# US 8,772,693 B2 · System and method for Hilbert phase imaging

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- Number: US 8,772,693 B2
- Document ID: US8772693B2
- Status: Granted
- Kind: B2
- Application number: 11/389,670
- Filed: 2006-03-24
- Published: 2006-12-28
- Granted: 2014-07-08
- Earliest claimed: 2005-03-25
- Assignees: Massachusetts Institute of Technology; Hamamatsu Photonics K.K.
- Applicants: Gabriel Popescu; Ramachandra Dasari; Takahiro Ikeda
- Inventors: Gabriel Popescu; Ramachandra Dasari; Michael Feld; Takahiro Ikeda
- Web page: https://patentgrep.com/patent/US8772693B2

## CPC

- H10F 39/80: {Constructional details of image sensors}
- H10F 39/806: {Optical elements or arrangements associated with the image sensors}
- G01N 15/10: Investigating individual particles
- G01N 15/1433: using image recognition
- G01N 21/453: {Holographic interferometry (for dimensional measurements G01B9/021 - G01B9/029)}
- G01N 33/49: Blood {(chemical methods for determining blood cell populations G01N33/5094; chemical analysis of blood groups or blood types G01N33/80)}
- G06V 20/693: {Acquisition}
- G01N 2015/012: {Red blood cells}
- G01N 2015/016: {White blood cells}
- G01N 2015/145: {by pattern of light, e.g. fringe pattern}
- G01N 2015/1454: {using phase shift or interference, e.g. for improving contrast}
- G01N 2015/1497: {Particle shape}
- H10F: INORGANIC SEMICONDUCTOR DEVICES SENSITIVE TO INFRARED RADIATION, LIGHT, ELECTROMAGNETIC RADIATION OF SHORTER WAVELENGTH OR CORPUSCULAR RADIATION
- 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)
- G06V: IMAGE OR VIDEO RECOGNITION OR UNDERSTANDING

## Abstract

Hilbert phase microscopy (HPM) as an optical technique for measuring high transverse resolution quantitative phase images associated with optically transparent objects. Due to its single-shot nature, HPM is suitable for investigating rapid phenomena that take place in transparent structures such as biological cells. A preferred embodiment is used for measuring biological systems including measurements on red blood cells, while its ability to quantify dynamic processes on the millisecond scale, for example, can be illustrated with measurements on evaporating micron-size water droplets.

## Claims

### Claim 1

A method of imaging an object comprising:
using a light source to emit light along a first optical path including an object to be imaged, and along a second optical path that is spatially separated by a splitter from the first optical path at the object to be imaged;
combining light from the first optical path and light from the second optical path such that the combined light is incident on a two-dimensional imaging device having a plurality of pixels, the two dimensional imaging device receiving light along the first optical path that is tilted at an angle relative to the second optical path such that the incident light at the two-dimensional imaging device is received at different angles;
detecting a two dimensional image of the object with the two-dimensional imaging device; and
processing the two dimensional image with a Hilbert transform to obtain quantitative image data.

### Claim 2 (depends on 1)

The method of claim 1 further comprising orienting the fringe structure at an angle of 45° with respect to orthogonal axes of the imaging device.

### Claim 3 (depends on 1)

The method of claim 1 further comprising imaging a biological material.

### Claim 4 (depends on 1)

The method of claim 1 further comprising imaging tissue.

### Claim 5 (depends on 1)

The method of claim 1 further comprising providing a laser light source and a charge coupled device in a Fourier plane of the first optical path and the second optical path that detects an image.

### Claim 6 (depends on 1)

The method of claim 1 further comprising obtaining a quantitative spatial phase image of the object.

### Claim 7 (depends on 1)

The method of claim 1 further comprising removing a tissue or blood ample from a mammalian body for measurement.

### Claim 8 (depends on 1)

The method of claim 1 further comprising determining a size of a sample from an image of the sample.

### Claim 9 (depends on 1)

The method of claim 1 further comprising determining a volume of an object in a sample.

### Claim 10 (depends on 1)

The method of claim 1 further comprising imaging a blood cell.

### Claim 11 (depends on 1)

The method of claim 1 further comprising measuring a characteristic of a red or white blood cell.

### Claim 12 (depends on 1)

The method of claim 1 further comprising measuring a noise component in an image and removing the noise component from the image.

### Claim 13 (depends on 1)

The method of claim 1 further comprising obtaining an image with light reflected by an object.

### Claim 14 (depends on 1)

The method of claim 1 further comprising measuring hemolysis of a red blood cell.

### Claim 15 (depends on 1)

The method of claim 1 further comprising using an optical modulator to modulate light from the light source.

### Claim 16 (depends on 1)

The method of claim 1 further comprising measuring a crystalline material.

### Claim 17 (depends on 1)

The method of claim 1 further comprising measuring an optical fiber.

### Claim 18 (depends on 6)

The method of claim 6 further comprising obtaining the spatial phase image of the object by:
Fourier-transforming and high-pass filtering the measured spectral data to obtain a real sinusoidal signal component of a complex analytic signal;
obtaining the complex analytic signal associated with the sinusoidal signal by computing a 2D Fourier transform and suppressing the negative frequencies by employing a Hilbert transformation;
taking an inverse Fourier transform to obtain a complex 2D signal that provides phase information about the object; and
obtaining the quantitative spatial phase image by subtracting a linear phase component.

### Claim 19 (depends on 13)

The method of claim 13 further comprising attaching a reflective material to an object being measured.

