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WO1997030338A1 - Systeme et procede d'analyse rapide de cellules par cytometrie spectrale - Google Patents

Systeme et procede d'analyse rapide de cellules par cytometrie spectrale Download PDF

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Publication number
WO1997030338A1
WO1997030338A1 PCT/US1997/001625 US9701625W WO9730338A1 WO 1997030338 A1 WO1997030338 A1 WO 1997030338A1 US 9701625 W US9701625 W US 9701625W WO 9730338 A1 WO9730338 A1 WO 9730338A1
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WO
WIPO (PCT)
Prior art keywords
cells
cell
spectra
present
spectrum
Prior art date
Application number
PCT/US1997/001625
Other languages
English (en)
Inventor
David S. Zakim
Max Diem
Original Assignee
Inphocyte, Inc.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Inphocyte, Inc. filed Critical Inphocyte, Inc.
Priority to AU18522/97A priority Critical patent/AU1852297A/en
Publication of WO1997030338A1 publication Critical patent/WO1997030338A1/fr

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Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/65—Raman scattering
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Optical investigation techniques, e.g. flow cytometry
    • G01N15/1456—Optical investigation techniques, e.g. flow cytometry without spatial resolution of the texture or inner structure of the particle, e.g. processing of pulse signals
    • G01N15/1459—Optical investigation techniques, e.g. flow cytometry without spatial resolution of the texture or inner structure of the particle, e.g. processing of pulse signals the analysis being performed on a sample stream
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Optical investigation techniques, e.g. flow cytometry
    • G01N15/149—Optical investigation techniques, e.g. flow cytometry specially adapted for sorting particles, e.g. by their size or optical properties
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Optical investigation techniques, e.g. flow cytometry
    • G01N2015/1486—Counting the particles
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/65—Raman scattering
    • G01N2021/651—Cuvettes therefore

Definitions

  • the limitauons of current diagnosuc pathology are illustrated in its inability to detect residual disease in patients.
  • Visual, microscopic examination is not able to delect leukemic cells in bone marrow samples, for example, when the concentration ot leukemic cells falls below 5% ol the total number of cells present.
  • This inability to delect the leukemic cells below 5% means that for the period of time it takes tor the level of diseased cells to amount to more than 5%, the physician and pauent can believe he or she is in remission (when in fact that is not the case). This can cause the physician to recommend the cessation of treatment when, in fact, the treatment should conunue.
  • the problem in detecting cancer cells in blood is similar to the one for detecung relauvely low levels of leukemic cells in bone marrow Cancer cells in the blood are mixed with the normal white cell elements.
  • the white cells elements outnumber the cancer cells (if present) on the order of thousands of normal white blood cells to one cancer cell.
  • the current best method of microscopic examinauon is to search for and detect cancer cells among a very large white blood cell fracuon.
  • detecung a relatively small number ot diseased cells among a iar larger number oi normal cells is ol increasing importance in the diagnosis ol disease
  • a given probe may bind with 10,000-told greater avidity to cancer cells versus normal cells
  • the number ol normal cells falsely detected as cancer cells with a probe even this selective will be 1 per 10,000 normal cells in the complete absence of cancer cells.
  • even a highly selective probe would not be useful for detecting cancer cells. Further, there is no way to determine what these results really mean because of the number of false negauves that also will obtain.
  • Rapid cytometry is a technique that causes cells to flow in a stream of fluid (usually water) such that the cells line up in smgle file. This is accomplished by forcing the cells into tubes with diameters on the order of the diameter of a single cell Once this separation of cells into a single line is achieved, the properties of each cell can
  • the examination tor properties of cells separated by the flow cytometry technique generally have to be optical because of the b ⁇ ef period ot time that each cell will be available tor testing.
  • the examination ot each cell in the stream is performed by lrradiauon with a laser at a predetermined point m the flow path.
  • the detecuon is performed by lrradiauon with a laser at a predetermined point m the flow path.
