Patent Issued for Ensemble-decision aliquot ranking (USPTO 11982678): University of Washington

2024 JUN 05 (NewsRx) -- By a News Reporter-Staff News Editor at Insurance Daily News -- University of Washington (Seattle, Washington, United States) has been issued patent number 11982678, according to news reporting originating out of Alexandria, Virginia, by NewsRx editors.

The patent’s inventors are Chiu, Daniel T. (Seattle, WA, US), Kuo, Jason S. (Seattle, WA, US), Schiro, Perry G. (Seattle, WA, US).

This patent was filed on October 26, 2018 and was published online on May 14, 2024.

From the background information supplied by the inventors, news correspondents obtained the following quote: “Body fluids are complex suspensions of biological particles in liquid. Blood, for example, includes plasma and cells (red blood cells, white blood cells, platelets) and the cells occupy about 55% of blood. Plasma is mostly water and it transfers proteins, ions, vitamins, enzymes, hormones, and other chemicals to cells in the body.

“Red blood cells are about 6 to 8 mm in size and serve to provide oxygen to cells. White blood cells are about 10 to 13 mm in diameter and they defend the body from disease as a part of an immune system by fighting against foreign viruses and bacteria. Platelets are the smallest cells, 1.5 to 3 mm, and they stop bleeding by forming blood clots.

“Fluids in addition to blood, such as saliva, tear, urine, cerebral spinal fluid as well as other body fluids in contact with various organs (e.g. lung) contain mixtures of cells and bioparticles. The type and amount of cells and bioparticles that are present in a particular body fluid (e.g. blood) reveals information about the health of the organism, and in the case of an infected individual, information about the diagnosis and prognosis of the disease.

“Some cells or bioparticles are present in rare quantities compared to the nominal concentrations of blood cells. Despite their rare occurrence, these cells or bioparticles may be intricately tied to significant events that take place in the body that alter the health state of an individual. These cells are commonly referred to as “rare cells”.

“For example, dissemination of cancer cells from the primary tumor is an important factor governing the probability of relapse and the survival rate in cancer patients. As cancer cells grow unregulated and lose their ability to adhere to each other, they can enter the blood and lymphatic circulation and circulate throughout the body. These cells are commonly referred to as Circulating Tumor Cells (CTC), Disseminated Tumor Cells (DTC), Circulating Cancer Cells (CCC), Circulating Epithelial Cells (CEC), Occult Tumor Cells (OTC), or other similar permutations to indicate the mobile nature of these cells, in contrast to the specimens obtained by direct biopsy of solid tumors. CTCs have been detected in the blood of patients suffering from all major cancers: prostate, ovarian, breast, gastric, colorectal, renal, lung, pancreatic, and others.

“In this fashion, tests that counts CTCs present in bodily fluids have been developed to assist with providing a prognosis for cancer patients. A “CTC test” can also be used to monitor a patient’s response to a particular treatment (e.g. radiation or chemotherapy) protocol. Based on the results of CTC test, a cancer patient may be able to avoid significant costs by minimizing additional unnecessary and expensive diagnostic tests and therapies, which are often times not covered by health insurance, for example Computed Tomography (CT) or Positron Emission Tomography (PET) scans, or shorten the drug treatments that are ineffective.

“However, CTCs are present in extremely low concentration in the peripheral blood, estimated to be on the order of one tumor cell per 10⁶ to 10⁷ mononuclear cells, which is equivalent to one tumor cell per 0.5 ml to 5 ml of peripheral blood. At such a low concentration, a sample with estimated 100 million mononuclear cells must be screened in order to detect at least one CTC with 99.995% certainty. Using conventional techniques, such as automatic digital microscopy (ADM) scanning at a typical speed of 800 cells/second, would require 18 hours to complete a sample that size, rendering it monetarily and temporally impractical for clinical use.

“For example, conventional flow cytometry may be employed to determine the presence or quantity of CTCs in a blood sample. However, flow cytometry requires that the cells are organized linearly in a row and detect each cell singularly because simultaneous detection of two cells cannot be interpreted correctly using existing technology. In flow cytometry, laser beams are focused such that they only illuminate a single particle at any given time. For example, if one nonfluorescent cell traverses the detection volume simultaneously with a fluorescent cell configured to be detectable by the flow cytometer, the nonfluorescent cell, being invisible to the flow cytometer, would be directed in the same trajectory as the fluorescent cell. To avoid misinterpretation in flow cytometry, cells suspensions are routinely either diluted or slowed down to allow sufficient distance between the cells to avoid overlapping of two cells within the detection volume. Current state-of-the-art flow cytometers have an upper limit of sorting 100,000 objects/second.

“As such, flow cytometry is not suitable for detecting or recovering rare cells. Detecting cells one at a time (serially) and making a decision on the trajectory of every cell is too time-consuming when analyzing a large number of cells. For example, since ten milliliter of blood contains approximately ten billion cells, at 100,000 cells per second, which is the highest sorting speed of state-of-the-art flow cytometer, it would take 100,000 second or 28 hours to completely sort the content of 10 mL. For rare cell, often 7-15 mL of blood is required to collect a statistically significant number of rare cells; using a flow cytometer to recover rare cells is an impractical consumption of clinical resources and can translate to a very high testing cost.

