 110 is deflected upward into the position shown in FIG. 2.
The diverting surface 112 is a sorting contour which redirects the
flow of the inlet channel 120 into the sort output channel 122. The
output channel 122 may lie in substantially the same plane as the
inlet channel 120, such that the flow within the sort channel 122
is also in substantially the same plane as the flow within the
inlet channel 120. There may be an angle .alpha. between the inlet
channel 120 and the sort channel 122, This angle may be any value
up to about 90 degrees. Actuation of movable member 110 may arise
from a force from force-generating apparatus 400, shown generically
in FIG. 2. In some embodiments, force-generating apparatus may be
an electromagnet, however, it should be understood that
force-generating apparatus may also be electrostatic,
piezoelectric, or some other means to exert a force on movable
member 110, causing it to move from a first position (FIG. 1) to a
second position (FIG. 2).
[0037] More generally, the micromechanical particle manipulation
device shown in FIGS. 1 and 2 may be formed on a surface of a
fabrication substrate, wherein the micromechanical particle
manipulation device may include a microfabricated, movable member
110 having a first diverting surface 112, wherein the movable
member 110 moves from a first position to a second position in
response to a force applied to the movable member, wherein the
motion is substantially in a plane parallel to the surface, a
sample inlet channel 120 formed in the substrate and through which
a fluid flows, the fluid including one or more target particles and
non-target material, wherein the flow in the sample inlet channel
is substantially parallel to the surface, and a plurality of output
channels 122, 140 into which the microfabricated member diverts the
fluid, and wherein the flow in at least one of the output channels
140 is not parallel to the plane, and wherein at least one output
channel 140 is located directly below at least a portion of the
movable member 110 over at least a portion of its motion.
[0038] In one embodiment, the diverting surface 112 may be nearly
tangent to the input flow direction as well as the sort output flow
direction, and the slope may vary smoothly between these tangent
lines. In this embodiment, the moving mass of the stream has a
momentum which is smoothly shifted from the input direction to the
output direction, and thus if the target particles are biological
cells, a minimum of force is delivered to the particles. As shown
in FIGS. 1 and 2, the micromechanical particle manipulation device
10 has a first diverting surface 112 with a smoothly curved shape,
wherein the surface which is substantially tangent to the direction
of flow in the sample inlet channel at one point on the shape and
substantially tangent to the direction of flow of a first output
channel at a second point on the shape, wherein the first diverting
surface diverts flow from the sample inlet channel into the first
output channel when the movable member 110 is in the first
position, and allows the flow into a second output channel in the
second position.
[0039] In other embodiments, the overall shape of the diverter 112
may be circular, triangular, trapezoidal, parabolic, or v-shaped
for example, but the diverter serves in all cases to direct the
flow from the inlet channel to another channel.
[0040] It should be understood that although channel 122 is
referred to as the "sort channel" and orifice 140 is referred to as
the "waste orifice", these terms can be interchanged such that the
sort stream is directed into the waste orifice 140 and the waste
stream is directed into channel 122, without any loss of
generality. Similarly, the "inlet channel" 120 and "sort channel"
122 may be reversed. The terms used to designate the three channels
are arbitrary, but the inlet stream may be diverted by the valve
110 into either of two separate directions, at least one of which
does not lie in the same plane as the other two. The term
"substantially" when used in reference to an angular direction,
i.e. substantially tangent or substantially vertical, should be
understood to mean within 15 degrees of the referenced direction.
For example, "substantially orthogonal" to a line should be
understood to mean from about 75 degrees to about 105 degrees from
the line.
[0041] FIGS. 3a and 3b illustrate an embodiment wherein the angle
.alpha. between the inlet channel 120 and the sort channel 122 is
approximately zero degrees. Accordingly, the sort channel 122 is
essentially antiparallel to the inlet channel 120, such that the
flow is from right to left in the inlet channel 120. With valve 110
in the un-actuated, quiescent position shown in FIG. 3a, the inlet
stream flows straight to the waste orifice 140 and vertically out
of the device 10.
[0042] In FIG. 3b, the valve 110 is in the actuated, sort position.
In this position, the flow is turned around by the diverting
surface 112 of the valve 110 and into the antiparallel sort channel
122. This configuration may have an advantage in that the field of
view of the detector 150 covers both the inlet channel 120 and the
sort channel 122. Thus a single set of detection optics may be used
to detect the passage of a target particle through the respective
channels. It may also be advantageous to minimize the distance
between the detection region and the valve 110, in order to
minimize the timing uncertainty in the opening and closing of the
valve.
