79 results about "Formalin fixed paraffin embedded" patented technology

The present invention provides novel methods for analyzing gene expression levels from fresh or aged (more than one year old) formalin-fixed, paraffin-embedded tissue (“FFPET”) samples that comprise pre-hybridizing a labeled nucleic acid sample prepared from the formalin-fixed, paraffin-embedded tissue sample with a first microarray, hybridizing the unbound labeled nucleic acid sample with a second microarray, and detecting the labeled nucleic acid sample bound to the second microarray. The pre-hybridization step results in an increase in the specific gene signals in subsequent hybridizations with high density gene expression arrays. The first microarray used for the pre-hybridization step can be either a new or used microarray. Importantly, from a cost-savings perspective, the inventors determined that when the first microarray used for the pre-hybridization step is a previously used microarray, the results of the subsequent hybridization on a second microarray are nearly identical to the results obtained when the pre-hybridization was carried out using a new or previously unused microarray.

The present invention relates to the field of cancer. More specifically, the present invention provides methods and compositions useful for diagnosing or predicting cancer in a patient. In one embodiment, a method for identifying a patient as having cancer comprises the steps of (a) providing a formalin-fixed, paraffin-embedded or fresh frozen sample of patient tissue; (b) steaming the sample in antigen retrieval buffer; (c) incubating the sample in hydrochloric acid (HCl); (d) incubating the sample with an affinity reagent specific for 5hmC under conditions to form a complex between the affinity reagent and 5-hydroxymethylcytosine (5hmC) present in the sample; (e) detecting the complexes formed between 5hmC and the affinity reagent with secondary detection reagents; (f) quantifying 5hmC levels; and (g) identifying the patient as having cancer if the 5hmC levels in the sample are reduced as compared to a control.

The present teachings provide methods, compositions, and kits for detecting micro RNAs (miRNAs). In some embodiments, the miRNAs are quantified from formalin-fixed paraffin-embedded tissue samples in which messenger RNA is degraded. The present teachings take advantage of the observation that most mature miRNAs in vivo are protected by degradation as a result of their association with RISC. Thus, novel methods of studying nucleic acids in archived tissues containing degraded messenger RNA are provided, wherein RISC-protected miRNAs are liberated, and analyzed.

The invention discloses an FFPE reference product for gene detection and a preparation method and application of the FFPE reference product. The preparation method comprises the following steps that S1, cell culture is performed on a tumor cell line, and cell pellets are collected; S2, formalin fixation is performed on the cell pellets, then sepharose gel wraps the cell pellets to form cell aggregates, and the cell aggregate are prepared into cell paraffin blocks; S3, genomic DNA of the tumor cell line in the cell paraffin blocks is extracted, and genetic mutation frequency determination is performed on the genomic DNA of the tumor cell line; S4, the genomic DNA of the tumor cell line containing a target mutation site is mixed into a DNA mixture of a target mutation frequency, and the DNAmixture is the FFPE reference product for gene detection. According to the technical scheme, formalin fixation and paraffin wrapping are performed on the cells cultured by tumor cell line, so that thesituation of clinical samples can be better simulated.

The invention relates to a method for extracting desoxyribonucleic acid from formalin fixed and paraffin embedded tissues, and belongs to the technical field of nucleic acid application in biology. The method comprises the following steps of: preparing special lysis solution, adding the lysis solution into paraffin section tissues, boiling at high temperature for 30 minutes, and centrifuging to take supernate; adding absolute ethanol for uniform mixing, and adding the mixed solution into a silicon membrane absorption column for centrifuging; rinsing by using protein-free liquid and salt-free rinsing liquid; and eluting by using elution buffer. In the method, a toxic reagent of dimethylbenzene is not used for dewaxing, and harm to bodies of experimenters is avoided; and precious protease Kis not needed to perform long-time incubation enzymolysis, the operation is simple and quick, the extracted desoxyribonucleic acid in genome is high in quality and stability, and the cost and time can be saved to the greatest degree. The extracted product is subjected to polymerase chain reaction (PCR) detection, long segments with about 750pb can be obtained through amplification, and the work in the aspects such as scientific research, biomedicine and the like is greatly facilitated.

The invention relates to imaging, such as in situ imaging by expansion microscopy, labelling, and analyzing biological samples, such as formalin fixed paraffin embedded (FFPE) cells and tissues, as well as reagents and kits for doing so.