“As such, simple and cost/time-effective techniques are needed for the detection and quantitation of rare particles and cells in a fluid sample. The present invention satisfies these and other needs by providing methods and apparatuses for the detection of rare particles in fluid samples.”

Supplementing the background information on this patent, NewsRx reporters also obtained the inventors’ summary information for this patent: “Among other aspects, the present invention provides methods and apparatuses that rapidly scan a large volume of a fluid for the detection and or quantitation of desired bioparticles by ranking aliquots. In one aspect, the concept employed, termed Ensemble-Decision Aliquot Ranking (“eDAR”), is particularly useful for detecting rare cells in biofluids.

“In one aspect, the present invention provides a method for detecting a rare particle in a fluid sample, the method comprising the steps of detecting the presence or absence of the rare particle in an aliquot of the fluid sample, assigning a value to the aliquot based on the presence or absence of the rare particle, and directing the flow or collection of the aliquot based on the assigned value.

“In a second embodiment, the present invention provides a method for providing a subject a diagnosis or prognosis for a condition associated with the presence of a rare particle in a biological fluid, the method comprising the steps of detecting the presence or absence of the rare particle in an aliquot of the biological fluid, assigning a value to the aliquot based on the presence or absence of the rare particle, and providing a diagnosis or prognosis to the subject based on the assigned value.

“In a third aspect, the present invention provides a device for detecting a rare particle in a fluid sample, the device comprising at least a first input channel, at least two exit channels, at least one detector capable of detecting one or more rare particles in an aliquot of the fluid sample, a mechanism for directing the flow of the aliquot, and a computer capable of assigning a value to the aliquot based on the presence, absence, identity, composition, or quantity of the rare particles in the aliquot, wherein the computer is in communication with the detector and the mechanism for directing the flow of the aliquot.”

The claims supplied by the inventors are:

“1. A method for detecting a rare cell in a fluid sample comprising other cell population, the method comprising the steps of: (a) partitioning the fluid sample into plurality of aliquots, wherein the rare cell is a fluorescently labelled cell configured to express a fluorescent protein or the rare cell is labeled with a fluorescent detection reagent and wherein the rare cell is a cell present in the fluid sample at a low level comprising less than about 1 part per 10³ of the total cell population in the fluid sample; (b) detecting, with a detector, the presence or absence of a rare cell in each of the plurality of aliquots of the fluid sample, wherein the aliquot comprises a plurality of cells randomly distributed in a three-dimensional detection volume of the detector during detection, wherein the step of detecting the presence or absence of the rare cell comprises the sub-steps of: (i) illuminating simultaneously the plurality of cells of the aliquot with an external source of electromagnetic radiation; and (ii) detecting in the illuminated aliquot fluorescence from the plurality of cells including the rare cell; and © ranking each of the plurality of aliquots as a null or nonzero based on the absence or presence respectively of the fluorescence detected in each of the plurality of aliquots, and directing the flow of the aliquot to discard when the aliquot is ranked as null and directing the flow to collection when the aliquot is ranked as nonzero.

“2. The method of claim 1, wherein the individual aliquots are not physically separated.

“3. The method of claim 1, wherein the aliquot comprises a discrete volume containing the plurality of cells.

“4. The method of claim 1, wherein the aliquot contains more than one rare cell.

“5. The method of claim 1, wherein the detection step is performed during continuous flow of the aliquet through a flow channel.

“6. The method of claim 1, wherein a plurality of aliquots of the fluid sample are physically separated prior to the detection step.

“7. The method of claim 6, wherein the aliquots are partitioned into separate flow channels or chambers prior to the detection step.

“8. The method of claim 1, further comprising performing flow cytometry after step ©.

“9. A method for detecting a rare cell in a fluid sample containing other cell population, the method comprising: (a) contacting the fluid sample with a fluorescent detection reagent under conditions suitable to transform the detection reagent into a complex comprising the detection reagent and a rare cell, wherein the rare cell is present in the fluid sample at a low level comprising less than 1 part per 10³ of the total cell population; and partitioning the contacted fluid sample into plurality of aliquots; (b) detecting, with a detector, the presence or absence of a complex formed in step (a) in each of the plurality of aliquots of the fluid sample, wherein the aliquot comprises a plurality of cells randomly distributed in a three-dimensional detection volume of the detector during detection, wherein the step of detecting the presence or absence of the complex comprises the sub-steps of: (i) illuminating simultaneously the plurality of cells of the aliquot with an external source of electromagnetic radiation; and (ii) detecting in the illuminated aliquot fluorescence from the plurality of cells including the rare cell; © ranking each of the plurality of aliquots as a null or nonzero based on the absence or presence respectively of the fluorescence detected in each of the plurality of aliquots and directing the flow of the aliquots by channeling the aliquots based on the ranking of null or nonzero, wherein the aliquot is directed into a first channel or waste outlet when the ranking is null and the aliquot is directed to second channel or a first collection chamber when the ranking is nonzero.”

For the URL and additional information on this patent, see: Chiu, Daniel T. Ensemble-decision aliquot ranking. U.S. Patent Number 11982678, filed October 26, 2018, and published online on May 14, 2024. Patent URL (for desktop use only): https://ppubs.uspto.gov/pubwebapp/external.html?q=(11982678)&db=USPAT&type=ids

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