# US 11,077,438 B2 · Apparatuses, systems and methods for imaging micro-objects

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- Number: US 11,077,438 B2
- Document ID: US11077438B2
- Status: Granted
- Kind: B2
- Application number: 16/415,894
- Filed: 2019-05-17
- Published: 2019-12-12
- Granted: 2021-08-03
- Earliest claimed: 2016-12-01
- Assignees: Berkeley Lights, Inc.
- Applicants: Berkeley Lights, Inc.
- Inventors: Paul M. Lundquist; Paul M. Lebel; Phillip Ronald Thomas Jess
- Web page: https://patentgrep.com/patent/US11077438B2

## CPC

- B01L 3/502715: {characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces}
- B01L 3/00: Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
- B01L 3/502792: {for moving individual droplets on a plate, e.g. by locally altering surface tension}
- B01L 7/00: Heating or cooling apparatus (autoclaves B01J3/04); Heat insulating devices
- G01J 3/0208: {using focussing or collimating elements, e.g. lenses or mirrors; performing aberration correction}
- G01J 3/0213: {using attenuators}
- G01J 3/0229: {using masks, aperture plates, spatial light modulators or spatial filters, e.g. reflective filters}
- G01J 3/10: Arrangements of light sources specially adapted for spectrometry or colorimetry
- G01J 3/2803: {using photoelectric array detector}
- G01J 3/4406: {Fluorescence spectrometry}
- G01N 1/40: Concentrating samples
- G01N 21/31: Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry {(G01N21/72 takes precedence)}
- G01N 21/6458: {Fluorescence microscopy (fluorescence microscopes per se G02B21/0076 and G02B21/16)}
- G02B 21/02: Objectives
- G02B 21/06: Means for illuminating specimens
- G02B 21/16: adapted for ultraviolet illumination {; Fluorescence microscopes (G02B21/0076 takes precedence)}
- G02B 21/361: {Optical details, e.g. image relay to the camera or image sensor (G02B21/364 takes precedence; illumination details G02B21/06 and subgroups)}
- B01L 2200/147: Employing temperature sensors
- B01L 2300/046: Function or devices integrated in the closure
- B01L 2300/0645: Electrodes
- B01L 2300/0654: Lenses; Optical fibres
- B01L 2300/0663: Whole sensors
- B01L 2300/0883: Serpentine channels
- B01L 2300/0887: Laminated structure
- B01L 2300/1822: using Peltier elements
- B01L 2400/0424: Dielectrophoretic forces
- G01J 2003/104: {Monochromatic plural sources}
- B01L: CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- G01J: MEASUREMENT OF INTENSITY, VELOCITY, SPECTRAL CONTENT, POLARISATION, PHASE OR PULSE CHARACTERISTICS OF INFRARED, VISIBLE OR ULTRAVIOLET LIGHT; COLORIMETRY; RADIATION PYROMETRY (light sources F21, H01J, H01K, H05B; investigating properties of materials by optical means G01N)
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)
- G02B: OPTICAL ELEMENTS, SYSTEMS OR APPARATUS

## Abstract

The present disclosure relates to an optical apparatus for imaging and/or manipulating micro-objects in a microfluidic device, such as a light-actuated microfluidic (LAMF) device, and related systems and methods. The optical apparatus can comprise a structured light modulator, a first and a second tube lens, an objective lens, a dichroic beam splitter, and an image sensor. The structured light modulator can be configured to receive unstructured light beams and transmit structured light beams for illuminating micro-objects located within an enclosure of the microfluidic device and/or selectively activating one or more of a plurality of dielectrophoresis (DEP) electrodes of the microfluidic device. The first tube lens can be configured to capture the structured light beams transmitted by the structured light modulator. The second tube lens can be configured to transmit image light beams from the dichroic beam splitter to the image sensor. The image sensor can be configured to receive image light beams from the second tube lens. The image light beams received by the image sensor can be used to form an image of at least a portion of the microfluidic device.

