SEARCH · PubMed Health

Results for “Paraffin Embedding”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 919 records · Page 51Linked to original sources

PubMed2004

Analysis of p53 mutations in cells taken from paraffin-embedded tissue sections of ductal carcinoma in situ and atypical ductal hyperplasia of the breast.

Mutations in the p53 tumor suppressor gene are frequent in breast tumors but the implication of p53 mutations in breast cancer development remains poorly understood. In this study, we applied laser capture microdissection (LCM) microscope to histologically review and sample cells from paraffin-embedded breast tissue sections obtained from six cases of ductal carcinoma in situ (DCIS) and ten cases of atypical ductal hyperplasia (ADH). p53 mutations were detected, using single stranded conformational polymorphism (SSCP) and sequencing, in cell samples of three cases with DCIS and five cases with ADH. p53 mutations are therefore present in DCIS and ADH of the breast, considered as pre-malignant precursors to breast cancer, and some of them may represent early events in breast cancer development.

Breast↗

PubMed1994

Detection of the Philadelphia chromosome in paraffin-embedded tissue by fluorescence in situ hybridization.

The bcr-abl fusion gene situated on the Philadelphia chromosome is a tumor-specific marker for chronic myelogenous leukemia. We evaluated the usefulness of two color fluorescence in situ hybridization with bcr and abl probes as a means of detecting the Philadelphia chromosome in formalin-fixed, paraffin-embedded sections of spleen and lymph node specimens from eight patients with myeloproliferative diseases showing clinical and morphological features of chronic myelogenous leukemia in accelerated phase. Our analysis showed co-localized hybridization signals corresponding to the bcr-abl fusion product in tissue sections from six patients previously found to have the Philadelphia chromosome by conventional cytogenetics and polymerase chain reaction. The two remaining specimens lacked bcr-abl fusion signals and were obtained from patients who were negative for the Philadelphia chromosome by cytogenetic and polymerase chain reaction analysis. We conclude that fluorescence in situ hybridization is a sensitive method for the detection of the bcr-abl fusion gene in histological specimens from patients with chronic myelogenous leukemia. The technique may become a useful tool in the evaluation of tissue specimens from patients with chronic myelogenous leukemia and related Philadelphia chromosome-positive hematologic malignancies.

Fusion Proteins, bcr-abl↗

PubMed2006

Incestuous paternity detected by STR-typing of chorionic villi isolated from archival formalin-fixed paraffin-embedded abortion material using laser microdissection.

Microscopic examination of a blood clot expelled by a physically and mentally disabled woman taken to the emergency room because of genital bleeding revealed the presence of chorionic villi encircled by decidua, hemorrhage, and necrosis. In order to identify the father of the product of conception, sections of formalin-fixed, paraffin-embedded abortion material were subjected to laser microdissection: DNA extraction from chorionic villi selectively isolated from the surrounding tissues allowed successful STR-typing of fetal cells, which was otherwise prevented by excess maternal DNA. The large number of homozygous genotypes in the fetal profile suggested incestuous paternity. Analysis of reference DNA samples from male relatives excluded the woman's father, paternal grandfather, and maternal grandfather, whereas the obligate paternal alleles of the fetus were constantly present in the genotypes of the woman's brother, clearly demonstrating brother-sister incest (probability of paternity > 99.99999%).

Aborted Fetus↗

PubMed1994

Detection of hepatitis C virus genome in formalin-fixed paraffin-embedded liver tissue by in situ reverse transcription polymerase chain reaction.

Detection of hepatic hepatitis C virus RNA and antigens is difficult since their expression is very low. The technique of in situ reverse transcription polymerase chain reaction (IS-RT-PCR) was developed for the detection and localization of HCV RNA in formalin-fixed paraffin-embedded liver sections. Key steps for achieving a good signal-to-background ratio include appropriate proteolytic digestion, DNase pretreatment, higher Mg2+ concentration, primers from the core region, "hot-start", and between 10-20 cycles of PCR. Using this approach, HCV RNA was detected in the liver in four of the six patients. In those positive samples, the percentage of cells positive for HCV RNA ranges from < 5 to 25%. Using RT-PCR of serum samples and RNA extracted from the corresponding tissue block (10 sections) as the gold standard and sections from four control subjects as negative controls, the sensitivity and specificity of this assay was 4/6 and 4/4, respectively. The specific positive signal was found mainly in the cytoplasm of the hepatocytes and occasionally in mononuclear cells but not in the bile duct epithelium or sinusoidal cells. These positive hepatocytes were scattered in the liver lobules with occasional clustering. These preliminary data confirm the hepatotropic nature of HCV and the localization of HCV RNA is consistent with cytoplasmic replication.