  • the 15 system is based on the fluorescence of mate ⁇ al in cells or the fluorescence of probe molecules attached to cells of interest.
  • the incident beam of light excites fluorescent molecules only in the single cell being irradiated.
  • the fluorescent signal is collected by a suitable detector, and the nature of the detection event is recorded. Scatte ⁇ ng of incident light, based on the size of the cell in the beam, also is used as a signal to detect and measure
  • Natural fluorescence oi normal cells and cancer cells will not discriminate between them To provide this discrimination, attempts have been made to label populations of different cells selectively For example, to detect small numbers of cancer cells in a large population ot normal cells, there would be an attempt to label only the cancer cells with a fluorescent molecule. When a tagged cancer cell passes through the beam ot incident light, a signal will be detected mdicaung the presence ot a cancer cell When a cell passes through the incident beam and no labeling is detected, this would be categorized as a normal cell Systems do exist tor detecting and counting normal cells in the blood in this manner.
  • J j when the fluorescent system can be constructed to collect relevant information about the cells of interest, which depends on the ability to label the cells of interest with high selectivity.
  • One convenuonally and widely used method is to mix the cell sample of interest with fluorescent antibodies that will bind specific molecular recogmuon sites on the surfaces of different populations ot cells.
  • Another is to make the DNA in the entire cell population fluorescent and then measure the amount ol DNA per cell.
  • the DNA content ot cancer cells differs from that ot normal cells. However, this difference is not always significant lor the purpose ot discriminating between different types of normal cells, or between cancer cells and normal cells.
  • the present invention is a system and method ot detecting diseased cells in a large population ol normal cells that uses flow cytometry, and Raman or resonance Raman spectroscopic techniques.
  • the present invenuon in essence, combines flow cytometry with a spectromet ⁇ c system.
  • This combined system does not depend on selecuve labeling of cells, nor the p ⁇ or treatment ot cells in any necessarily prescribed manner, nor the labeling ot ceils with probe molecules in order to discriminate between normal cells and diseased cells.
  • the present invention depends, as is highly desired, on detecting inherent properties ot each cell examined, inherent properties that not only discriminate between the presence or absence ot disease but also the degree to which a cell is diseased
  • the combined system ot the present mvenuon can be used to sort cells as they exit the flow system. Cells sorted in this manner into homogenous groups then may be subjected to more extensive study ot their properties.
  • the present invention combines the use of flow cytometry to separate cells of the sample populauon and Raman or resonance Raman spectroscopic techniques for detecting inherent differences in cells for the purpose ot determining whether cells are diseased and level of disease in the cells. This is accomplished by formmg a stream of cells by flow cytometry into the sampling region at which each cell is irradiated, lor example, by an incident beam ol a laser. The scattered Raman spectrum trom each cell is detected by a suitable detector More specifically, a complete Raman spectrum or resonance Raman spectrum is detected tor each cell flowing past the incident beam. This will provide information as to whether the specific cell m the beam ot the incident light is normal or diseased The level of disease also may be determined.
  • An object ot the present mvention is to provide a simple, easy, and inexpensive system and method for determining the presence or absence of disease in a population ot cells, and the level of disease in the cells
  • Another object ot the present invention is to provide a system and method that combines flow cytrometry and Raman and/or resonance Raman spectroscopic techniques to determine whether a given cell is diseased, normal, or somewhere between the two
  • Figure 1 shows a tirst embodiment ot the system ot the present invention
  • FIG. 2 shows a second embodiment ot the system ol the present invention Detailed Description ol the Drawings
  • the present invenUon is a detection system that combines flow cytrometry, and Raman or resonance Raman spectroscopy tor detecting the presence or absence ol diseased cells in a sample population ot cells.
  • Figure 1 generally at 100, shows a tirst embodiment ot the system ot the present mvenuon tor detecung diseased cells in a large population ot normal cells in the blood, bone marrow, body fluid, Ussue (treated to release cells for examination one by one in a flow system) or the like.