## Claims

### Claim 1

An optical apparatus for imaging micro-objects in an enclosure of a microfluidic device, the optical apparatus comprising:
a structured light modulator configured to receive unstructured light beams from a first light source and reflect or transmit structured light beams suitable for illuminating micro-objects located in the enclosure of the microfluidic device;
a first tube lens configured to capture and transmit the structured light beams from the structured light modulator, comprising:
a first surface having a convex shape and a first radius of curvature;
a second surface having a second radius of curvature;
a third surface having a concave shape and a third radius of curvature;
a fourth surface having a concave shape and a fourth radius of curvature; and
a clear aperture with a diameter of at least 45 mm, wherein the first radius of curvature is positive, the third radius of curvature is negative, and the fourth radius of curvature is negative, and
wherein a front focal point and a back focal point of the tube lens are not equally spaced from a midpoint of the tube lens;
an objective lens configured to capture and transmit image light beams from a field of view comprising at least a portion of the enclosure of the microfluidic device;
a first dichroic beam splitter configured to receive and reflect or transmit the structured light beams from the first tube lens, and further configured to receive and transmit or reflect the image light beams from the objective lens;
a second tube lens configured to receive and transmit the image light beams from the first dichroic beam splitter; and
an image sensor configured to receive the image light beams from the second tube lens, wherein the image sensor forms an image of the field of view based upon the image light beams received from the second tube lens.

### Claim 2 (depends on 1)

The optical apparatus of claim 1, wherein the structured light modulator comprises an active area of at least 15 mm.

### Claim 3 (depends on 1)

The optical apparatus of claim 1, wherein the first tube lens has an effective focal length of about 162 mm or shorter.

### Claim 4 (depends on 1)

The optical apparatus of claim 1, wherein the first tube lens has an effective focal length of about 155 mm.

### Claim 5 (depends on 4)

The optical apparatus of claim 4, wherein the first tube lens has a numerical aperture of about 0.071 to about 0.085.

### Claim 6 (depends on 1)

The optical apparatus of claim 1, wherein the second tube lens has an effective focal length of about 180 mm or longer.

### Claim 7 (depends on 6)

The optical apparatus of claim 6, wherein the second tube lens has an effective focal length of about 200 mm.

### Claim 8 (depends on 1)

The optical apparatus of claim 1, wherein the second tube lens has a numerical aperture of about 0.063 to about 0.077.

### Claim 9 (depends on 1)

The optical apparatus of claim 1, wherein the image sensor comprises an active area of at least 18.0 mm.

### Claim 10 (depends on 1)

The optical apparatus of claim 1, wherein the apparatus is characterized by an aperture stop at the back of the objective lens, wherein the aperture stop is at least 25 mm.

### Claim 11 (depends on 1)

The optical apparatus of claim 1, wherein the objective lens is configured to minimize aberration in the image of the field of view formed by the image sensor.

### Claim 12 (depends on 11)

The optical apparatus of claim 11, wherein the second tube lens is configured to correct a residual aberration of the objective lens.

### Claim 13 (depends on 11)

The optical apparatus of claim 11, further comprising a correction lens configured to correct a residual aberration of the objective lens.

### Claim 14 (depends on 1)

The optical apparatus of claim 1, wherein the structured light modulator is disposed at a conjugate plane of the image sensor.

### Claim 15 (depends on 1)

The optical apparatus of claim 1 further comprising a first light source.

### Claim 16 (depends on 15)

The optical apparatus of claim 15, wherein the first light source has a power of at least 10 Watts.

### Claim 17 (depends on 15)

The optical apparatus of claim 15 further comprising a second light source.

### Claim 18 (depends on 17)

The optical apparatus of claim 17, wherein the second light source is configured to provide unstructured bright field illumination or wherein the second light source comprises a laser.

### Claim 19 (depends on 1)

The optical apparatus of claim 1, further comprising a second dichroic beam splitter.

### Claim 20 (depends on 17)

The optical apparatus of claim 17, further comprising a third light source, wherein the third light source comprises a laser, and wherein the laser of the third light source is configured to heat an internal surface of the microfluidic device and/or a liquid medium located within the enclosure of the microfluidic device.

### Claim 21 (depends on 1)

The optical apparatus of claim 1, further comprising a nest, wherein the nest is configured to hold the microfluidic device, provide at least one electrical connection to the microfluidic device, or provide fluidic connections to the microfluidic device.

### Claim 22 (depends on 1)

The optical apparatus of claim 1, further comprising a control unit for providing instructions to the structured light modulator, wherein the instructions cause the structured light modulator to produce one or more illumination patterns.

