246 results about "Rare cell" patented technology

The present invention relates to methods for detecting, enriching, and analyzing rare cells that are present in the blood, e.g. fetal cells. The invention further features methods of analyzing rare cell(s) to determine the presence of an abnormality, disease or condition in a subject, e.g. a fetus by analyzing a cellular sample from the subject.

Described herein are methods to diagnose or prognose cancer in a subject by enriching, detecting, and analyzing individual rare cells, e.g., epithelial cells, in a sample from the subject. Also described are methods for labeling regions of genomic DNA in individual cells in said mixed sample with different labels wherein each label is specific to each cell and quantifying the labeled regions of genomic DNA from each cell in the mixed sample. More particularly the method includes detecting the presence of gene mutations in individual rare cells in a subsample.

The present invention provides systems, apparatuses, and methods to detect the presence of fetal cells when mixed with a population of maternal cells in a sample and to test fetal abnormalities, e.g. aneuploidy. The present invention involves labeling regions of genomic DNA in each cell in said mixed sample with different labels wherein each label is specific to each cell and quantifying the labeled regions of genomic DNA from each cell in the mixed sample. More particularly the invention involves quantifying labeled DNA polymorphisms from each cell in the mixed sample.

The invention features devices and methods for the deterministic separation of particles. Exemplary methods include the enrichment of a sample in a desired particle or the alteration of a desired particle in the device. The devices and methods are advantageously employed to enrich for rare cells, e.g., fetal cells, present in a sample, e.g., maternal blood and rare cell components, e.g., fetal cell nuclei. The invention further provides a method for preferentially lysing cells of interest in a sample, e.g., to extract clinical information from a cellular component, e.g., a nucleus, of the cells of interest. In general, the method employs differential lysis between the cells of interest and other cells (e.g., other nucleated cells) in the sample.

The present invention includes methods of enriching rare cells, such as cancer cells, from biological samples, such as blood samples. The methods include performing at least one debulking step on a blood sample and selectively removing at least one type undesirable component from the blood sample to obtain a blood sample that is enriched in a rare cell of interest. In some embodiments magnetic beads coupled to specific binding members are used to selectively removed components.

The present invention recognizes that diagnosis and prognosis of many conditions can depend on the enrichment of rare cells from a complex fluid sample. In particular, the enrichment of fetal cells from maternal samples, such as maternal blood samples, can greatly aid in the detection of fetal abnormalities or a variety of genetic conditions. In addition, the present invention recognizes that the enrichment of rare malignant cells from patient samples, can aid in diagnosis, prognosis, and development of therapeutic modalities for patients. The invention includes microfabricated filters for filtering fluid samples and methods of enriching rare cells of fluid samples using microfabricated filters of the present invention. The invention also includes solutions for the selective sedimentation of red blood cells (RBCs) from a blood sample and methods of using selective RBC sedimentation solutions for enriching rare cells of a fluid sample. Yet another aspect of the invention is an automated system for processing a fluid sample that includes: at least one filtration chamber that includes a microfabricated filter; automated means for directing fluid flow through at least one filtration chamber of the automated system, and means for collecting enriched rare cells. The present invention also includes methods of using automated systems for separating rare cells from fluid samples. Preferred fluid samples are blood, effusion, or urine samples, and rare cells that can be enriched from such sample include nucleated red blood cells and cancer cells.

A microflow apparatus for separating or isolating cells from a bodily fluid or other liquid sample uses a flow path where straight-line flow is interrupted by a pattern of transverse posts. The posts are spaced across the width of a collection region in the flow path, extending between the upper and lower surfaces thereof; they have rectilinear surfaces, have arcuate cross-sections, and are randomly arranged so as to disrupt streamlined flow. Sequestering agents, such as Abs, are attached to all surfaces in the collection region via a hydrophilic coating, preferably a hydrogel containing isocyanate moieties or a PEG or polyglycine of substantial length, and are highly effective in capturing cells or other targeted biomolecules as a result of such streamlined flow disruption.

The methods and reagents described in this invention are used to analyze circulating tumor cells, clusters, fragments, and debris. Analysis is performed with a number of platforms, including flow cytometry and the CellSpotter® fluorescent microscopy imaging system. Analyzing damaged cells has shown to be important. However, there are two sources of damage: in vivo and in vitro. Damage in vivo occurs by apoptosis, necrosis, or immune response. Damage in vitro occurs during sample acquisition, handling, transport, processing, or analysis. It is therefore desirable to confine, reduce, eliminate, or at least qualify in vitro damage to prevent it from interfering in analysis. Described herein are methods to diagnose, monitor, and screen disease based on circulating rare cells, including malignancy as determined by CTC, clusters, fragments, and debris. Also provided are kits for assaying biological specimens using these methods.