[0043] The movable member or valve 110 may be attached to the
substrate with a flexible spring 114. The spring may be a narrow
isthmus of substrate material. In the example set forth above, the
substrate material may be single crystal silicon, which is known
for its outstanding mechanical properties, such as its strength,
low residual stress and resistance to creep. With proper doping,
the material can also be made to be sufficiently conductive so as
to avoid charge build up on any portion of the device, which might
otherwise interfere with its movement. The spring may have a
serpentine shape as shown, having a width of about 1 micron to
about 10 microns and a spring constant of between about 10 N/m and
100 N/m, and preferably about 40 N/m
[0044] FIGS. 4a, 4b, 4c are cross sectional views illustrating the
operation of the out-of-plane waste channel 140. FIG. 4c is
slightly enlarged relative to FIGS. 4a and 4b, in order to show
detail of the flow around the movable member 110 and into the waste
channel 142 through waste orifice 140. In this embodiment, the
waste channel 142 is vertical, substantially orthogonal to the
inlet stream 120 and sort stream 122. It should be understood that
other embodiments are possible other than orthogonal, but in any
event, the flow into waste channel 142 is out of the plane of the
flow in the inlet channel 120 and/or sort channel 122. As shown in
FIG. 4a, with the valve in the sort, actuated position, the inlet
stream and target particle may flow into the sort stream, which in
FIG. 4a is out of the paper, and the waste orifice 140 is largely,
though not completely, blocked by the movable member 110. The area
144 (shown more clearly in FIG. 4c) on top of the valve or movable
member 110 may be relieved to provide clearance for this flow.
[0045] When the valve or movable member 110 is un-actuated as in
FIG. 4b, the flow of the inlet channel 120 may flow directly into
the waste channel 142 by going over, around or by the movable
member or valve 110. The area 144 on top of the valve or movable
member 110 may be relieved to provide clearance for this flow. The
relieved area 144 is shown in greater detail in the enlarged FIG.
4c. Thus when the movable member is un-actuated, the flow will be
sent directly to the waste channel. When the movable member is
actuated, most of the fluid will be directed to the sort channel,
although liquid may still flow over and under the movable
member.
[0046] Thus, the purpose of providing flow both under and over the
movable member 110 is to reduce the fluid pressure produced by the
actuator motion in the region behind the valve or movable member
110. In other words, the purpose is to provide as short a path as
possible between the high pressure region in front of the valve 110
and the low pressure region behind the valve. This allows the valve
to operate with little pressure resisting its motion. As a result,
the movable valve 110 shown in FIGS. 1-4c may be substantially
faster than valves which have all channels disposed in the same
plane.
[0047] Another advantage of the vertical waste channel 142 is that
by positioning it directly underneath a stationary permeable
feature 130 and movable permeable feature 116, the magnetic gap
between the permeable features 116 and 130 can be narrower than if
the fluidic channel went between them. The narrower gap enables
higher forces and thus faster actuation compared to prior art
designs. A description of the magnetic components and the magnetic
actuation mechanism will be given next, and the advantages of the
out-of-plane channel architecture will be apparent.
[0048] FIG. 5 is a plan view of another exemplary embodiment of
device 100 of the device 10, showing the disposition of a
stationary permeable feature 130 and further detail of the movable
member 110. In this embodiment, the movable member 110 may include
the diverting surface 112, the flexible hinge or spring 114, and a
separate area 116 circumscribed but inside the line corresponding
to movable member 110. This area 116 may be inlaid with a permeable
magnetic material such as nickel-iron permalloy, and may function
as described further below.
[0049] A magnetically permeable material should be understood to
mean any material which is capable of supporting the formation of a
magnetic field within itself. In other words, the permeability of a
material is the degree of magnetization that the material obtains
in response to an applied magnetic field.
[0050] The terms "permeable material" or "material with high
magnetic permeability" as used herein should be understood to be a
material with a permeability which is large compared to the
permeability of air or vacuum. That is, a permeable material or
material with high magnetic permeability is a material with a
relative permeability (compared to air or vacuum) of at least about
100, that is, 100 times the permeability of air or vacuum which is
about 1.26.times.10.sup.-6 Hm.sup.-1. There are many examples of
permeable materials, including chromium (Cr), cobalt (Co), nickel
(Ni) and iron (Fe) alloys. One popular permeable material is known
as Permalloy, which has a composition of between about 60% and
about 90% Ni and 40% and 10% iron. The most common composition is
80% Ni and 20% Fe, which has a relative permeability of about
8,000.