Base Sequence↗

PubMed1989

Flow DNA-cytometric findings of paraffin-embedded primary cutaneous melanomas related to prognosis.

In a retrospective study flow cytometry was performed in 82 paraffin-embedded clinical stage I cutaneous melanomas. A significant correlation of ploidy to melanoma thickness was found. Using a log rank test ploidy was found significant for prognosis. However, using the Cox's proportional hazard test the influence of ploidy vanished and melanoma thickness or Clark level of invasion together with sex and location were the factors most important for prognosis.

Adult↗

PubMed2006

Accuracy of BRCA1 and BRCA2 founder mutation analysis in formalin-fixed and paraffin-embedded (FFPE) tissue.

BACKGROUND: A major limitation in counseling unaffected women from families with inherited breast and ovarian cancer is that a "true-negative" interpretation of wild type BRCA analysis of the proband cannot be inferred in the absence of demonstration of a BRCA mutation segregating in the kindred. Documentation of familial BRCA mutations from paraffin-derived DNA of deceased patients has been limited due to reports of technical complications leading to lack of reproducibility of BRCA testing of archival material. METHODS: DNA was extracted from formalin-fixed paraffin-embedded (FFPE) morphologically normal tissue of 161 blinded, coded samples from women previously genotyped for the three Ashkenazi Jewish BRCA founder mutations from lymphocyte-derived DNA. Multiplex PCR followed by denaturing polyacrylamide gel electrophoresis was performed for the three founder mutations to determine if analysis on FFPE tissue could produce results concordant with those of the lymphocyte-derived DNA. RESULTS: After disclosure of the sample codes, the results were compared with the original lymphocyte-derived DNA genotypes. Excluding one sample unevaluable due to PCR failure, there was 100% concordance of 160 genotypes (120 mutation samples) derived from DNA from archival FFPE tissue compared to peripheral lymphocytes. CONCLUSIONS: The method described reliably detected BRCA founder mutations in archival DNA derived from FFPE tissue. These results suggests that this technique may be useful in clinical settings to inform wild type BRCA results of unaffected probands, leading to avoidance of unnecessary intensified surveillance or risk-reducing surgery. With further validation this approach can also be applied to other populations where founder mutations are observed.

DNA↗

PubMed1998

Immunohistochemical evaluation of estrogen and progesterone receptors in paraffin-embedded, formalin-fixed endometrial tissues: comparison with enzyme immunoassay and immunohistochemical analysis of frozen tissue.

Monoclonal antibody (MoAb) 1D5 with specificity to the estrogen receptor (ER), MoAb 1A6, and a polyclonal antibody (PoAb) (the latter two with specificity to the progesterone receptor [PgR]) were used to stain microwave-pretreated sections of formalin-fixed, paraffin-embedded normal and malignant endometrial tissues (n = 60). The tissues were previously evaluated for ER and PgR by enzyme immunoassay (EIA) (n = 44) and immunohistochemical analysis of frozen tissue (ICAfroz, n = 59). With results of EIA as a reference, the ER-1D5 method yielded a better agreement on receptor status, i.e., positive versus negative (74 vs. 51%) and a higher sensitivity (71 vs. 45%) but a similar high specificity (100%) than the ER-ICA method. Compared with results of PgR-EIA, the immunohistochemical assays for PgR gave similar results as to agreement (86-95%) and sensitivity (95-97%). Quantitative agreement on the fraction of cells stained for ER and PgR by immunohistochemical analysis in frozen and formalin-fixed tissue was obtained in approximately 60% of the cases. The results of semiquantitation were correlated with the results of both ICA and EIA. The MoAbs 1D5 and 1A6, as well as the anti-PgR PoAb, thus seem to be valid for evaluation of ER and PgR status in formalin-fixed endometrial tissue. Differences in the specificity of the Abs and in the sensitivity of the methods used to demonstrate ER and PgR might explain some of the discordant findings.