The invention discloses a method for extracting total ribonucleic acid (RNA) from paraffin-embedded tissues, which is an improvement method provided on the basis of a kit. The method comprises the following steps: deparaffinating, digesting, extracting and purifying. By adopting the method, the RNA in a tissue sample stored by being embedded by paraffin can be extracted, wherein, the longest storage time reaches up to 6 years; the RNA is analyzed by a nucleic acid trace tester and an internationally recognized Agilent 2100bioanalyzer; the extraction concentration is 211-3116ng / mu l, and the extraction purity is A260 / 280:1.75-2.04, which are all very ideal; and the expression research of microarray high-abundance genes of qRT-PCR and miRNA in mRNA and miRNA level can be carried out, which provides an experimental basis for developing a large amount of research on formalin fixed paraffin-embedded (FFPE) genes, and has wide application value.

An ultra-sensitive, specific methodology for detecting PIK3CA mutations in biological samples of cancer patients, comprises a combination of allele-specific, asymmetric rapid PCR and melting analysis in a DNA sample from Circulating Tumor Cells, cell-free DNA in plasma / serum, or Formalin-Fixed Paraffin-Embedded tissues. Using the allele-specific primers for hotspot mutations in exons 9 and 20 (E545K and H1047R), detection can enhance amplification of mutant PIK3CA allele sequence, whereas presence of corresponding competitive blocking unlabeled probes for each exon can avoid non-specific amplification of wild-type PIK3CA sequence increasing the sensitivity and the specificity of method. The mutational detection is completed with melting curve analysis of the unlabeled probe and DNA template of the mutant PIK3CA sequence. Evaluation of PIK3CA mutational status on CTC in peripheral blood and cfDNA in plasma / serum of patients has potential for clinical applications and therapeutic interventions, since presence of PIK3CA mutations is associated with response to molecular targeted therapies.

The present invention relates to a system and method of genomic profiling and is particularly useful in genomic differentiation of heterogeneous and polyclonal neoplastic cell populations, preferably of flow sorted formalin fixed paraffin embedded samples. The present invention includes methods of improving resolution for identifying aberration in variable carcinoma genomes and / or heterogeneous cell populations. The present invention also includes kits configured to improve genomic resolution and the ability to identify genomic aberration in variable and / or heterogeneous cell populations.

A dissection system has a dissection platform which has a frame, an agitation platform, a tissue section tray, a solution dispenser unit, an airflow drying unit and a waste collection unit and a specimen collector which has a tubular body, a plunger button, a hollow shaft, a piston cylinder, a motor unit, a piston, a piston spring and a piston rod. The agitation platform, the solution dispenser unit, the airflow drying unit and the waste collection unit are disposed on the frame, and the tissue section tray is removably disposed on the agitation platform. The plunger button, the hollow shaft and the piston cylinder are movably inserted in the tubular body. The piston is slidably inserted within the piston cylinder, the piston spring is biased in between the piston cylinder and the motor unit, and the piston rod is connected in between the motor unit and the piston.

Methods and reagents are provided for the rapid extraction of nucleic acids from a cell or tissue sample. In certain embodiments the sample comprises a formalin fixed paraffin embedded sample (e.g., aFFPET sample), or a fine needle aspirate and / or a cell / tissue smear. In some embodiments, the methods comprise incubating one or more sections of said tissue sample in a lysis solution comprising a buffer sufficient to maintain the pH of said solution at a pH ranging from about pH 4 to about pH 9; a chaotropic agent; a chelating agent; and a detergent; where the incubating is at a temperature ranging from about 50 DEG C to about 100 DEG C; and recovering the nucleic acid from said lysis solution.

Disclosed is an efficient and scalable method for targeted resequencing and variant identification of nucleic acids such as genomic DNA found in single stranded, fragmented form, such as in a clinical sample of formalin-fixed, paraffin-embedded (FFPE) tissue. The method uses a large number of capture probes mixed with the sample in the presence of a 5′ to 3′ exonuclease, a 3′ to 5′ exonuclease, a ligase, and a universal amplification oligonucleotide that hybridizes to the various capture probes. The nucleases act on ssDNA, not dsDNA. A single stranded circle is formed by the ligase, and is then amplified to produce a population (library) of double stranded linear DNA molecules that are suitable for sequencing. It is shown that the library produces a high degree of fidelity to the original sample, and predictable base changes are shown.

The present invention relates to methods for the extraction of long fragment RNA from fixed tissue specimens. In particular, the present invention relates to methods for the extraction of RNA from formalin-fixed paraffin-embedded tissue specimens for use in biologic applications, including assays based on oligonucleotide hybridization.

