641 results about "In situ hybridization" patented technology

Conjugate compositions are disclosed that include a specific-binding moiety covalently coupled to a nanoparticle through a heterobifunctional polyalkyleneglycol linker. In one embodiment, a conjugates is provided that includes a specific-binding moiety and a fluorescent nanoparticle coupled by a heterobifunctional PEG linker. Fluorescent conjugates according to the disclosure can provide exceptionally intense and stable signals for immunohistochemical and in situ hybridization assays on tissue sections and cytology samples, and enable multiplexing of such assays.

Disclosed are new methods comprising the use of in situ hybridization to detect abnormal nucleic acid sequence copy numbers in one or more genomes wherein repetitive sequences that bind to multiple loci in a reference chromosome spread are either substantially removed and / or their hybridization signals suppressed. The invention termed Comparative Genomic Hybridization (CGH) provides for methods of determining the relative number of copies of nucleic acid sequences in one or more subject genomes or portions thereof (for example, a tumor cell) as a function of the location of those sequences in a reference genome (for example, a normal human genome). The intensity(ies) of the signals from each labeled subject nucleic acid and / or the differences in the ratios between different signals from the labeled subject nucleic acid sequences are compared to determine the relative copy numbers of the nucleic acid sequences in the one or more subject genomes as a function of position along the reference chromosome spread. Amplifications, duplications and / or deletions in the subject genome(s) can be detected. Also provided is a method of determining the absolute copy numbers of substantially all RNA or DNA sequences in subject cell(s) or cell population(s).

A spectral imaging method for simultaneous detection of multiple fluorophores aimed at detecting and analyzing fluorescent in situ hybridizations employing numerous chromosome paints and / or loci specific probes each labeled with a different fluorophore or a combination of fluorophores for color karyotyping, and at multicolor chromosome banding, wherein each chromosome acquires a specifying banding pattern, which pattern is established using groups of chromosome fragments labeled with various fluorophore or combinations of fluorophores.

The present invention provides methods for diagnosing and treating cancer, and in particular methods for determining the susceptibility of subjects suspected of having breast cancer (or known to have breast cancer) to treatment with topoisomerase II inhibitors. The present invention also provides in situ hybridization probes for specifically detecting topoIIα gene sequences.

Antibody / signal-generating moiety conjugates are disclosed that include an antibody covalently linked to a signal-generating moiety through a heterobifunctional polyalkyleneglycol linker. The disclosed conjugates show exceptional signal-generation in immunohistochemical and in situ hybridization assays on tissue sections and cytology samples. In one embodiment, enzyme-metallographic detection of nucleic acid sequences with hapten-labeled probes can be accomplished using the disclosed conjugates as a primary antibody without amplification.

The present invention provides methods and compositions for specifically identifying a fetal cell. An initial screening of approximately 400 candidate genes by digital PCR in different fetal and adult tissues identified a subset of 24 gene markers specific for fetal nucleated RBC and trophoblasts. The specific expression of those genes was further evaluated and verified in more defined tissues and isolated cells through quantitative RT-PCR using custom Taqman probes specific for each gene. A subset of fetal cell specific markers (FCM) was tested and validated by RNA fluorescent in situ hybridization (FISH) in blood samples from non-pregnant women, and pre-termination and post-termination pregnant women. Applications of these gene markers include, but are not limited to, distinguishing a fetal cell from a maternal cell for fetal cell identification and genetic diagnosis, identifying circulating fetal cell types in maternal blood, purifying or enriching one or more fetal cells, and enumerating one or more fetal cells during fetal cell enrichment.

The invention provides a method for more effective treatment of patients susceptible to or diagnosed with tumors overexpressing HER2, as determined by a gene amplification assay, with a HER2 antibody. Such method comprises administering a cancer-treating dose of the HER2 antibody, preferably in addition to chemotherapeutic agents, to a subject in whose tumor cells her2 has been found to be amplified e.g., by fluorescent in situ hybridization.

The invention enables in situ genomic and transcriptomic assessment of nucleic acids to be conducted in biological specimens that have been physically expanded. The invention leverages the techniques for expansion microscopy (ExM) to provide new methods for in situ genomic and transcriptomic assessment of nucleic in a new process referred to herein as “expansion fluorescent in situ hybridization” (ExFISH).