Carcinoma↗

PubMed2005

Whole genome SNP arrays using DNA derived from formalin-fixed, paraffin-embedded ovarian tumor tissue.

Array-based genotyping platforms, such as the Affymetrix mapping array, have been validated as reliable methods for obtaining high-resolution copy number and allele status information when using DNA derived from fresh tissue sources. However, the suitability of such systems for the examination of DNA derived from formalin-fixed, paraffin-embedded (FFPE) tissues has not been tested. Therefore, we analyzed DNA derived from five matching fresh frozen and FFPE ovarian tumors for gene copy number changes and loss of heterozygosity using the Affymetrix GeneChip Human Mapping 10 K Array Xba 131. The data was analyzed using Affymetrix proprietary software, GeneChip DNA Analysis Software, and Chromosome Copy Number Tool. The average SNP call rate (rate of successful genotype identification) of the fresh samples was 89.44% (range 78.72-96.22%, median 92.72%) and was only slightly lower for the matching FFPE samples at 83.48% (range 76.93-93.17%, median 82.60%). The average concordance (rate of agreement between successful genotype calls) between the fresh and matching FFPE samples was 97.06% (range 92.70-99.41%, median 97.52%). Loss of heterozygosity (LOH) profiles of the fresh and FFPE samples were essentially identical across all chromosomes. Copy number data was also comparable, although the quantification of copy number changes appears overstated in the FFPE samples. In conclusion, we have shown that it is possible to achieve high-performance outcomes using FFPE-derived DNA in the Affymetrix 10 K mapping array. This advance will open up vast archival tissue resources to high-resolution genetic analysis and unlock a wealth of biological information.

DNA↗

PubMed2006

Effects of fixative type and fixation time on the detection of Maedi Visna virus by PCR and immunohistochemistry in paraffin-embedded ovine lung samples.

In doubtful cases, the histopathological diagnosis of lesions induced by Maedi Visna virus (MVV), a chronic multisystemic lentiviral disease of sheep, needs to be confirmed by the demonstration of MVV in the tissues. The influence of fixatives and the duration of fixation on the detection of MVV by immunohistochemistry (IHC) and PCR in paraffin-embedded tissues was assessed in lung samples with lesions in different degree, from five sheep serologically positive. Samples were fixed in 10% neutral buffered formalin (NBF), Bouin's solution (BS) and a zinc salts-based fixative (ZSF), for different periods of time between 24 h and 30 days. The three fixatives preserved the morphology of the tissues, although in ZSF-fixed samples an increase in the number of desquamated cells was seen in the alveoli. Tissues showed a similar degree of immunolabelling, irrespective of the duration of fixation using ZSF and NBF fixatives. MVV nucleic acids could be detected in samples fixed up to 14 days in NBF and 30 days in ZSF. However, in BS fixed tissues, immunostaining was weak and non-specific signals were observed after 4 days of fixation. Amplification of proviral DNA could not be obtained by PCR in these samples. IHC detected viral antigens in all sheep whereas one sheep with mild lesions was always negative by PCR.

Acetic Acid↗

PubMed2004

Improved immunocytochemical identification of neural, endothelial, and inflammatory cell types in paraffin-embedded injured adult rat spinal cord.