The present invention provides for a mass spectrometry proteomic approach to distinguishing Spitz nevi from Spitzoid malignant melanoma. Histology directed mass spectral profiling allows for targeted analysis of sites of melanocytic lesion within formalin-fixed, paraffin embedded excisional biopsies. The classification system identified 5 peptide peaks, of which two have been identified as originating from vimentin and actin. A sensitivity and specificity for Spitz nevi of 97% and 90%, respectively, were achieved.

The invention provides a kit for co-separation of (deoxyribonucleic acid) and RNA (ribonucleic acid) from FFPE (formalin fixed and paraffin embedded tissues) as well as a method. The kit can be used for simultaneously extracting DNA and RNA from an FFPE sample, and has the advantages that not only can nucleic acid extraction be performed simultaneously, time is saved, and the extraction is economical and fast, but also the sample size can be saved, the problem of sample lack in a test can be solved effectively, and further, identical source of the DNA and the RNA used in study can be guaranteed. The kit and the method have the advantages that the operation is simple and direct, the repeatability is high, the yield is high, and enough DNAs and RNAs can be obtained simultaneously only through 2-5 pieces of tumor tissue sections; the obtained nucleic acids are high in purity and has no interference to follow-up detection; and compared with the common kit, the kit has remarkable advantages.

The invention provides a method for deparaffinizing a formalin-fixed paraffin-embedded tissue, including: providing a formalin-fixed paraffin-embedded tissue sample; mixing the formalin-fixed paraffin-embedded tissue sample with an organic solvent and water or with an organic solvent and an aqueous solution to form a mixture, wherein a density of the organic solvent is less than that of the water or the aqueous solution, and the organic solvent is immiscible with the water or the aqueous solution; and separating the mixture into an organic solution layer and an aqueous solution layer, wherein a paraffin dissolved from the formalin-fixed paraffin-embedded tissue sample is in the organic solution layer and a deparaffinized tissue from the formalin-fixed paraffin-embedded tissue sample is in the aqueous solution layer and / or an interlayer between the organic solution layer and the aqueous solution layer.

The invention provides a specimen pretreatment method for reducing auto-fluorescence interference in FISH (fluorescence in situ hybridization) and a reagent kit therefor. The specimen pretreatment method comprises a first step of formalin fixation of the paraffin-embedded tissue, a second step of sectioning and drying, a third step of functioning of sodium thiocyanate, a fourth step of addition of 20[mu]l of collagenase type IV with the concentration of 5-10 mg / mL or elastinase with the concentration of 1mg / mL in the tissue with the area of 24*32mm and post-sealing functioning at the temperature of 37 DEG C for 0.5-1hr for degradation of the extracellular matrix with the auto-fluorescence characteristics, and a fifth step of functioning of pepsin at the temperature of 37 DEG C. The invention also provides a reagent kit for the specimen pretreatment method for reducing auto-fluorescence interference in FISH (fluorescence in situ hybridization). Through the specimen pretreatment based on the invention, the auto-fluorescence phenomena of the tissue is remarkably reduced, the ratio of fluorescent probe signals to background fluorescence is increased, and the detection quality and accuracy of FISH are improved.

The invention provides a kit for detecting gene variation in a tumor FFPE (formalin-fixed paraffin-embedded) sample. The kit comprises probes for gene trapping, and the probes for gene trapping include a probe designed for an SNV (single nucleotide variant) area, a probe designed for a CNV (copy number variation) area, and a probe designed for a FUSION area; public primers include nucleotide sequence primers shown in SEQ ID NO:1; and tag primers include nucleotide sequence primers shown in SEQ ID NO:2-SEQ ID NO:5. According to the invention, in a method for detecting gene variation in the tumor FFPE sample, the genes SNV, CNV and FUSION can be stably detected at the same time.

The present invention relates to: a method for separating nucleic acids from a sample containing formalin-fixed, paraffin-embedded (FFPE) tissue sections; a nucleic acid separation kit; and a lysis buffer for separating nucleic acids.

Methods are disclosed for isolating nucleic acids from formalin-fixed paraffin embedded (FFPE) tissue samples. Each of tissue samples contains paraffin and a target biological tissue or material, and the method includes the steps of: adding a first reagent and a second reagent to the FFPE tissue sample, the first reagent dissolving the paraffin material and the second reagent lysing the biological tissue; mixing the first reagent, the second reagent, and the FFPE tissue sample to form a first mixture; (2) heating the first mixture at 50-80° C. for 30-90 minutes; and then heating the first mixture at 80-95° C. for 30-90 minutes to fractionize the first mixture to form an aqueous phase and an oil phase; (3) collecting an aqueous solution from the aqueous phase; and (4) isolating nucleic acids from the aqueous solution. The method improves the efficiency and convenience of isolating nucleic acids from FFPE tissue samples.