  • the system includes a flow cytometer 102 which causes the flow of cells in a fluid solution to separate into a smgle line in Ime 104. In line 104, the cells move one at one time in direction A.
  • Flow cytometer 104 includes sample chamber 106.
  • the sample chamber is at a locauon at which each cell in line 104 is subjected to an incident beam of light from laser 108.
  • the incident light from laser 108 is focused on each cell as it passes through sample chamber 106 of flow cytometer 102.
  • the beam excites the cells in sample chamber 106 with light at predetermined frequencies.
  • the scattered light responses from the cell bemg irradiated by laser 108 are
  • the demodulator separates the total response into the individual lrequencies representative ot the scattered light ot the irradiated cells in sample chamber 106.
  • the scattered light is collected at right angles to the incident beam that is focused on each cell so that there will be the rapid demodulauon of the frequencies of the scattered light.
  • the demodulator may be a graung, or such devices as an interferometry or 15 tunable acoustico-optical filters.
  • Detector 112 receives the demodulated signal and generates the vibrational spectra for scattered light and inputs it into computer 1 14.
  • Computer 1 14 controls the entire system. This includes flow cytometer 104, laser 108, demodulator 1 10, detector 112, and cell sorter 116.
  • Computer 114 upon receiving the vibra ⁇ onal spectra from detector 1 12, compares it with stored information to 0 determine if the cell bemg examined is a diseased cell or normal cell. If it is determined that it is diseased, it is also determines the level at which the cell is diseased. The results ot the comparison is output to display 1 15 for display.
  • computer 1 14 makes its determination as to the type ot cell that is being examined, it sends an approp ⁇ ate signal to cell sorter 1 16 to direct the cell at issue to a particular after examination location.
  • the cells can be directed to line 1 18 and 120 depending on the category in which the cell is placed. This includes normal and diseased cell locauons.
  • Raman spectroscopy is used to generate the necessary vibrauonal spectra lor each cell because it is sensitive lor detecung changes in the amount and in the chemical and physical properties ot molecules in the cell, which includes both simple and complex
  • Raman spectra in the above indicated manner for each cell has inherent advantages for diagnosis of rare events.
  • a first is that a characteristic vibrational spectrum will not be generated for debris that contains less than the complete components of a cell.
  • a second is that a marker of disease is an inherent property of the cell being examined and will be present if the complete components of a cell are present and the cell is diseased.
  • a third is that the Raman spectrum provides information on the inherent properties of the cells examined and is independent of any need to label cells to make them detectable to the analytical system.
  • a fourth is that the Raman spectrum of a cell contains a far larger amount of information about that cell than can be obtained with any current method, and, in particular, the significant differences in the spectral responses between types of cells.
  • a fifth is that a pulsed resonance Raman laser on a rapid time scale allows for rapid screening of cells. This method also reduces the fluorescence of cells, avoids heating of the cells, and degradation of the inherent spectral information within them.
  • a sixth is that any type of cell, even within the normal or diseased categories, can be differentiated from each other by the Raman spectrum for the cells because of the sensitivity of the response.
  • a seventh is that in addition to using vibrational (Raman) spectroscopy for diagnostic purposes, the spectra identified according to the present invention are usable for sorting cells.
  • the color of the incident beam of light may be selected to coincide with one or another molecular absorption for one of the components in cells.
  • These components could be proteins of the cell or the nucleic acids (DNA) of the cell.
  • the spectra obtained are resonance Raman spectra.
  • Resonance Raman spectroscopy will produce the spectra for selected types of molecules in the cell. These spectra will not have interfering, overlaping spectral bands from other molecules in the cells.
  • the intensity of the scattered light in resonance Raman spectroscopy can be as much as a million-times greater than in conventional Raman spectroscopy. This enhances signal strength and the signal-to-noise ratio of the generated signal. As such, there will be rapid collection of the spectra and rapid detection of cellular, spectral signatures for the purpose of disc ⁇ mination.
  • Resonance Raman spectra can be obtained at more than a single excitation frequency.