[0051] It is well known from magnetostatics that permeable
materials are drawn into areas wherein the lines of magnetic flux
are concentrated, in order to lower the reluctance of the path
provided by the permeable material to the flux. Accordingly, a
gradient in the magnetic field urges the motion of the movable
member 110 because of the presence of inlaid permeable material
116, towards areas having a high concentration of magnetic flux.
That is, the movable member 110 with inlaid permeable material 116
will be drawn in the direction of positive gradient in magnetic
flux.
[0052] An external source of magnetic field lines of flux may be
provided outside the device 100, as shown in FIG. 6. This source
may be an electromagnet 500. The electromagnet 500 may include a
permeable core 512 around which a conductor 514 is wound. The wound
conductor or coil 514 and core 512 generate a magnetic field which
exits the pole of the magnet, diverges, and returns to the opposite
pole, as is well known from electromagnetism. Accordingly, the
movable member 110 is generally drawn toward the pole of the
electromagnet 500 as shown in FIG. 7.
[0053] However, the performance of the device 100 can be improved
by the use of a stationary permeable feature 130. The term
"stationary feature" should be understood to mean a feature which
is affixed to the substrate and does not move relative to the
substrate, unlike movable member or valve 110. A stationary
permeable feature 130 may be shaped to collect these diverging
lines of flux and refocus them in an area directly adjacent to the
movable member 110 with inlaid permeable material. The stationary
permeable feature may have an expansive region 132 with a narrower
throat 134. The lines of flux are collected in the expansive region
132 and focused into and out of the narrow throat area 134.
Accordingly, the density of flux lines in the throat area 134 is
substantially higher than it would be in the absence of the
stationary permeable feature 130. Thus, use of the stationary
permeable feature 130 though optional, allows a higher force,
faster actuation, and reduces the need for the electromagnet 500 to
be in close proximity to the device 10. From the narrow throat area
134, the field lines exit the permeable material and return to the
opposite magnetic pole of the external source 500. But because of
the high concentration of field lines in throat area 134, the
permeable material 116 inlaid into movable member 110 may be drawn
toward the stationary permeable feature 130, bringing the rest of
movable member with it.
[0054] When the electromagnet is quiescent, and no current is being
supplied to coil 514, the restoring force of spring 114 causes the
movable member 110 to be in the "closed" or "waste" position. In
this position, the inlet stream passes unimpeded through the device
100 to the waste channel 140. This position is shown in FIG. 5.
When the electromagnet 500 is activated, and a current is applied
through coil 514, a magnetic field arises in the core 512 and exits
the pole of the core 512. These lines of flux are collected and
focused by the stationary permeable feature 130 and focused in the
region directly adjacent to the throat 134. As mentioned
previously, the permeable portion 116 of the movable member 110 is
drawn toward the throat 134, thus moving the movable member 110 and
diverting surface 112 such that the inlet stream in inlet channel
120 is redirected to the output or sort channel 122. This position
is shown in FIG. 7.
[0055] Permalloy may be used to create the permeable features 116
and 130, although it should be understood that other permeable
materials may also be used. Permalloy is a well known material that
lends itself to MEMS lithographic fabrication techniques. A method
for making the permeable features 116 and 130 is described further
below.
[0056] As mentioned previously, having the waste channel 140 and
142 directly beneath the movable member or valve 110 allows the
movable permeable feature 116 to be disposed much closer to the
stationary permeable feature 130. If instead the waste channel were
in the same plane, this gap would have to be at least large enough
to accommodate the waste channel, along with associated tolerances.
As a result, actuation forces are higher and valve opening and
closing times are much shorter. This in turn corresponds to either
faster sorting or better sorting accuracy, or both.
[0057] With the use of the electromagnetic actuation technique
described above, actuation times on the order of 10 microseconds
can be realized. Accordingly, the particle sorting device is
capable of sorting particles at rates in excess of 50 kHz or
higher, assuming 10 microseconds required to pull the actuator in,
and 10 microseconds required to return it to the as-manufactured
position.
[0058] For any particle sorting mechanism however, there is an
inherent trade-off between sort purity and sort speed. One can only
increase the fluid speed to a certain point, after which one runs
into physical limitations of the sorter, for example, when the
valve speed is such that there is insufficient time to open the
valve or flap when a cell is detected. Beyond that limitation, the
most obvious way to achieve more events per second is to increase
the cell density. But, with increased cell density, the incidence
of sort conflicts, wherein both a desired and an undesired cell are
collected, also increases.