This invention relates to imaging, such as by expansion microscopy, labelling, and analyzing biological samples, such as cells and tissues, as well as reagents and kits for doing so.

The present invention relates to a novel method for the amplification of DNA, this method being particularly useful for the amplification of the DNA or the whole genome of a single cell, chromosomes or fragments thereof. Described is also the use of the method in DNA analysis for medical, forensic, diagnostic or scientific purposes, like comparative genomic hybridization (CGH)-, fluorescence in situ hybridization (FISH)-, polymerase chain reaction (PCR)-, single strand conformation polymorphism (SSCP)-, DNA sequence-, "loss of heterozygosity" (LOH)-, fingerprint- and / or restriction fragment length polymorphism (RFLP)-analysis.

The present inventors have shown that the gene and protein expression profiles of ADAM8, ADAM10 and ADAM12 in different grades and stages of bladder cancer.ADAM12 gene expression was evaluated in tumors from 96 patients with bladder cancer using a customized Affymetrix GeneChip. Gene expression in bladder cancer was validated using reverse transcription-polymerase chain reaction (RT-PCR), quantitative PCR, and in situ hybridization. Protein expression was evaluated by immunohistochemical staining on tissue arrays of bladder cancers.The presence and relative amount of ADAM12 in the urine of cancer patients were determined by Western blotting and densitometric measurements, respectively.Particularly ADAM12 mRNA expression was significantly upregulated in bladder cancer, as determined by microarray analysis, and the level of ADAM12 mRNA correlated with disease stage. ADAM12 protein expression correlated with tumor stage and grade. ADAM12 was present in higher levels in the urine from bladder cancer patients than in urine from healthy individuals. Significantly, following removal of tumor by surgery, in most bladder cancer cases examined the level of ADAM12 in the urine decreased and, upon recurrence of tumor, increased.

The invention disclosed herein improves upon existing tissue microarray technology by using frozen tissues embedded in tissue embedding compound as donor samples and arraying the specimens into a recipient block comprising tissue embedding compound. Tissue is not fixed prior to embedding, and sections from the array are evaluated without fixation or post-fixed according to the appropriate methodology used to analyze a specific gene at the DNA, RNA, and / or protein levels. Unlike paraffin tissue arrays which can be problematic for immunohistochemistry and for RNA in situ hybridization analyses, the disclosed methods allow optimal evaluation by each technique and uniform fixation across the array panel. The disclosed arrays work well for DNA, RNA, and protein analyses, and have significant qualitative and quantitative advantages over existing methods.

Disclosed is a method for in situ detection of one or more target nucleic acids based on a combination of an in situ hybridization (ISH) assay method and a general ISH signal amplification method. This new method produces high signal intensity and while keeps low background noise of signal amplification. The result can be consistently reproduced and the method can be easily adopted for routine clinic diagnostic use. Further, the invention relates to a kit, comprising the components of the ISH assay and a general ISH signal amplification assay, for sensitive detection of one or more target nucleic acids.

A method and system for automated digital fluorescent in situ hybridization (FISH) image analysis. Luminance parameters from a digital image of a biological tissue sample to which a fluorescent compound (e.g., LSI-HER-2 / neu and CEP-17 dyes) have been applied are analyzed to determine plural regions of interest. Fluorescent color signals in the plural regions of interest including plural cell nuclei are identified, classified and grouped into plural groups. Each of the plural groups is validated based on pre-defined conditions. A medical diagnosis or prognosis or medical, life science or biotechnology experiment conclusion determined using a count of plural ratios of validated fluorescent color signals within each of the cell nuclei within the plural groups.

The present invention provides reagents for use in an automated environment for cell conditioning of biological samples wherein the cells or tissues are predisposed for access by reagent molecules for histochemical and cytochemical staining procedures. The components of the reagents are optimized to facilitate molecular access to cells and cell constituents within the biological sample. The present invention also provides reagents for use in an automated environment for removing or etching embedding media by exposing a biological sample to be stained in histochemical or cytochemical procedures without the dependence on organic solvents. The components of the reagents are optimized to facilitate removal or etching of the embedding media from the biological sample.