  • the incident light can be chosen at a frequency that is absorbed by protein, e.g., 280 nm, or in a frequency absorbed by DNA.
  • the selection of frequencies for resonance Raman spectroscopy may not be limited as long as it will excite a desired cell component or components. The use ot multiple frequencies will permit the selective acquisition of informa ⁇ on via resonance Raman spectroscopy for multiple types of molecules in cells.
  • incident beams of the desired different frequencies are arranged in tandem.
  • the beams will irradiate cells in a predetermined sequence.
  • the alternative method for using different frequencies to excite the cells is to arrange the excitation beams in parallel by splitting the stream of flowing cells into more than one channel.
  • the flow of cells is to more than one sample chamber when the chambers are arranged in parallel.
  • the resonance Raman spectra at different frequencies are not recorded for idenucal cells as in the first method.
  • the parallel arrangement also may be used for collecting Raman spectra at a single excitation frequency. This arrangement allows taster throughput of cells in the system and reduces the time for examination ot a single sample of cells.
  • FIG. 2 The embodiment of the present invention that uses two parallel paths is shown in Figure 2, generally at 200.
  • the flow of cells of interest are input to flow cytometer 202 and flow in direction A.
  • the cells enter line 204 one by one.
  • the cell flow is split by conventional means so that the flow is divided substantially equally in lines 206 and 208.
  • the cells in line 206 pass through sample chamber 210 where they are irradiated at a predetermined frequency by Raman laser 214.
  • the scattered light is demodulated by demodulator 218, as described for demodulator 110, and the demodulated signal is input to detector 220.
  • the vibrational spectra representative of the cell is output by detector 220 and is input to computer 222 where it is compared to determine if the irradiated cell is normal or diseased, and the level of disease in the cell. Based on the results of the comparison, the computer can control cell sorter 226 to send the cell under test to either output line 230 or hne 232 so that cells with the same characteristics can be placed together for further study if desired. Although only two output lines are shown, it is within the scope of the present invention that more lines can be added so that a more selective sort may take place.
  • the cells in line 208 undergo processing similar to the cells in line 206. Specifically, the cells in line 206 pass through sample chamber 212 where they are irradiated at a predetermined frequency (which may be the same or different from the trequency of laser 214) by laser 216. The scattered light is demodulated by demodulator 218. Detector 218 receives the output from demodulator 218 and generates a vibrational spectra representation of the cell i ⁇ adiated. The spectra is input to computer 222 where it is compared and analyzed to determine if the cell under test is normal or diseased, and the level at which it is diseased if it is in fact diseased. Computer 222 controls cell sorter 228 to output the cell under test to output line 234 or line 236 depending on the type cell it is. Again, there may be more output lines if it is desirable that the sorting be more selective.
  • Computer 222 controls the system elements so that both of the parallel Iines have access time to the computer, demodulators, detectors, and all other system elements without problem.
  • the system also includes display 224 to display desired information.
  • the computer operaung in real-time provides the operator with a real-time summary of the recording of events displayed on the display.
  • the events of importance may be the number of cells counted, the number of rare events corresponding to cancer cells, the number of rare events corresponding to characteristic spectra of other known types, and the number of events nol corresponding to normal events but not charactenstic of cancer cells.
  • Spectra for events already recorded can be down loaded for any pu ⁇ ose.
  • spectra can be in real time. However, if desired the spectra can be collected, stored, and analyzed at a later u e. It has been found that a complete spectrum of each cell will contain about 1-2 kb of data. The run time will vary depending on the sample size and testing criteria.
  • the system operator may select to save only a certain amount of spectra data collected. As such, representative spectra for normal cells, for diseased cells, or for particular types of cells may be saved for a given sample. Once this is done, when the comparison is completed, the count for a particular type of cell will be incremented to show that a match was found (but the actual spectra are not saved). However, if desired, the computer can save all spectra or just spectra which do not match anything that has been saved. This also includes saving more than one type of spectrum that is categorized as diseased so that the desired grading may be accomplished.