The present invention relates to methods for detecting, enriching, and analyzing rare cells that are present in the blood, e.g., epithelial cells. The invention further features methods of analyzing rare cell(s) to determine the presence of an abnormality, disease or condition in a subject by analyzing a cellular sample from the subject.

The invention features a device for the deterministic separation of analytes coupled to a reservoir containing a reagent that alters a magnetic propert of the analyte. Exemplary methods include the enrichment of a sample in a desired analyte (e.g., using deterministic separation) or the alteration of a desired analyte in the device. The devices and methods may be advantageously employed to enrich for rare cells, e.g., fetal cells or epithelial cells, present in a sample, e.g., maternal blood.

The present invention recognizes that diagnosis and prognosis of many conditions can depend on the enrichment of rare cells from a complex fluid sample. In particular, the enrichment of fetal cells from maternal samples, such as maternal blood samples, can greatly aid in the detection of fetal abnormalities or a variety of genetic conditions. In addition, the present invention recognizes that the enrichment of rare malignant cells from patient samples, can aid in diagnosis, prognosis, and development of therapeutic modalities for patients.A first aspect of the present invention is a microfabricated filter for filtering a fluid sample. A microfabricated filter of the present invention comprises at least one tapered pore, and preferably comprises at least two tapered pores whose variation in size is 20% or less. The present invention also includes a method of enriching rare cells of a fluid sample using a microfabricated filter of the present invention.Another aspect of the invention is solutions for the selective sedimentation of red blood cells (RBCs) from a blood sample comprising a red blood cell aggregating agent and at least one specific binding member that selectively binds RBCs. Solutions of the present invention include a combined solution for rare cell enrichment that comprise RBC aggregating agents, at least one specific binding member that selectively binds RBCs, and at least one additional specific binding member for the removal of undesirable sample components other than RBCs. The invention also includes methods of using selective RBC sedimentation solutions and combined solutions for enriching rare cells of a fluid sample.Yet another aspect of the invention is an automated system for processing a fluid sample that includes: at least one filtration chamber that comprises or engages one or more microfabricated filters of the present invention; automated means for directing fluid flow through the one or more filtration chambers of the automated system, and means for collecting enriched rare cells. The present invention also includes methods of using an automated system for separating rare cells from a fluid sample. Preferred fluid samples are effusion, blood, or urine samples, and rare cells that can be enriched from such sample include nucleated red blood cells and cancer cells.

The present invention recognizes that diagnosis and prognosis of many conditions can depend on the enrichment of rare cells, especially tumor cells, from a complex fluid sample such as a blood sample. In particular, the present invention is directed to methods and compositions for detecting a non-hematopoietic cell, e.g., a non-hematopoietic tumor cell, in a blood sample via, inter alia, removing red blood cells (RBCs) from a blood sample using a non-centrifugation procedure, removing white blood cells (WBCs) from said blood sample to enrich a non-hematopoietic cell, if any, from said blood sample; and assessing the presence, absence and / or amount of said enriched non-hematopoietic cell.

A highly sensitive assay is disclosed which utilizes a method for gene specific primed amplification of mRNA libraries from rare cells and rare transcripts found in blood. The assay allows detection of rare mRNA (10 copies / cell) found in 1 to 10 cells isolated through immunomagnetic enrichment. The assay is an improvement over multiplex PCR and allows efficient detection of rare coding sequences for circulating carcinoma cells in the blood. The methods are useful in profiling of cells isolated from tissues or body fluids and serves as an adjunct to clinical diagnosis of diverse carcinomas including early stage detection and classification of circulating tumor cells. Monitoring of nucleic acid and protein profiles of cells either in conventional or microarray formats, facilitates management of therapeutic intervention including staging, monitoring response to therapy, confirmation of remission and detection of regression.