[0059] In order to overcome this limitation, a cell sample may
theoretically be processed multiple times in a sequential sort
strategy--initially a very rapid, crude sort followed by a--slower,
high precision sort. This is generally not a practical option with
a traditional FACS system as a result of massive cell dilution
(from sheath fluid), slow processing speeds and unacceptable cell
damage resulting from multiple passes through the high pressure
electrostatic sorting mechanism. A single pass through a flow
cytometer is exceptionally violent, with 10 msec velocities,
explosive decompression from 60 psi to 0 psi. Cells are unlikely to
survive such treatment on multiple passes without significant loss
of viability. Even if one is willing to accept the dilution, manual
processing and cell death, the yield losses on a FACS would be
overwhelming. Also, the time constant per cycle for processing,
cleaning, sterilization and certification is untenable and the
sterility of the sample is completely compromised. As a result,
this sequential sorting is not a practical approach for FACS-based
clinical cell sorting.
[0060] In contrast, for the microfabricated particle sorting system
described above, using the microfluidic channel architecture, a
multi-stage, "sequential" sort may be performed in a
straightforward way as described below. A plurality of particle
manipulation operations may take place using a plurality of MEMS
sorting devices 10 or 100. The sorting devices may be on separate
MEMS chips and enclosed in a disposable cartridge, or multiple
valves may be formed on a single substrate using MEMS fabrication
techniques. In one embodiment, the plurality of MEMS sorting chips
are separated by some extent, such that by laterally shifting the
device, the additional MEMS chips may become operational. This
embodiment is described further below, and illustrated in FIG. 8.
More broadly, the sorting device system may include a secondary
manipulation device or sorting stage 200 downstream of the first
manipulation device or sorting stage 100. Sorting stage 100
connotes a stage using either device 10 or device 100 for example,
as illustrated in FIGS. 1 and 5, respectively.
[0061] The first sorting stage 100 and second sorting stage 200 are
both preceded by a laser interrogation region 170 and 270,
respectively. In this region, a laser is used to irradiate the
particles in the sample stream. Those particles bearing a
fluorescent tag may fluoresce as a result of the laser irradiation.
This fluorescence signal is detected and is indicative of the
presence of a target particle in the sample stream. Upon detection
of the target particle, a signal is sent to the controller
controlling the electromagnet 500, energizing the electromagnet and
thus opening the movable member or valve 110. The target particle
is thus directed into the sort channel 122. This functionality is
described in further detail below with respect to the full particle
sorting system shown in FIG. 12. The sorting stages 100 and 200 may
also be accompanied by a third laser interrogation region 280
downstream of the last sorting stage 200. This interrogation may be
performed to evaluate the accuracy of the sort, or in order to
adjust various sorting parameters. Although only two sorting
operations arranged sequentially are shown in FIG. 12, it should be
understood that this basic concept may be extended to any number of
additional sorting stages, and that the stages may be arranged in a
parallel configuration, instead of, or in addition to, the serial
configuration.
[0062] Accordingly, a first sort may be run rapidly through a first
sorting stage 100, to enrich target cells with negligible yield
losses. The output of the first sorting stage 100 may flow into
either a waste channel 140 or a sort channel 122, based on the
output of a discriminator or detector located in region 170. If the
stream flows to the sort channel 122, it then flows on to a second
sorting stage 200, which may have its own associated detection area
270. Similarly to sort stage 100, the flow may be direct to a waste
channel 240 or a sort channel 222. Using this approach, the sample
remains sterile and gently handled through the entire sequential
sorting process. It should be understood that although difficult to
depict in a two dimensional drawing, the waste channel 140 and 240
may lie in a different plane relative to the inlet channel 120, and
sort channels 122 and 222. In FIG. 8 waste channels 140 and 240 are
depicted flowing into the paper.
[0063] In another embodiment, using the architecture shown in FIG.
1, 3, or 5, a dual output, dual position particle manipulation
device may also be envisioned. Such a device is shown in FIG. 9.