Methods that facilitate the accurate counting of specific neural cell types would be of substantial value in evaluating the efficacy of treatments applied to spinal cord injury. This report describes reliable procedures for identification of neurons, oligodendrocytes, astrocytes, endothelial cells and inflammatory cells (neutrophils and activated macrophage/microglial cells) in paraformaldehyde-fixed, paraffin-embedded injured adult rat spinal cord. Antigen retrieval techniques (enzymatic and thermal) were used to improve antibody access to masked epitopes. To decrease background immunofluorescence and autofluorescence of hemoglobin, the tissue sections were pretreated with 0.1% sodium borohydride in PBS (30min), followed by 1-5min incubation in 0.5% Sudan black in 70% ethanol. Commercially available techniques to amplify the primary signal such as tyramide signal amplification (TSA) and avidin/biotin/peroxidase/DAB/nickel/cobalt amplification (ABP/DABA) were also tested. Hoechst 33342 nuclear staining was used to indicate cell location, number, and integrity, thereby avoiding misidentification of cells. The best antibodies were: anti-NeuN antibody for neurons, anti-S100 for astrocytes, and anti-S100 and APC-7 antibodies in combination for oligodendrocytes, anti-laminin (LN) for endothelial cells, and ED1 antibody for activated macrophages and microglia. Amplification of the primary signal with TSA or ABP/DABA was also found to be beneficial.

Animals↗

PubMed1995

Determination of Ki-67 defined growth fraction by monoclonal antibody MIB-1 in formalin-fixed, paraffin-embedded prostatic cancer tissues.

The applicability of MIB-1, a monoclonal antibody directed against the Ki-67 antigen, was studied in the PC-82 and LNCaP prostatic tumor models at various levels of proliferative activity. Statistically significant correlations were found in LNCaP cultures between Ki-67 and MIB-1 scores (r = 0.84, P < 0.001), and in PC-82 tumors between MIB-1 scores and paraffin tissue Ki-67 (pKi-67) (r = 0.90, P < 0.001), frozen tissue (fKi-67) (r = 0.86, P < 0.001), and BrdU uptake (r = 0.70, P < 0.001), respectively. pKi-67 scores were double the fKi-67 scores, which may be due to methodological differences. MIB-1 scores exceeded both the fKi-67 and pKi-67 scores. The affinity of MIB-1 for the antigen is much higher than the affinity of Ki-67, which may explain the differences. MIB-1 is a promising means of evaluating the presence of only minute amounts of the Ki-67 antigen in paraffin-embedded human tumor material, especially in relatively slowly growing tumors.

Adenocarcinoma↗

PubMed2002

The value of anti-pax-5 immunostaining in routinely fixed and paraffin-embedded sections: a novel pan pre-B and B-cell marker.

Whereas L26 (anti-CD20) is well established as a B-cell marker of high specificity for use in paraffin-embedded tissues and JCB117 (anti-CD79a) is increasingly used, a comparable additional pan-B-cell antibody has hitherto not yet been identified. Here we have studied the use of a novel anti-pan-B-cell marker Pax-5 for use in diagnostic pathology. Pax-5 encodes for BSAP (Pax-5), a B-cell-specific transcription factor, the expression of which is detectable as early as the pro-B-cell stage and subsequently in all further stages of B-cell development until the plasma cell stage where it is downregulated. Pax-5 is essential for B-lineage commitment in the fetal liver, whereas in adult bone marrow this transcription factor is required for progression of B-cell development beyond the early pro-B (pre-BI) cell stage. Among the B-cell genes that are present in early B-cell development and are upregulated by Pax-5 are CD19 and Igalpha (CD79a). We have tested a commercially available anti-Pax-5 antibody (anti-BSAP, clone 24) in a series of 592 routinely fixed and paraffin wax-embedded biopsies, including lymph nodes, bone marrow, and various other organs containing lymphoid tissues. Pax-5 protein (BSAP) was detected in all cases of precursor and mature B-cell non-Hodgkin lymphomas/leukemias. In addition, in 97% of classic Hodgkin lymphomas, Reed-Sternberg cells expressed Pax-5. However, Pax-5 was not detected in any of the multiple myelomas, solitary plasmacytomas, and 4% of diffuse large B-cell lymphomas. Among those diffuse large B-cell lymphomas not expressing Pax-5 were only those with terminal B-cell differentiation. All T-cell non-Hodgkin lymphomas, including ALCL and lymphoblastic lymphomas and leukemias, were negative. There was a strong association between Pax-5 and CD20 expression. We conclude that anti-Pax-5 is an excellent pan-B and pan-pre-B-cell marker. We have found that anti-Pax-5 is superior to anti-CD20 in the diagnosis of pre-B acute lymphoblastic leukemia and classic Hodgkin lymphoma versus ALCL of T and "null" cell type. It was also useful in differential diagnosis between lymphoplasmacytic lymphoma and plasmacytoma. Even though there is an excellent correlation between CD20 and Pax-5 expression, anti-Pax-5 exceeds the specificity and sensitivity of L26 (anti-CD20) because of its earlier expression in B-cell differentiation and its ability to detect all committed B cells, including classic Hodgkin lymphoma.