The methods and reagents described in this invention are used to analyze circulating tumor cells, clusters, fragments, and debris. Analysis is performed with a number of platforms, including flow cytometry and the CellSpotter® fluorescent microscopy imaging system. Analyzing damaged cells has shown to be important. However, there are two sources of damage: in vivo and in vitro. Damage in vivo occurs by apoptosis, necrosis, or immune response. Damage in vitro occurs during sample acquisition, handling, transport, processing, or analysis. It is therefore desirable to confine, reduce, eliminate, or at least qualify in vitro damage to prevent it from interfering in analysis. Described herein are methods to diagnose, monitor, and screen disease based on circulating rare cells, including malignancy as determined by CTC, clusters, fragments, and debris. Also provided are kits for assaying biological specimens using these methods.

Methods of detecting multiple nucleic acid targets in single cells through indirect capture of labels to the nucleic acids are provided. Methods of assaying the relative levels of nucleic acid targets through normalization to levels of reference nucleic acids are also provided. Methods of detecting individual cells, particularly rare cells from large heterogeneous cell populations, through detection of nucleic acids are described. Related compositions, systems, and kits are also provided.

A microflow apparatus for separating or isolating cells from a bodily fluid or other liquid sample uses a flow path where straight-line flow is interrupted by a pattern of transverse posts. The posts are spaced across the width of a collection region in the flow path, extending between the upper and lower surfaces thereof; they have rectilinear surfaces, have arcuate cross-sections, and are randomly arranged so as to disrupt streamlined flow. Sequestering agents, such as Abs, are attached to all surfaces in the collection region via a hydrophilic coating, preferably a hydrogel containing isocyanate moieties or a PEG or polyglycine of substantial length, and are highly effective in capturing cells or other targeted biomolecules as a result of such streamlined flow disruption.

Compositions and methods for stabilizing rare cells in blood specimens, preserving the quality of blood specimens, and also serving as cell fixatives are disclosed which minimize losses of target cells (for example, circulating tumor cells) and formation of debris and aggregates from target cells, non-target cells and plasma components, thereby allowing more accurate analysis and classification of circulating tumor cells (CTC) and, ultimately, of tumor burdens in cancer patients. Stabilization of specimens is particularly desirable in protocols requiring rare cell enrichment from blood specimens drawn from cancer patients. Exposure of such specimens to potentially stressful conditions encountered, for example, in normal processing, mixing, shaking, delays due to transporting the blood, has been observed to not only diminish the number of CTC but also to generate debris and aggregates in the blood specimens that were found to interfere with accurate enumeration of target cells, if present. Stabilizers are necessary to discriminate between in vivo CTC disintegration and in vitro sample degredation.

Methods of detecting multiple nucleic acid targets in single cells through indirect capture of labels to the nucleic acids are provided. Methods of assaying the relative levels of nucleic acid targets through normalization to levels of reference nucleic acids are also provided. Methods of detecting individual cells, particularly rare cells from large heterogeneous cell populations, through detection of nucleic acids are described. Related compositions, systems, and kits are also provided.

A cell separation process for the isolation of pathogenic cells in a small concentration in a biological fluid specimen, including treating the biological specimen to modify in a differential manner selected rare cells and other cells and causing differential migration of cells that reacted during treatment versus cells that did not react during treatment.

Method of enriching specific cells from cellular samples are disclosed, comprising contacting in solution a cellular sample with affinity-tagged ligands (ATLs) each comprising a first ligand linked to an affinity tag, wherein the ligand selectively binds a cellular marker of the rare cells and the affinity tag can be selectively captured by a capture moiety, wherein the affinity tags do not comprise a magnetic particle; and flowing the sample through a microfluidic device comprising the capture moiety to selectively retain ATL-bound cells. Methods for enriching circulating tumor cells, and devices for enriching specific cells from cellular samples are also disclosed.

The present invention provides methods and compositions for isolating and detecting rare cells from a biological sample containing other types of cells. In particular, the present invention includes a debulking step that uses a microfabricated filters for filtering fluid samples and the enriched rare cells can be used in a downstream process such as identifies, characterizes or even grown in culture or used in other ways. The invention also include a method of determining the aggressiveness of the tumor or of the number or proportion of cancer cells in the enriched sample by detecting the presence or amount of telomerase activity or telomerase nucleic acid or telomerase expression after enrichment of rare cells. This invention further provides an efficient and rapid method to specifically remove red blood cells as well as white blood cells from a biological sample containing at least one of each of red blood cells and white blood cells, resulting in the enrichment of rare target cells including circulating tumor cells (CTC), stromal cells, mesenchymal cells, endothelial cells, fetal cells, stem cells, non-hematopoietic cells etc from a blood sample. The method is based upon combination of immuno-microparticles (antibody coated microparticles) and density-based separation. The final enriched target cells can be subjected to a variety of analysis and manipulations, such as flowcytometry, PCR, immunofluorescence, immunocytochemistry, image analysis, enzymatic assays, gene expression profiling analysis, efficacy tests of therapeutics, culturing of enriched rare cells, and therapeutic use of enriched rare cells. In addition, depleted plasma protein and white blood cells can be optionally recovered, and subjected to other analysis such as inflammation studies, gene expression profiling, etc.