FIG. 9 shows a dual output device 800 wherein a single inlet
channel 820 can feed either of two separate sort output channels
822 and 824, depending on the position of movable member 810. Dual
output device 800 may have two permeable areas 816 and 818, which
may be drawn toward either of two stationary permeable features 830
and 850, respectively. For example, if a source of external
magnetic flux such as electromagnet 500 is positioned near
stationary permeable feature 830, the flux emitted from
electromagnet 500 is concentrated by stationary permeable feature
830 and movable permeable feature 816 is drawn toward it. The
situation is as depicted in FIG. 10. When the movable feature
rotates clockwise, opening sort channel 822 to the flow from inlet
channel 820 by diverting surface 842. When another external magnet
(not shown) is energized above device 800 and upper stationary
permeable feature 850, the movable member 810 rotates
counterclockwise, directing the flow in inlet channel 820 into the
upper sort channel 824 by sort diverting surface 812. The waste
channel orifice 840 may be enlarged compared to 140, such that it
is disposed directly under at least a portion of movable member
810, but does not interfere with the motion of sort diverting
surfaces 812 or 842.
[0064] Although the embodiments shown in FIGS. 1-11 are described
with respect to an electromagnetic actuation mechanism, it should
be understood that other actuation forces may be used instead. For
example, if permeable features 116 and 130 are made from an
electrically conductive rather than permeable magnetic material, a
voltage potential may be placed across elements 116 and 130,
producing an electrostatic force to move the movable member 110.
Piezoelectric forces may also be used.
[0065] Because of the microfabricated architecture of particle
manipulation device 10 and 100, it lends itself to techniques that
can make use of such an enclosed, well defined architecture. One
such technique is illustrated in FIG. 11, wherein the
microfabricated particle manipulation device may have at least one
additional channel that provides a sheath fluid to the sample
stream and also a focusing element coupled to the inlet channel.
The sheath fluid may be used to adjust the concentration or
positioning of the target particles within the inlet channel. The
focusing element may be configured to urge the target particles
into a particular portion of the sample inlet channel, as described
further below. The focusing element may be disposed in
substantially the same plane as the movable member 110, and may be
formed in the same substrate surface as the movable member 110 and
inlet channel 120.
[0066] FIG. 11 depicts a microfabricated fluidic manifold 300 which
may be used to focus the particles in a certain area within the
fluid stream. Techniques for designing such a manifold may be found
in, for example, "Single-layer planar on-chip flow cytometer using
microfluidic drifting based three-dimensional (3D) hydrodynamic
focusing," by Xiaole Mao et cl, Journal of Royal Society of
Chemistry, Lab Chip, 2009, 9, 1583-1589. The manifold may include a
sample inlet 310 and sheath fluid channel 320. As the name
suggests, the sheath channel adds a sheath fluid to the sample
stream, which is a buffering fluid which tends to dilute the flow
of particles in the stream and locate them in a particular portion
of the stream. The combined fluid then flows around a focusing
element coupled to the inlet channel 120, here a z-focusing channel
330, which tends to herd the particles into a particular plane
within the flow. This plane is substantially in the plane of the
paper of FIG. 11. The combined fluid then passes another
intersection point, a "y-intersection point" 350, which introduces
additional sheath fluid above and below the plane of particles. At
the y-intersection point 350, two flows may join the z-focus
channel 330 from substantially antiparallel directions, and
orthogonal to the z-focus channel 330. This intersection may
compress the plane of particles into a single point, substantially
in the center of the stream. Accordingly, at the y-intersection
point 350 the target particles may be compressed from a plane to a
stream line near the center of the z-focus channel 330 and sample
inlet channel 120. Focusing the particles into a certain volume
tends to decrease the uncertainly in their location, and thus the
uncertainty in the timing of the opening and closing of the movable
member or valve 110. Such hydrodynamic focusing may therefore
improve the speed and/or accuracy of the sorting operation.
[0067] In one exemplary embodiment of the microfabricated particle
manipulation device 10 or 100 with hydrodynamic focusing
illustrated in FIG. 11, the angular sweep of z-bend 330 is a curved
arc of about 180 degrees. That is, the approximate angular sweep
between the junction of the sheath inlet with the cell inlet and
the y-intersection point 350, may be about 180 degrees. Generally,
the radius of curvature of the z-bend 330 may be at least about 100
microns and less than about 500 microns, and the characteristic
dimension, that is the width, of the channels is typically about 50
microns to provide the focusing effect. In one embodiment, the
radius of curvature of the channel may be about 250 microns, and
the channel widths, or characteristic dimensions, for the sample
inlet channel 120 and z-bend channel are on the order of about 50
microns. These characteristic dimensions may provide a curvature
sufficient to focus the particles, such that they tend to be
confined to the plane of the paper upon exit from the z-focus
channel 330 at y-intersection point 350. This plane is then
compressed to a point in the channel at the y-intersection point
350.