  • the system operator may choose to store spectra of diseased cells from each examination for archival pu ⁇ oses along with a typical spectrum of normal cells present in the examination.
  • This storage by the computer may be either internal or at an external location.
  • These data also can be transmitted to a remote site or an examining laboratory.
  • the examining laboratory may store the appropriate spectra on disks, which include identifying information about the patient and the source of the sample that was examined.
  • the examining laboratory may prepare a personal storage device card for the patient, which contains all relevant spectral data, inte ⁇ retations, basis for analysis, etc. that could be useful for following the subsequent course of the patient's disease.
  • the files in the storage card can be read by any facility equipped to conduct examinations according to the present invention. Therefore, there will be rapid and easy access to all relevant examinations done in the past at any time new examinations are conducted, whether or not the latter are carried out at the same facility as earlier examinations.
  • the data files being in the personal storage devices they will be available to the medical professional when needed to be able to compare changes in spectra of diseased cells as the patient's disease progresses or regresses.
  • the cells that are processed by the first or second embodiments of the present invention may be fresh.
  • the cells may be examined promptly after collection or after storage under conditions which preserve their freshness.
  • cells fixed with a variety of chemicals known to those skilled in the arts may be processed by the present invention.
  • the effects of fixation on the spectrum of a cell being examined are controlled by comparing the spectra of an unknown sample of cells that have been fixed prior to examination with stored data of spectra from known, well-studied, normal and diseased cells that were fixed in the same way as the cells in the unknown sample.
  • the present invention provides a system and method for searching for cancer in patients suspected of having cancer but in whom careful examination by other modalities may fail to find such cancer.
  • a major problem in the treatment of leukemia is the inability to find leukemic cells in the bone marrow when the concentration of leukemic cells fall below 5%.
  • the cancer cell burden in patients with acute myelogenous leukemia is on the order of 10 13 cells
  • a residual of 5% diseased cells represents 10- l -10 12 cancer cells present in the patient's bone marrow.
  • the present invention will permit treatment decisions to be driven by precise knowledge of the effect specific treatments have on the numbers of cancer cells left in die patient's bone marrow at each stage during the course of treatment.
  • the present invention will determine if a specific treatment has reached the limits of usefulness as reflected, for example, by no further decrease or increase in the numbers of cancer cells in the bone marrow.
  • the effect of diat treatment on the spectral properties of the cancer cells becomes a marker of therapeutically positive effects.
  • the disappearance of these markers in the spectra of cancer cells during treatment can be used as a basis for judging the efficacy of therapy in real time. This has implications for therapeutic decisions because it provides oncologists with a criterion. about a cancer, on which to base judgments on real-time to change treatment.
  • the present invention may be practical for determining the onset of a relapse of disease.
  • the sensitivity of the system and method of the present invention will determine the number of cancer cells present even though they fall into less for 5% category. This is, the present mvention will determine if the number of cancer cells at levels less than 5% of marrow cells is increasing despite conunued treatment.
  • the present invention will allow patients in remission who are no longer being actively treated, for example, with chemotherapy, to be able to detect early any relapse by the reappearance of cancer cells in the bone marrow.
  • the present invention also is very useful for detecting a very small number of cells in a large fluid volume, such as cells in urine. It further includes specimens in sputum, or cells scraped from the cervix, or cells removed from solid tissue by free needles aspiration (of a suspicious nature).
  • the system and method of the present invention have general application to cytological examinations of any type.
  • This present invenuon is favorably applied when cells in a given sample are derived from anatomically separate regions of an organ, for example, in cells in sputa that derive from all segments of the bronchial tree, or when cells in a sample are derived from multiple organs, as occurs for cells in urine (derived from kidney, bladder, collecting system of the urogenital tract, prostate) or cells in a sample removed from the cervix (cells potentially derived from the exocervix, the endocervix, uterus, and ovary.)