Amplification and overexpression of theHER-2 oncogene in breast cancer is felt to be stable over the course of disease and concordant between the primary tumor and metastases. Therefore, patients with HER-2 negative primary tumors will rarely receive anti-HER-2 antibody therapy. A very sensitive blood test is used to capture circulating tumor cells (CTC's) and evaluate their HER-2 gene status by FISH evaluation. The HER-2 status of the primary tumor and corresponding CTC's is used to assess the ratio of CTC's as a reliable surrogate marker. HER-2 expression of 10 CTC's is sufficient to make a definitive diagnosis of the HER-2 gene status for the whole population of CTC's in patients with recurrent breast cancer.

Provided is an HPV E6, E7 mRNA assay, referenced herein as the “In Cell HPV Assay,” that is capable of sensitive and specific detection of normal cervical cells undergoing malignant transformation as well as abnormal cervical cells with pre-malignant or malignant lesions. The In Cell HPV Assay identifies HPV E6, E7 mRNA via in situ hybridization with oligonucleotides specific for HPV E6, E7 mRNA and quantitates the HPV E6, E7 mRNA via flow cytometry. The In Cell HPV Assay can be carried out in less than three hours directly from liquid-based cervical (“LBC”) cytology specimens. The In Cell HPV Assay provides an efficient and highly sensitive alternative to the Pap smear for determining abnormal cervical cytology.

Scanning and analysis of cytology and histology samples uses a flatbed scanner to capture images of the structures of interest such as tumor cells in a manner that results in sufficient image resolution to allow for the analysis of such common pathology staining techniques as ICC (immunocytochemistry), IHC (immunohistochemistry) or in situ hybridization. Very large volumes of such material are scanned in order to identify cells or clusters of cells which are positive or warrant more detailed examination, and if analysis at higher resolution is necessary, information regarding these positive events is transferred to a secondary microscope, such as a conventional scanning microscope, to allow further analysis and review of the selected regions of the slide containing the sample.

The invention provides a method for more effective treatment of patients susceptible to or diagnosed with tumors overexpressing ErbB, as determined by a gene amplification assay, with an ErbB antagonist. Such method comprises administering a cancer-treating dose of the ErbB antagonist, preferably in addition to chemotherapeutic agents, to a subject in whose tumor cells ErbB has been found to be amplified e.g., by fluorescent in situ hybridization. ErbB antagonists described include an anti-HER2 antibody. Pharmaceutical packaging for providing the components for such treatment is also provided.

A computer controlled method for detecting and diagnosing a rare cell type in a tissue sample is provided, said method comprising treating the tissue sample such that it generates a first signal indicative of the presence at a location of a rare cell, detecting the first signal, treating the location at which the first signal is detected to generate a second signal indicative of a diagnostically useful cellular characteristic and detecting the second signal. The first signal can be morphological or a color present in a sought cell either before or after staining. The second signal can be generated by in situ PCR or PCR in situ hybridization. In one preferred embodiment, the rare cell type is a fetal cell in a maternal blood tissue sample, said sample consisting of a smear of unenriched maternal blood. In another embodiment, the method is used to diagnose or genotype cancer cells in a blood or tissue biopsy sample.

Methods and apparatuses for gently removing embedding media from biological samples at temperatures below the embedding medium melting point with liquid composition using batch methods or automated instruments prior to immunohistochemical (IHC), in situ hybridization (ISH) or other special staining or histochemical or cytochemical manipulations.

Novel methods of probing multiple targets in a biological sample are provide whereby the targets are DNA, RNA and protein. The method comprises subjecting the sample to an in situ hybridization reaction using a labeled nucleic acid probe that binds an RNA target, observing a signal, and optionally removing the signal. The method further comprises an antigen retrieval protocol, observing a signal, removing the signal, and optionally applying a protease treatment to access the sample's DNA targets by subjecting the sample to an in situ hybridization reaction using a labeled nucleic acid probe, observing a signal from the labeled DNA targets, and optionally removing the signal.

This invention relates to the original position cross checking method of a Hence Kong waters disease type Pamela Ci's body and its reagent box. This invention includes the following steps: composing the three oligoribonucleotides probes of the High Sim markings according to the special DNA list design of the Pamela Ci's body's 16S rDNA list in the bacteria taxonogy, this unusual probe crosses and combines with the Pamela Ci's body 16s rRNA of the Hence Kong waters slice up, and then enlarges signal through the antigen antibody reaction, and displays colour through enzyme reaction. This checking method can combine the pathogeny checking with its pathology together; it can analyze the infection rate and infection intension on the stylebook slice when effiently and correctly checking the Pamela Ci's body; and the checking result is sensitive and precisious; and the slice after being checked can be preserved perennially.