[0068] The microfabricated particle manipulation device 10 or 100
may be used in a particle sorting system 1000 enclosed in a housing
containing the components shown in FIG. 12. The MEMS particle
manipulation devices 10, 100 or 800 may be enclosed in a plastic,
disposable cartridge which is inserted into the system 1000. The
insertion area may be a movable stage with mechanisms available for
fine positioning of the particle manipulation device 10, 100 or 800
and associated microfluidic channels against one or more data,
which orient and position the detection region and particle
manipulation device 10, 100 or 800 with respect to the collection
optics 1100. If finer positioning is required, the inlet stage may
also be a translation stage, which adjusts the positioning based on
observation of the location of the movable member 110 relative to a
datum.
[0069] It should be understood that although FIG. 12 shows a
particle sorting system 1000 which uses a plurality of laser
sources 1400 and 1410, only a single laser may be required
depending on the application. For the plurality of lasers shown in
FIG. 12, one of the laser sources 1410 may be used with an
associated set of parallel optics (not shown in FIG. 12) to
illuminate the at least one additional laser interrogation region
170 and/or 270. This setup may be somewhat more complicated and
expensive to arrange than a single laser system, but may have
advantages in that the optical and detection paths may be separated
for the different laser interrogation regions. For this embodiment,
it may not be necessary to alter the trajectory, spectral content,
timing or duration of the laser 1410 light. Although not shown
explicitly in FIG. 12, it should be understood that the detection
path for additional laser(s) 1410 may also be separate from the
detection path for laser 1400. Accordingly, some embodiments of the
particle sorting system may include a plurality of laser sources
and a plurality of optical detection paths, whereas other
embodiments may only use a single laser source 1400 and collection
optics 1100. In the embodiment described here, a plurality of
excitation lasers uses a common optical path, and the optical
signals are separated electronically by the system shown in FIG.
12.
[0070] The embodiment shown in FIG. 12 is based on a FACS-type
detection mechanism, wherein one or more lasers 1400, 1410 excites
one or more fluorescent tags affixed to the target particles. The
laser excitation may take place in multiple interrogation regions,
such as regions 170, 270 and 280. The fluorescence emitted as a
result are detected and the signal is fed to a computer 1900. The
computer then generates a control signal that controls the
electromagnet 500, or multiple electromagnets if multiple sorters
are used such as in FIG. 8. It should be understood that other
detection mechanisms may be used instead, including electrical,
mechanical, chemical, or other effects that can distinguish target
particles from non-target particles.
[0071] Accordingly, the MEMS particle sorting system 1000 shown in
FIG. 12 may include a number of elements that may be helpful in
implementing the additional interrogation regions 170 and 270, or
more. First, an optical manipulating means 1600 may alter the
trajectory, spectral content, timing or duration of the laser
radiation from laser 1400 to the second or third interrogation
spots. Examples of items that may be included in optical
manipulating means 1600 are a birefringent crystal, spinning prism,
mirror, saturable absorber, acousto-optic modulator, harmonic
crystal, Q-switch, for example. More generally, optical
manipulating means 1600 may include one or more items that alter
laser frequency, amplitude, timing or trajectory along one branch
of the optical path to an additional interrogation region.
[0072] For example, optical manipulating means 1600 may include a
beamsplitter and/or acousto-optic modulator. The beam splitter may
separate a portion of the incoming laser beam into a secondary
branch or arm, where this secondary branch or arm passes through
the modulator which modulates the amplitude of the secondary beam
at a high frequency. The modulation frequency may be, for example,
about 2 MHz or higher. The light impinging on the first laser
interrogation region 101 may, in contrast, be continuous wave
(unmodulated). The secondary branch or arm is then directed to the
additional laser interrogation region 170 or 270. This excitation
will then produce a corresponding fluorescent pattern from an
appropriately tagged cell.
[0073] This modulated fluorescent pattern may then be picked up by
the detection optics 1600, which may recombine the detected
fluorescence from interrogation region 170 and/or 270 with
fluorescence from laser interrogation region 170. The combined
radiation may then impinge on the one or more detectors 1300.
[0074] An additional optical component 1700 may also alter the
frequency, amplitude, timing or trajectory of the second beam path,
however, it may perform this operation upstream (on the detector
side) of the collection optics 1100 rather than downstream (on the
sample side) of it, as does optical component 1600.
