Provided herein are optical systems and methods for detecting and characterizing particles. Systems and method are provided which increase the sensitivity of an optical particle counter and allow for detection of smaller particles while analyzing a larger fluid volume. The described systems and methods allow for sensitive and accurate detection and size characterization of nanoscale particles (e.g., less than 50 nm, optionally less than 20 nm, optionally less than 10 nm) for large volumes of analyzed fluids.

Spatially distributed optical excitation and integrated waveguides are used for ultrasensitive particle detection based on individual electrokinetic velocities of particles. In some embodiments, chip-integrated systems are used to identify individual particles (e.g., individual molecules) based on their velocity as they move through an optically interrogated channel. Molecular species may be identified and quantified in a fully integrated setting, allowing for particle analysis including molecular analysis that can operate at low copy numbers down to the level of single-cell lysates. In some embodiments, the single-particle velocimetry-based identification and/or separation techniques are applied to various diagnostic assays, including nucleic acids, metabolites, macromolecules, organelles, cell, synthetic markers, small molecules, organic polymers, hormones, peptides, antibodies, lipids, carbohydrates, inorganic and organic microparticles and nanoparticles, whole viruses, and any combination thereof.

A particle detection device includes: a first light source to emit first irradiation light; a first light-collection member; a second light-collection member facing the first reflection surface; a second light source to emit second irradiation light; and a first light-reception element. When the first light source emits the first irradiation light, the first light-reception element detects, as the first incident light, scattered light generated when a particle existing at a detection position in a target space is irradiated with the first irradiation light. When the second light source emits the second irradiation light, the first light-reception element detects, as the first incident light, a light ray of the second irradiation light that is reflected by the first reflection surface and a light ray of the second irradiation light that is reflected by both the first reflection surface and the second reflection surface.

A method for characterizing a particle present in a sample, the sample lying between an image sensor and a light source and the sensor lying in a detection plane, includes illuminating the sample with the light source which emits an incident light wave propagating along a propagation axis, and acquiring an image of the sample with the sensor. The sensor is exposed to an exposure light wave. The image includes a plurality of elementary diffraction patterns each corresponding to one particle. The method also includes reconstructing a complex image representative of a complex amplitude of the light wave on a reconstruction surface passing through the sample, based on the acquired image; selecting a region of interest of the complex image corresponding to a particle of interest; forming an extracted image based on the region of interest; and characterizing the particle of interest depending on the extracted region of interest.

In one embodiment, a flow cytometer is disclosed having a compact light detection module. The compact light detection module includes an image array with a transparent block, a plurality of micro-mirrors in a row coupled to a first side of the transparent block, and a plurality of filters in a row coupled to a second side of the transparent block opposite the first side. Each of the plurality of filters reflects light to one of the plurality of micro-mirrors and passes light of a differing wavelength range and each of the plurality of micro-mirrors reflects light to one of the plurality of filters, such that incident light into the image array zigzags back and forth between consecutive filters of the plurality of filters and consecutive micro-mirrors of the plurality of micro-mirrors. A radius of curvature of each of the plurality of micro-mirrors images the fiber aperture onto the odd filters and collimates the light beam on the even filters.

The present invention discloses a three-dimensional diffraction tomography microscopy imaging method based on LED array coded illumination. Firstly, acquiring the raw intensity images, three sets of intensity image stacks are acquired at different out-of-focus positions by moving the stage or using electrically tunable lens. And then, after acquiring the intensity image stacks of the object to be measured at different out-of-focus positions, the three-dimensional phase transfer function of the microscopy imaging system with arbitrary shape illumination is derived. Further, the three-dimensional phase transfer function of the microscopic system under circular and annular illumination with different coherence coefficients is obtained as well, and the three-dimensional quantitative refractive index is reconstructed by inverse Fourier transform of the three-dimensional scattering potential function. The scattering potential function is converted into the refractive index distribution. Thus, the quantitative three-dimensional refractive index distribution of the test object is obtained. The invention realizes high-resolution and high signal-to-noise ratio 3D diffraction tomography microscopic imaging of cells, tiny biological tissues and other samples.

The present disclosure provides methods and systems for ghost cytometry (GC), which may be used to produce an image of an object without using a spatially resolving detector. This may be used to perform image-free ultrafast fluorescence “imaging” cytometry, based on, for example, a single pixel detector. Spatial information obtained from the motion of cells relative to a patterned optical structure may be compressively converted into signals that arrive sequentially at a single pixel detector. Combinatorial use of the temporal waveform with the intensity distribution of the random or pseudo-random pattern may permit computational reconstruction of cell morphology. Machine learning methods may be applied directly to the compressed waveforms without image reconstruction to enable efficient image-free morphology-based cytometry. Image-free GC may achieve accurate and high throughput cell classification as well as selective sorting based on cell morphology without a specific biomarker, which have been challenging using conventional flow cytometers.

A method includes launching, from an optical fiber, fluorescent light of differing wavelengths generated by different fluorochromes attached to particles in a sample fluid; magnifying an image size from an end of the optical fiber to a first dichroic filter of a row of a plurality of dichroic filters in a de-multiplexing imaging array; alternatively reflecting the fluorescent light between the plurality of dichroic filters and a plurality of micro-mirrors to collimate the fluorescent light on odd numbered dichroic filters and re-image the fluorescent light on even numbered dichroic filters; band passing different wavelength ranges of the fluorescent light at each of the plurality of dichroic filters to de-multiplex the wavelength spectrum of the wavelength range of the fluorescent light; and detecting fluorescent light in each of the different wavelength ranges to count a number of each of the different particles in the sample fluid.

Methods, systems, and devices are disclosed for imaging particles and/or cells using flow cytometry. In one aspect, a method includes transmitting a light beam at a fluidic channel carrying a fluid sample containing particles; optically encoding scattered or fluorescently-emitted light at a spatial optical filter, the spatial optical filter including a surface having a plurality of apertures arranged in a pattern along a transverse direction opposite to particle flow and a longitudinal direction parallel to particle flow, such that different portions of a particle flowing over the pattern of the apertures pass different apertures at different times and scatter the light beam or emit fluorescent light at locations associated with the apertures; and producing image data associated with the particle flowing through the fluidic channel based on the encoded optical signal, in which the produced image data includes information of a physical characteristic of the particle.

Aspects of the present disclosure include methods for producing an output laser beam having two or more angularly deflected laser beams (e.g., for irradiating a sample in a flow stream) with a predetermined intensity profile. Systems for practicing the subject methods having a laser, an acousto-optic device, a radiofrequency generator and a controller for adjusting the amplitude of the radiofrequency drive signals to produce an output laser beam of angularly deflected laser beams with a predetermined intensity profile are also described.