### Claim 20 (depends on 19)

The method of claim 19 further comprising attaching polystyrene particles to the object.

### Claim 21

A Hilbert phase imaging device comprising:
a light source that is optically coupled to a first optical path that is separated from a second optical path with a splitter;
a two dimensional imaging device positioned to receive light from the first optical path and the second optical path, the two dimensional imaging device having a plurality of pixels along x and y axes of the imaging device;
a modulating element that positions light from the light source such that light on the first optical path is tilted at an angle relative to light on the second optical path, the light from the separated first optical path being combined with light from the separated second optical path with a combiner to be incident on the imaging device at different angles to detect two dimensional image data with the two-dimensional imaging device; and
a processor that receives the image data from the two-dimensional imaging device and performs a Hilbert phase transformation of two dimensional image data to form a quantitative phase image of an object.

### Claim 22 (depends on 21)

The device of claim 21 further comprising a fiber optic device that couples light from the light source to the first optical path and the second optical path.

### Claim 23 (depends on 21)

The device of claim 21 wherein a beam splitter combines a modulated reference image that optically interferes with a sample image.

### Claim 24 (depends on 21)

The device of claim 21 wherein the modulating device comprises a movable minor.

### Claim 25 (depends on 21)

The device of claim 21 wherein the modulating element comprises a optical element that tilts the first light path relative to the second light path.

### Claim 26 (depends on 21)

The device of claim 21 wherein a sample is mounted relative to an inverted microscope.

### Claim 27 (depends on 21)

The device of claim 21 wherein the processor includes a program that performs a Fourier transform on image data, applies a filter to the transformed image data and performs an inverse Fourier transform on the filtered image data.

### Claim 28 (depends on 21)

The device of claim 21 further comprising a blood or tissue sample holder.

### Claim 29 (depends on 21)

The device of claim 21 wherein light is reflected by a sample.

### Claim 30 (depends on 21)

The device of claim 21 wherein light is transmitted through the sample.

### Claim 31 (depends on 21)

The device of claim 21 wherein a reflective material is positioned on the sample.

### Claim 32 (depends on 21)

The device of claim 21 wherein the imaging device comprises a CCD or CMOS imaging device leaving at least 480×640 pixels and operating at least 10 frames per second.

### Claim 33 (depends on 26)

The device of claim 26 wherein the inverted microscope includes a sample holder, an objective lens, a beam splitter and a lens that couples light onto the imaging device positioned in a common Fourier plane.

### Claim 34 (depends on 22)

The device of claim 22 further comprising a fiber optic splitter, a fiber coupler, a first collimator, a second collimator and a second objective lens.

### Claim 35

A method of imaging an object comprising:
providing a light source, a first optical path including an object to be imaged, a second optical path and an imaging device;
positioning the imaging device relative to the first optical path and the second optical path;
tilting the first optical path relative to the second optical path such that light directed along the first optical path is incident upon the imaging device at a different angle than light directed along that second optical path that is incident upon the imaging device;
obtaining a two dimensional (2D) image of the object;
Fourier-transforming and filtering the image to obtain a real sinusoidal signal component of a complex analytic signal;
obtaining the complex analytic signal associated with the sinusoidal signal by computing a 2D Fourier transform and performing a Hilbert transformation;
taking an inverse Fourier transform to obtain a complex 2D signal that provides phase information about the object; and
obtaining a quantitative spatial phase image of the object.

### Claim 36 (depends on 35)

The method of claim 35 further comprising providing a laser light source and the imaging device in a Fourier plane of the first optical path and the second optical path that detects an image.

### Claim 37 (depends on 35)

The method of claim 35 further comprising imaging a biological material.

### Claim 38 (depends on 35)

The method of claim 35 further comprising imaging a fringe structure.

### Claim 39 (depends on 35)

The method of claim 35 further comprising imaging a blood or tissue sample.

### Claim 40 (depends on 35)

The method of claim 35 further comprising detecting 2D images at a rate of at least 10 frames per second with the imagine device.

### Claim 41

A method of imaging an object comprising:
illuminating an object to be imaged with light from a light source;
detecting light from the object with a two dimensional imaging device having a plurality of pixels, the two-dimensional imaging device receives light along a first optical path that interferes with light along a second optical path, the first optical path being tilted at an angle relative to the second optical path such that light is received at the two-dimensional imaging device along the first optical path and the second optical path at different angles; and
processing a two dimensional image that is detected by the two-dimensional imaging device with a Hilbert transform to obtain quantitative image data of the object.

### Claim 42 (depends on 41)

The method of claim 41 further comprising orienting the fringe structure at an angle of 45° with respect to orthogonal axes of the imaging device.

### Claim 43 (depends on 41)

The method of claim 41 further comprising imaging a biological material.

### Claim 44 (depends on 41)

The method of claim 41 further comprising imaging tissue.

### Claim 45 (depends on 41)

The method of claim 41 further comprising using a laser light source to illuminate the object and detecting the image with a charge coupled device in a Fourier plane of the first optical path and the second optical path that detects the image of the object.

### Claim 46 (depends on 21)

The device of claim 21 wherein the first optical path is tilted at an angle relative to the second optical path at the imaging device, such imaging device having at least 200,00 pixels to detect the two dimensional image of an object illuminated by the light source.

### Claim 47 (depends on 41)

The device of claim 41 further comprising coupling light from the light source to the object with a fiber optic device.

### Claim 48 (depends on 41)

The device of claim 41 further comprising processing the image with a data processor.