[0075] The output of detectors 1300 may be analyzed to separate the
content corresponding to laser interrogation region 280 from the
content corresponding to laser interrogation region 170 or 270.
This may be accomplished by applying some electronic distinguishing
means to the signals from detectors 1300. The details of electronic
distinguishing means 1800 may depend on the choice for optical
manipulation means 1600. For example, the distinguishing means 1800
may include a high pass stage and a low pass stage that is
consistent with a photoacoustic modulator that was included in
optical manipulating means 1600. Or electronic distinguishing means
1800 may include a filter (high pass and/or low pass) and/or an
envelope detector, for example.
[0076] Therefore, depending on the choice of optical manipulating
means 1600, the unfiltered signal output from detectors 1300 may
include a continuous wave, low frequency portion and a modulated,
high frequency portion. After filtering through the high pass
filter stage, the signal may have substantially only the high
frequency portion, and after the low pass stage, only the low
frequency portion. These signals may then be easily separated in
the logic circuits of computer 1900. Alternatively, the high pass
filter may be an envelope detector, which puts out a signal
corresponding to the envelop of the amplitudes of the high
frequency pulses.
[0077] Other sorts of components may be included in electronic
distinguishing means 1800 to separate the signals. These components
may include, for example, a signal filter, mixer, phase locked
loop, multiplexer, trigger, or any other similar device that can
separate or distinguish the signals. Component 1800 may also
include the high pass and/or low pass electronic filter or the
envelope detector described previously. The two sets of signals
from the electronic distinguishing means 1800 may be handled
differently by the logic circuits 1900 in order to separate the
signals.
[0078] Thus, a MEMS particle manipulation system may be used in
conjunction with one or more additional downstream laser
interrogation regions, wherein the additional laser interrogation
regions are used to confirm the effectiveness or accuracy of a
manipulation stage in manipulating a stream of particles. The
downstream evaluation from laser interrogation region 280 past the
sorting stage 100 and 200 may allow the operator to measure one
event number (e.g. the captured event rate post-sort) divided by
another event number (e.g. the initial event rate pre-sort) for
individual particle types, and to feedback to adjust initial
interrogation parameters (e.g. such as x, y, z position and also
"open window" length in time) based on this ratio. This method may
be used to optimize the yield or accuracy of the system 1000.
Alternatively, the operator could measure the event rate post-sort
of target cells, divided by total event rate post-sort feedback to
adjust initial laser interrogation parameters such as x, y, z
position and also "open window" length in time, in order to
optimize the purity of the sorting system 1000. These sorting
parameters may be adjusted by changing control signal 2000 which is
sent by computer 1900 to electromagnet 500, or by changing the
optical detection parameters or by changing the laser control
signals, as shown in FIG. 12.
[0079] One example of how the system depicted in FIG. 12 may be
used to adjust the sorting parameters, is via the control signal
waveform 2000 delivered to the electromagnet 500. This waveform
2000 may be fine-tuned to adjust the sorting performance of the
valve or movable member 110 or 810, and may be produced by logic
circuits 1900. FIG. 13 depicts a control signal waveform 2000 with
additional features that may be used to control the motion of
movable member 110 or 810. This control signal waveform 2000 may be
generated by computer 1900, and thus may be made essentially
arbitrarily complex. The control signal waveform 2000 may be either
a voltage waveform or a current waveform. The control signal
waveform 2000 may be applied to coil 510 of electromagnet 500, for
example, to drive current through the coil to produce the actuating
magnetic field. The control signal 2000 may include an acceleration
phase 2110 which has a substantially larger magnitude than the
remainder of the control signal waveform 2000, and lasts for tens
of microseconds.
[0080] The larger magnitude of the current in the acceleration
phase may be used to overcome the back electromotive force produced
in the coils by the moving magnets. It may also produce a higher
force, which may be needed to break the movable member 110, 810
from its rest position and overcome any stiction forces that may be
hindering motion. After this initial acceleration phase, the
control signal may have a maintenance phase during which the
current is essentially constant and lasts for tens of microseconds.