Antibodies, Monoclonal↗

PubMed1994

Cathepsin D immunostaining in paraffin-embedded breast cancer cells and macrophages: correlation with cytosolic assay.

High cathepsin D (cath-D) concentration in breast cancer cytosol is associated with increased risk of metastasis. To specify the relative contribution of the different cells types responsible for cath-D level in cytosol, we validated semiquantitative cath-D immunoperoxidase staining on formalin-fixed, paraffin-embedded sections, using the M1G8 monoclonal antibody, one of the two antibodies of the cytosolic assay. Using computer-aided image analysis, cath-D level in cancer cells was estimated by integrating both staining intensity in each cell and proportion of stained cells. We confirmed on 41 primary breast cancers a higher expression of cath-D in cancer cells compared with peritumoral mammary glands. Cancer cell staining was mostly in lysosomes and for some invasive ductal carcinomas in large vesicles corresponding to phagosomes. Lymphocytes and fibroblasts were not or were only weakly stained. Macrophages also were stained for cath-D, generally on the periphery of the tumor area. The cytosolic cath-D level was correlated with cath-D expression in cancer cells (r = .76; P = 1 x 10(-4)) rather than with the number of macrophages in the tumor (r = .29; P = .09), as determined by use of the specific anti-CD68 antibody. There was a significant increase in the tissue cath-D level in tumors containing large vesicles compared with tumors without large vesicles. This approach provides a means to separately estimate the prognostic significance of cath-D expression in cancer cells and macrophages when evaluating risk of metastasis.

Breast Neoplasms↗

PubMed1994

Detection of ptc in archival formalin-fixed, paraffin-embedded tissues. Comparison of radiolabeled DNA hybridization and direct incorporation of digoxigenin-11-dUTP into RT-PCR products.

Archival pathological specimens are a source of RNA and DNA for clinical surveillance or retrospective studies. We employed a modification of the acid guanidium thiocyanate-phenol-chloroform extraction method for the recovery of total RNA from formalin-fixed, paraffin-embedded neoplastic thyroid tissue. The extracted RNA was used for reverse transcription of ptc and subsequent amplification of the complementary DNA (cDNA) by the reverse transcription-polymerase chain reaction (RT-PCR). In lieu of 32P-labeled DNA for hybridization studies, we supplemented the nucleotide pool in the amplification reaction with a modified pyrimidine, digoxigenin-11-dUTP. Digoxigenin-11-dUTP was incorporated directly into the PCR product, eliminating the need for hybridization, posthybridization washes, and prolonged autoradiography. These products were resolved by electrophoresis on agarose gels, Southern blotted to nylon membranes, and rapidly detected by chemiluminescence. This nonradioisotopic method has expedited and reduced the cost for molecular investigations with archival pathological specimens by providing equal sensitivity to or greater sensitivity than that of DNA-labeled radionuclides without the associated biological hazards.

Carcinoma, Papillary↗

PubMed2005

Reliable high-throughput genotyping and loss-of-heterozygosity detection in formalin-fixed, paraffin-embedded tumors using single nucleotide polymorphism arrays.

Most human cancers show genetic instabilities leading to allelic imbalances, including loss of heterozygosity (LOH). Single nucleotide polymorphism (SNP) arrays can be used to detect LOH. Currently, these arrays require intact genomic DNA as obtained from frozen tissue; however, for most cancer cases, only low-quality DNA from formalin-fixed, paraffin-embedded (FFPE) tissue is available. In this study, we tested Illumina BeadArrays to genotype FFPE tissue and detect LOH/allelic imbalances in matched colorectal tumor and normal tissue. Genotypes were compared between leukocyte and FFPE normal tissue as well as between frozen and FFPE tumor tissue. Identical genotypes and LOH profiles were obtained from normal and tumor isolates. LOH was mainly observed on chromosomes 4, 5q, 12q, 14q, 15q, 17p, 18, and 20p, which are commonly detected regions in colorectal cancer. LOH profiles of the BeadArrays were compared with profiles obtained by Affymetrix GeneChip 10K arrays, showing identical LOH patterns. These data show that genome-wide genotyping of FFPE tissue with the BeadArray gives reliable results and is a powerful technique for LOH analysis.