Formalin fixation causes cross-linkage between nucleic acids and proteins and covalently modifies RNA. As a result, the molecules are rigid and may comprise subsequent RNA extraction. The invention provides a method for recovering RNA from formalin fixed paraffin-embedded tissue, including a short additional step of incubation with proteinase K after the first digestion step that makes a significant enhancement of the quality and quantity of the extracted RNA and subsequently, an improvement in the detection of gene expression is achieved. The method of the invention has the advantage of minimizing the number of manipulations, eliminating the need for potentially toxic solvents, and increasing significantly the amount of RNA recovered, and therefore the sensibility, when compared with previous methods.

A microfluidic apparatus, method, and associated applications utilize and apply to a formalin-fixed paraffin-embedded (FFPE) tissue sample and performing a liquid-liquid extraction to remove the paraffin from the tissue sample prior to a nucleic acid purification step. A microfluidic device includes a dedicated liquid-liquid extraction process vessel, a nucleic acid purification process component, and a nucleic acid amplification reactor. A liquid-liquid extraction and nucleic acid purification kit includes a microfluidic device capable of performing both a liquid-liquid extraction process and a nucleic acid purification process, including a dedicated liquid-liquid extraction process vessel, an immiscible liquid or a precursor phase thereof disposed in the vessel, a nucleic acid purification process component, a nucleic acid amplification reactor fluidically, and a supply of reagents suitable to enable the liquid-liquid extraction process and the nucleic acid purification process.

Disclosed are detergent-free protein extract buffers to extract proteins from formalin-fixed paraffin-embedded tissue sections, kit comprising the buffer, and methods for using the buffer to extract proteins from formalin-fixed, paraffin-embedded tissue sections.

The present invention relates to methods for the extraction of long fragment RNA from fixed tissue specimens. In particular, the present invention relates to methods for the extraction of RNA from formalin-fixed paraffin-embedded tissue specimens for use in biologic applications, including assays based on oligonucleotide hybridization.

Pretreating a tissue sample with an effective amount of collagenase results in increased immunodetection of collagen and enhanced preservation of tissue morphology. Such an improved method for immunodetection of collagen can be used in immunohistochemical studies of formalin-fixed, paraffin-embedded tissue sections.

The present disclosure provides methods, arrays and kits for assessing the quality of genomic DNA samples, especially those obtained from formalin-fixed paraffin-embedded (FFPE) samples. The methods, arrays and kits provided herein use primer pairs specific to regions in the genomes of the organisms from which genomic DNA samples are obtained that have identical or nearly identical copies distributed across multiple chromosomes.

The present invention concerns an in vitro method for detecting the susceptibility of a tumor cell to a chemotherapy, said method comprising the step of the measurement of the ERCC1 protein by immunohistochemistry in a formalin-fixed paraffin-embedded tumor sample.

The present invention provides specific nucleic acid sequences for use in the identification, classification and diagnosis of various sub-types of lung cancers. The present invention permits one to accurately classify lung cancers based on their miR expression profile without further manipulation. Using microRNA microarray data generated from over two hundred formalin-fixed paraffin-embedded (FFPE) resection samples, fine needle aspiration (FNA) samples and fine needle biopsy (FNB) samples of primary lung cancer, microRNA expression profiles were identified that differ significantly for various sub-types of lung cancer.

The invention discloses a method for constructing a circRNA high-throughput sequencing library of an FFPE (formalin-fixed and paraffin-embedded) sample. The method concretely comprises the following steps: (1) extracting total RNA from paraffin sections; (2) adding a PolyA end to the sample RNA; (3) capturing and separating linear RNA by Oligo dT magnetic beads; (4) further removing residual linear RNA; and (5) constructing the circRNA high-throughput sequencing library. The FFPE sample RNA has the characteristics of damages, low content and poor integrity, and the method for constructing thecircRNA high-throughput sequencing library of the FFPE sample has the advantages of simplicity in operation, short time, low cost, stableness in obtaining the library meeting high-throughput sequencing requirements, and avoiding of the problems of large sample size, high rRNA removal cost and low efficiency in the traditional library constructing methods.