During this period, the movable member 110 or 810 travels from its
closed position in FIG. 1, 5 or 9 to actuated positions shown in
FIG. 2, 7 or 10. Although the current may be constant during this
period, the force on the movable member may be variable, a function
of the closing distance between movable permeable feature 116, 816
and 840 and the respective stationary permeable features 130, 840
and 850. Reversing the polarity of the control signal as shown in
2130 reverses the direction of the magnetic field, and demagnetizes
the permeable portions. After the reversal period 2130, a quiescent
period 2140 lasting several microseconds may follow, during which
there is no magnetic field produced, and the spring force of spring
element 114 or 814 on movable member 110 or 810 may return the
movable member to its un-actuated state. This may be in the waste
or reject position. After a period when the actuator is closing and
about to reach the as-manufactured position, a short "braking"
pulse 2150 may slow the velocity of the movable member. This may
avoid an undesirable bounce off the hard stop, which may otherwise
allow a non-target particle to enter the sort channel 122. Or if
there is no hard stop, this may allow the fastest return to the
un-actuated position.
[0081] Using the downstream confirmation of the sort channel
contents as described above with respect to FIG. 12, any of the
adjustable parameters of the current profile shown in FIG. 13, such
as amplitude and duration of the acceleration phase, amplitude and
duration of the opening phase, duration of the quiescent phase, or
amplitude and duration of the braking phase, may be adjusted to
improve the sort performance of the system.
[0082] The description now turns to the fabrication of the devices
shown in FIGS. 1-11. Fabrication may begin with the inlaid
permeable features 116 and 130 formed in a first substrate. The
substrate may be a single crystal silicon substrate, for example.
To form these structures, depressions may be formed in these areas
of the substrate surface by etching. First, photoresist may be
deposited over the substrate surface and removed over the areas
corresponding to 116 and 130. Then, the trenches may be formed by,
for example, etching the substrate in potassium hydroxide (KOH) to
form a suitable depression. A seed layer may be deposited
conformally over the first substrate surface and patterned to
provide the seed layer for plating NiFe into the trenches. The seed
layer may be, for example, Ti/W or Cr/Au may then be deposited by
sputtering, CVD or plasma deposition. This layer may be covered
with photoresist and patterned according to the desired shape of
the areas 116 and 130. Unwanted areas of photoresist and seed layer
may then be removed by chemical etching. The permeable features may
then be deposited over the patterned seed layer by sputtering,
plasma deposition or electrochemical plating. It is known that
permalloy (80% Ni and 20% Fe), for example, can readily be
deposited by electroplating.
[0083] Alternatively, a liftoff method may be used to deposit a
sheet of permeable material, most of which is then lifted off areas
other than 116 and 130. Further details into the lithographic
formation of inlaid, magnetically permeable materials may be found
in, for example, U.S. Pat. No. 7,229,838. U.S. Pat. No. 7,229,838
is hereby incorporated by reference in its entirety. The substrate
may then be planarized by chemical mechanical polishing (CMP),
leaving a flat surface for the later bonding of a cover plate.
[0084] Having made the permeable features 116 and 130, the movable
member or valve 110 and 810 may be formed. The surface may again be
covered with photoresist and patterned to protect the inlaid
permeable features 116 and 130. The inlet channel 120 and output
channels 122 and relieved area 144 may be formed simultaneously
with the movable member 110 and 810. With movable member 110, 810
and other areas whose topography is to be preserved covered with
photoresist, the features 110, 810, 120, 122 and 144 may be formed
by deep reactive ion etching (DRIE) for example.
[0085] To form the fluidic channels, a cover plate may be bonded to
the surface of the substrate which was previously planarized for
this purpose. The cover plate may be optically transparent to allow
laser light to be applied to the particles in the fluid stream
flowing in the inlet channel 120, and for fluorescence emitted by
the fluorescent tags affixed to the particles to be detected by the
optical detection system described above. A hole formed in this
transparent material may form the waste channel 142. Alternatively,
a waste channel 142 may be formed in a second substrate, such as a
second silicon substrate, and bonded to the surface of the first
substrate. Alternatively, output channel 142 may be formed on the
opposite surface of the first substrate using a
silicon-on-insulator (SOI) substrate, with waste channel 142 and
orifice 140 formed in the handle layer and dielectric layer of the
SOI substrate, and the movable feature formed in the device
layer.
[0086] Additional details for carrying out this process outlined
above are well known to those skilled in the art, or readily found
in numerous lithographic processing references.
[0087] While various details have been described in conjunction
with the exemplary implementations outlined above, various
alternatives, modifications, variations, improvements, and/or
substantial equivalents, whether known or that are or may be
presently unforeseen, may become apparent upon reviewing the
foregoing disclosure. Accordingly, the exemplary implementations
set forth above, are intended to be illustrative, not limiting.
* * * * *