Colorectal Neoplasms↗

PubMed2006

Improved RT-PCR amplification for molecular analyses with long-term preserved formalin-fixed, paraffin-embedded tissue specimens.

Recently, in addition to DNA, RNA extracted from archival tissue specimens has become an invaluable source of material for molecular biological analysis. Successful amplification with PCR/RT-PCR is problematic when using amplicons of short size due to degradation of DNA or RNA. We established an improved method for efficient RT-PCR amplification of RNA extracted from archival formalin-fixed, paraffin-embedded tissue by the elimination of RNA modification and the restoration of RNA template activity. Namely, the preheating in citrate buffer (pH 4.0) of RNA extracted from long-term preserved tissue specimens resulted in significantly increased efficiency of RT-PCR.

Buffers↗

PubMed1997

Epstein-Barr virus infection, Hodgkin's disease, non-Hodgkin's lymphoma, and reactive follicular hyperplasia in Japanese children: evaluation of paraffin-embedded specimens using polymerase chain reaction and immunohistochemistry.

Epstein-Barr virus (EBV) infections are common in Japanese children, with infectious by EBV type 1. The relationships between EBV infection and lymphadenopathies in Hodgkin's disease (HD), non-Hodgkin's lymphomas (NHL), reactive follicular hyperplasia (RFH), and infectious mononucleosis (IM) in 37 Japanese children were evaluated. Formalin-fixed, paraffin-embedded lymph node specimens that were obtained at surgical resection or biopsy were evaluated for the presence of EBV DNA and the latent membrane protein-1 (LMP-1) using polymerase chain reaction (PCR) and immunohistochemical staining. The PCR detected EBV DNA in nine of 13 (69.2%) patients with RFH, including a case of IM, all three (100%) patients with HD, and one of 21 (4.8%) patients with NHL. All EBV-positive samples contained EBV type 1. Reed-Sternberg's cells in HD were immunohistochemically positive for LMP-1, whereas all cases of RFH and NHL were negative for LMP-1. Results suggest that EBV infection may be related to HD. Although no proof exists that EBV infection contributes to the transformation of cells, thus causing RFH or NHL, the present authors suggest that the EBV-positive cases in Japanese children demonstrate a relationship between the clinical and histopathological features of the lymphadenopathy and EBV-type 1 infection.

Child↗

PubMed2004

FISH analysis of BRCA1 copy number in paraffin-embedded ovarian cancer tissue samples.

We investigated the BRCA1 gene copy number in unselected ovarian malignancies. Both additional genes (amplification) as well as deletion (loss of heterozygosity, LOH) are often thought to have a role in the initiation or progression of cancer. In addition, if there were little change, deletion studies might help identify BRCA1 mutation carriers. Forty-seven paraffin-embedded ovarian tissue blocks obtained between 1984 and 1997 were used for this study. A sample was "deletion-positive" when BRCA1-deleted cells in the tumor area were significantly different from the benign area. Twenty-five (53%) cases were found to be "deletion-positive". The average age of onset of "deletion-positive" patients was 50.8 years and of "deletion-negative" 57.8 years (P < 0.05). There was no statistical difference between groups in the staging, histology, or prognosis. A Kaplan-Meier study did show a trend towards poorer survival for "deletion-positive" patients. FISH permits unique molecular characterization of malignancies at a cellular level. Double amplification of HER-2 and c-myc predicts poor ovarian cancer survival. There appears to be a definite role for BRCA1 deletion in reducing the age of ovarian cancer onset and possibly in overall survival. Further FISH studies of this and other patient sets using additional molecular markers are needed.

Adult↗