  • the strength of the present invenuon is for cytologic examinauon of specimens obtained from organs (like the lung), the cervix, or u ⁇ ne is the ability to distinguish between each type of cell no matter the rauo of the cells present.
  • the present invention also may be used to detect and quantitate any type ot cell with a Raman spectrum or a resonance Raman spectrum that differenuates it from other types of cells in the specimen.
  • the present invention may be used to detect selectively and to count the numbers of B and T lymphocytes in peripheral blood and to report on the presence of abnormalities in the T lymphocytes.
  • the present invention may be used to detect evidence for infection as reflected by abnormalities/reactive changes in white cells in blood.

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Abstract

L'invention concerne un système et un procédé pour détecter des cellules malades dans une grande population de cellules normales, et utilisant un cytomètre de flux et des techniques spectroscopiques vibrationnelles. Le système combine un cytomètre de flux avec un appareil spectrométrique pour déterminer les propriétés spécifiques de chaque cellule examinée, pour pouvoir non seulement déceler une maladie éventuelle, mais également l'importance des atteintes cellulaires. Plus particulièrement, le procédé combine la cytométrie de flux pour séparer les cellules de la population d'un échantillon et des techniques de spectroscopie Raman ou de spectroscopie Raman de résonance, pour détecter les différences dans les cellules afin de déterminer si les cellules sont malades et à quel degré elles sont atteintes. Ceci est réalisé en amenant un flux de cellules par un cytomètre de flux dans la région d'échantillonnage où chaque cellule est éclairée par un faisceau de laser Raman incident. Le spectre Raman diffusé de chaque cellule est détecté par spectroscopie Raman. Plus particulièrement, on détecte un spectre Raman complet ou un spectre Raman de résonance, pour chaque cellule traversant le faisceau incident. Le spectre va indiquer si une cellule particulière dans le faisceau de lumière incidente est normale ou malade et l'importance de l'atteinte peut être déterminée à partir de détails de ce spectre.
PCT/US1997/001625 1996-02-16 1997-02-04 Systeme et procede d'analyse rapide de cellules par cytometrie spectrale WO1997030338A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
AU18522/97A AU1852297A (en) 1996-02-16 1997-02-04 System and method for rapid analysis of cells using spectral cytometry

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US60227896A 1996-02-16 1996-02-16
US08/602,278 1996-02-16

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WO2003060486A1 (fr) * 2002-01-10 2003-07-24 Board Of Regents, The University Of Texas System Systeme de tri de flux et procedes associes
US7586604B2 (en) 1997-01-31 2009-09-08 Xy, Inc. Optical apparatus
US7629113B2 (en) 1997-12-31 2009-12-08 Xy, Inc Multiple sexed embryo production system for bovine mammals
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EP1815231B1 (fr) * 2004-11-17 2010-09-22 Honeywell International Inc. Cytometre en flux a base de detection raman
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WO2012059748A1 (fr) * 2010-11-04 2012-05-10 The University Of Nottingham Procédé, appareil et logiciel pour l'identification de cellules
US8253936B2 (en) 2008-08-08 2012-08-28 Chemimage Corporation Raman characterization of transplant tissue
US8416405B2 (en) 2008-08-08 2013-04-09 Chemimage Corporation Raman chemical imaging of implantable drug delivery devices
US8486618B2 (en) 2002-08-01 2013-07-16 Xy, Llc Heterogeneous inseminate system
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US9365822B2 (en) 1997-12-31 2016-06-14 Xy, Llc System and method for sorting cells
US20210215592A1 (en) * 2010-11-16 2021-07-15 1087 Systems, Inc. Use of vibrational spectroscopy for microfluidic liquid measurement
US11230695B2 (en) 2002-09-13 2022-01-25 Xy, Llc Sperm cell processing and preservation systems
US11796449B2 (en) 2013-10-30 2023-10-24 Abs Global, Inc. Microfluidic system and method with focused energy apparatus
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US12135270B2 (en) 2020-11-23 2024-11-05 Abs Global, Inc. Modular flow cytometry systems and methods of processing samples
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