Antibody / signal-generating moiety conjugates are disclosed that include an antibody covalently linked to a signal-generating moiety through a heterobifunctional polyalkyleneglycol linker. The disclosed conjugates show exceptional signal-generation in immunohistochemical and in situ hybridization assays on tissue sections and cytology samples. In one embodiment, enzyme-metallographic detection of nucleic acid sequences with hapten-labeled probes can be accomplished using the disclosed conjugates as a primary antibody without amplification.

A computer controlled method for detecting and diagnosing a rare cell type in a tissue sample is provided, said method comprising treating the tissue sample such that it generates a first signal indicative of the presence at a location of a rare cell, detecting the first signal, treating the location at which the first signal is detected to generate a second signal indicative of a diagnostically useful cellular characteristic and detecting the second signal. The first signal can be morphological or a color present in a sought cell either before or after staining. The second signal can be generated by in situ PCR or PCR in situ hybridization. In one preferred embodiment, the rare cell type is fetal cell in a maternal blood tissue sample, said sample consisting of a smear of unenriched maternal blood. In another embodiment, the method is used to diagnose or genotype cancer cells in a blood or tissue biopsy sample.

The present invention provides reagents for use in an automated environment for cell conditioning of biological samples wherein the cells or tissues are predisposed for access by reagent molecules for histochemical and cytochemical staining procedures. The reagents comprise components optimized to faciliate molecular access to cells and cell constituents within the biological sample. The present invention also provides reagents for use in an automated environment for removing or etching embedding media by exposing a biological sample to be stained in histochemical or cytochemical procedures without the dependence on organic solvents. The reagents comprise components optimized to facilitate removal or etching of the embedding media from the biological sample.

This invention relates to methods for the detection of one or more mRNA transcripts in paraffin-embedded tissue by “mRNA liberation in fixed-treated tissue or ‘MLIFTT’”. This method includes treating the tissue with ammonia-ethanol and sodium borohydride combined with pressure cooking of the tissue. The chemical treatments reduce the tissue autofluorescence and the physical treatments overcome the interference created by the fixation-induced chemical bonds. The methods of the present invention can be utilized to identify a plurality of mRNA transcripts in a microarray format.

The present invention provides methods of detecting chromosomal or genetic abnormalities associated with various diseases or with predisposition to various diseases. In particular, the present invention provides advanced methods of performing DNA hybridization, capture, and detection on solid support. Invention methods are useful for the detection, diagnosis, predicting response to therapy, detecting minimal residual disease, prognosis, or monitoring of disease treatment or progression of particular disease conditions such as cell proliferative disorders

Provided is an HPV E6, E7 mRNA assay, referenced herein as the “In Cell HPV Assay,” that is capable of sensitive and specific detection of normal cervical cells undergoing malignant transformation as well as abnormal cervical cells with pre-malignant or malignant lesions. The In Cell HPV Assay identifies HPV E6, E7 mRNA via in situ hybridization with oligonucleotides specific for HPV E6, E7 mRNA and quantitates the HPV E6, E7 mRNA via flow cytometry. The In Cell HPV Assay can be carried out in less than three hours directly from liquid-based cervical (“LBC”) cytology specimens. The In Cell HPV Assay provides an efficient and highly sensitive alternative to the Pap smear for determining abnormal cervical cytology.

A fluorescent in situ hybridization method including the steps of (a) obtaining a chromosome spread of a species; (b) preparing a hybridization composite containing a plurality of chromosomal paints each of the plurality of chromosomal paints being labeled with a different fluorophore-or-combination-of-fluorophores, such that an averaged specific activity of highly repetitive sequences in the hybridization composite substantially equals an averaged specific activity of unique sequences in the hybridization composite; (c) denaturing the hybridization composite and subjecting the hybridization composite to conditions for allowing at least a part of the highly repetitive sequences in the hybridization composite to reanneal while at least a part of the unique sequences in the hybridization composite remaining single stranded; (d) contacting under hybridization conditions the hybridization composite with the chromosome spread; (e) washing away excess of the hybridization composite; and (d) analyzing and presenting images of the now hybridized chromosome spread.