### Claim 23

A method of manipulating one or more micro-objects of a sample, the method comprising:
loading the sample containing the one or more micro-objects into a microfluidic device having an enclosure comprising a substrate, wherein the substrate comprises a plurality of light-actuated dielectrophoresis (DEP) electrodes disposed on or comprised by a surface of the substrate;
applying a voltage potential across the microfluidic device;
selectively activating a DEP force adjacent to at least one micro-object located within the microfluidic device by using an optical apparatus to project structured light onto a first position on the surface of the substrate of the microfluidic device, wherein the first position comprises one or more of the plurality of light-actuated DEP electrodes and is located adjacent to a second position on the surface of the substrate, the second position located beneath the at least one micro-object, and wherein the optical apparatus comprises:
a first light source;
a structured light modulator configured to receive unstructured light beams from the first light source and transmit structured light beams suitable for selectively activating the one or more DEP electrodes at the first position on the surface of the substrate of the microfluidic device;
a first tube lens configured to capture and transmit the structured light beams from the structured light modulator;
an objective lens configured to capture the structured light beams transmitted from the first tube lens and project the structured light beams onto the first position on the surface of the substrate of the microfluidic device, and wherein the objective lens is further configured to capture and transmit image light beams reflected or emitted from a field of view comprising at least a portion of the enclosure of the microfluidic device, the field of view encompassing the first and second positions on the surface of the substrate;
a first dichroic beam splitter configured to reflect or transmit to the objective lens the structured light beams received from the first tube lens, and further configured to transmit or reflect image light beams received from the objective lens;
a second tube lens configured to receive and transmit the image light beams from the first dichromic beam splitter; and
an image sensor configured to receive the image light beams from the second tube lens, wherein the image sensor records an image of the field of view based upon the image light beams received from the second tube lens; and
shifting the location of the DEP force generated adjacent to at least one micro-object by using the optical apparatus to move the projected structured light from the first position on the surface of the substrate of the microfluidic device to a third position on the surface of the substrate, wherein the third position also comprises one or more of the plurality of light-actuated DEP electrodes.

### Claim 24 (depends on 1)

The optical apparatus of claim 1, wherein a Back Focal Length (BFL) of the first tube lens is minimized.

### Claim 25 (depends on 1)

The optical apparatus of claim 1, wherein the first tube lens has an Effective Focal Length (EFL) of about 155 mm (e.g., 155 mm+/−1 mm) and a Back Focal Length (BFL) of about 135 mm (e.g., 135 mm+/−1 mm).

### Claim 26 (depends on 1)

The optical apparatus of claim 1, wherein the first tube lens has an Effective Focal Length (EFL) of about 162 mm (e.g., 162 mm+/−1 mm) and a Back Focal Length (BFL) of about 146 mm (e.g., 146 mm+/−1 mm).

### Claim 27 (depends on 1)

The optical apparatus of claim 1, wherein the second tube lens has an Effective Focal Length (EFL) of about 180 mm (e.g., 180 mm+/−1 mm) and a Back Focal Length (BFL) of about 164 mm (e.g., 164 mm+/−1 mm).

### Claim 28 (depends on 1)

The optical apparatus of claim 1, wherein the second tube lens has an Effective Focal Length (EFL) of about 200 mm (e.g., 200 mm+/−1 mm) and a Back Focal Length (BFL) of about 191 mm (e.g., 191 mm+/−1 mm).

### Claim 29

An optical apparatus for imaging micro-objects in an enclosure of a microfluidic device, the optical apparatus comprising:
a structured light modulator configured to receive unstructured light beams from a first light source and reflect or transmit structured light beams suitable for illuminating micro-objects located in the enclosure of the microfluidic device;
a first tube lens configured to capture and transmit the structured light beams from the structured light modulator;
an objective lens configured to capture and transmit image light beams from a field of view comprising at least a portion of the enclosure of the microfluidic device;
a first dichroic beam splitter configured to receive and reflect or transmit the structured light beams from the first tube lens, and further configured to receive and transmit or reflect the image light beams from the objective lens;
a second tube lens configured to receive and transmit the image light beams from the first dichroic beam splitter, wherein the second tube lens comprises:
a first surface having a convex shape and a first radius of curvature;
a second surface having a second radius of curvature;
a third surface having a concave shape and a third radius of curvature;
a fourth surface having a concave shape and a fourth radius of curvature; and
a clear aperture with a diameter of at least 45 mm;
wherein the first radius of curvature is positive, the third radius of curvature is negative, and the fourth radius of curvature is negative, and
wherein a front focal point and a back focal point of the tube lens are not equally spaced from a midpoint of the tube lens; and
an image sensor configured to receive the image light beams from the second tube lens, wherein the image sensor forms an image of the field of view based upon the image light beams received from the second tube lens.