The invention encompasses methods for diagnosing, theranosing, or prognosing a condition in a patient based on the results of one or more analysis methods. The methods can comprise enriching a sample obtained from the patient for one or more rare cells. The analysis methods can include performing enumeration of the one or more rare cells or cell subtypes, performing nucleic acid analysis, or detecting a serum marker.

The present invention relates to microfluidic devices and methods for detecting rare cells. The disclosed microfluidic devices and methods integrate and automate sample preparation, cell labeling, cell sorting and enrichment, and DNA / RNA analysis of sorted cells.

Provided herein, among other aspects, are methods and apparatuses for ranking aliquots from a suspension containing bioparticles. In certain embodiments, the bioparticles may be cells, organelles, proteins, DNAs, debris of biological origin, microbeads coated with biological compounds, or viral particles. As such, the methods and apparatuses provided herein may be used to quantify rare cells such as circulating cancer cells, fetal cells and other rare cells present in bodily fluids for disease diagnosis, prognosis, or treatment.

Method of enriching specific cells from cellular samples are disclosed, comprising contacting in solution a cellular sample with affinity-tagged ligands (ATLs) each comprising a first ligand linked to an affinity tag, wherein the ligand selectively binds a cellular marker of the rare cells and the affinity tag can be selectively captured by a capture moiety, wherein the affinity tags do not comprise a magnetic particle; and flowing the sample through a microfluidic device comprising the capture moiety to selectively retain ATL-bound cells. Methods for enriching circulating tumor cells, and devices for enriching specific cells from cellular samples are also disclosed.

Fluorescent dye-tagged rare cells are disposed in a biological fluid layer contained in an annular gap (12) between an inside test tube wall (14) and a float wall (16). One or more analysis images are acquired at one or more focus depths using a microscope system (10, 10′, 10″, 10′″). Each acquired analysis image is image processed each to identify candidate rare cell images. The test tube (72) is selectively rotated. The test tube and the microscope system are selectively relatively translated along a test tube axis (75). The acquiring and image processing are repeated for a plurality of fields of view accessed by the selective rotating and selective relative translating and substantially spanning the annular gap.

The invention encompasses methods and devices for diagnosing, theranosing, or prognosing a condition in a patient by enriching a sample in rare cells or other particles. The devices can be a microfluidic device comprising an array of obstacles and one or more binding moieties. The devices and methods can allow for enrichment of cells based on size and affinity, recovery of cells or other particles in locations on the microfluidic device, release of cells or other particles from the microfluidic device, flow of sample through the microfluidic device, and retention of rare cells or other particles from a sample obtained from a patient having a condition.

Spheroidal beads present an exterior surface of a hydrophilic hydrogel, which is an isocyanate-functional polymer that is polymerized by urethane bonds and cross-linked by urethane and urea bonds. Sequestering agents present at the surface are covalently bound to isocyanate groups or to intermediate linkers that are so bound. These beads allow sequestering agents to retain their native three-dimensional configuration, and as a result of such surface characteristics and hydrophilicity, they achieve highly effective capture of very small subpopulations of rare cells from bodily fluids or the like and very effectively deter nonspecific binding of other biomaterials present in such bodily fluid. They may be all-hydrogel spheroids or hydrogel-coated substrates.

Disclosed herein are methods for identifying rare cells containing particular markers and / or alleles from biological samples that have not been substantially pre-processed (e.g., unprocessed whole blood). The methods described herein provide a system for digital enrichment of target cells from a biological sample and detection of such target cells, thereby allowing accurate and efficient detection and / or enumeration of such cells in the sample.

The present invention provides systems, apparatuses, and methods to detect the presence of fetal cells when mixed with a population of maternal cells in a sample and to test fetal abnormalities, e.g. aneuploidy. The present invention involves labeling regions of genomic DNA in each cell in said mixed sample with different labels wherein each label is specific to each cell and quantifying the labeled regions of genomic DNA from each cell in the mixed sample. More particularly the invention involves quantifying labeled DNA polymorphisms from each cell in the mixed sample.