AIMS: Formalin-fixed paraffin-embedded (FFPE) samples, routinely used in neuropathology, represent an invaluable resource for studying rare diseases like transmissible spongiform encephalopathies (TSE). Despite fixation-induced protein cross-linking, prion seeding activity can be effectively detected using the seeding amplification assays. In this study, we employed the second-generation real-time quaking-induced conversion (RT-QuIC) assay to analyse and quantify human prion seeding activity in FFPE brain tissues. METHODS: FFPE frontal brain tissues were deparaffinised in xylene, followed by rehydration through descending concentrations of ethanol. The prion seeding activity in tissue homogenates was assessed by RT-QuIC assay utilising short recombinant hamster prion protein (rHaPrP90-231) as a substrate. RESULTS: A total of 60 samples, including 30 cases of confirmed TSE, comprising both sporadic and genetic forms, as well as 30 non-TSE controls, were analysed. Prion seeding activity has been detected in all TSE samples except one sCJD (VV2) and one GSS (P102L) case, corresponding to an assay sensitivity of 93.3%. Conversely, we did not detect any RT-QuIC positivity in the control group, resulting in 100% specificity. The mean 50% prion seeding dose of FFPE sporadic TSE samples was 107.8/g of brain tissue. CONCLUSION: Our study emphasises high sensitivity and specificity of RT-QuIC assay for prion detection in archival human FFPE brain tissues and demonstrates its diagnostic reliability comparable to other tissue types even after years of storage. The applicability of FFPE samples in RT-QuIC assays facilitates retrospective diagnostics and provides logistical advantages for sample preservation and transportation.

• Klíčová slova
• FFPE, RT‐QuIC, SAA, formalin‐fixed paraffin‐embedded, prion, prion disease, real‐time quaking‐induced conversion assay, seeding amplification assay,
• MeSH
• fixace tkání metody   MeSH
• formaldehyd MeSH
• lidé MeSH
• mozek * patologie   metabolismus   MeSH
• prionová bílkovina MeSH
• prionové nemoci * patologie   diagnóza   metabolismus   MeSH
• priony * metabolismus   MeSH
• zalévání tkání do parafínu metody   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• formaldehyd MeSH
• prionová bílkovina MeSH
• priony * MeSH

MicroRNAs are considered as promising prognostic and diagnostic biomarkers of human cancer since their profiles differ between tumor types. Most of the tumor profiling studies were performed on rarely available fresh frozen (FF) samples. Alternatively, archived formalin-fixed paraffin-embedded (FFPE) tissue samples are also well applicable to larger-scale retrospective miRNA profiling studies. The aim of this study was to perform systematic comparison of the miRNA expression profiles between FF and macrodissected FFPE tonsillar tumors using the TaqMan Low Density Array system, with the data processed by different software programs and two types of normalization methods. We observed a marked correlation between the miRNA expression profiles of paired FF and FFPE samples; however, only 27-38% of the differentially deregulated miRNAs overlapped between the two source systems. The comparison of the results with regard to the distinct modes of data normalization revealed an overlap in 58-67% of differentially expressed miRNAs, with no influence of the choice of software platform. Our study highlights the fact that for an accurate comparison of the miRNA expression profiles from published studies, it is important to use the same type of clinical material and to test and select the best-performing normalization method for data analysis.

• MeSH
• analýza hlavních komponent MeSH
• fixace tkání MeSH
• fixativa MeSH
• formaldehyd MeSH
• krční mandle metabolismus   MeSH
• kryoprezervace * MeSH
• lidé MeSH
• mikro RNA metabolismus   MeSH
• mikročipová analýza MeSH
• počítačové zpracování signálu MeSH
• software MeSH
• stanovení celkové genové exprese MeSH
• tonzilární nádory metabolismus   MeSH
• zalévání tkání do parafínu * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH
• Názvy látek
• fixativa MeSH
• formaldehyd MeSH
• mikro RNA MeSH

Formalin-fixed, paraffin-embedded (FFPE) tissue is the most common tissue specimen available after microscopic examination. Molecular methods, such as polymerase chain reaction (PCR) and gene expression examination, serve as a source of diagnostic and prognostic information but require high-quality RNA. However, the increasing application of RNA extracted from FFPE tissue frequently results in very small and degraded quantities of nucleic acid. This study targets gene expression analysis from FFPE specimens using real-time quantitative PCR. The whole protocol consists of several steps, that is, RNA extraction and its quality control, reverse transcription, and fluorescence detection during real-time quantitative PCR. We compared several methods in each step, chose the most effective, and with that combination we successfully examined 95% (62 from 65) FFPE samples for our genes of interest. We reached the best results with RNA isolation by using a commercial kit, carefully interpreted UV spectrophotometric values, and meticulously chose reverse transcriptase and TaqMan fluorescence detection. Our protocol improves the utility of FFPE tissue for molecular profiling studies.

• MeSH
• fixativa farmakologie   MeSH
• formaldehyd farmakologie   MeSH
• lidé MeSH
• odběr biologického vzorku metody   MeSH
• patologie metody   MeSH
• polymerázová řetězová reakce metody   MeSH
• RNA genetika   izolace a purifikace   MeSH
• stanovení celkové genové exprese metody   MeSH
• uchovávání tkání MeSH
• zalévání tkání do parafínu MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH
• práce podpořená grantem MeSH
• Názvy látek
• fixativa MeSH
• formaldehyd MeSH
• RNA MeSH

• Klíčová slova
• formalin-fixed paraffin-embedded, immunoglobulin heavy chain, mantle cell lymphoma, minimal residual disease, quantitative PCR,
• MeSH
• cyklin D1 metabolismus   MeSH
• imunohistochemie metody   MeSH
• lidé MeSH
• lymfatické uzliny metabolismus   patologie   MeSH
• lymfom z plášťových buněk metabolismus   patologie   MeSH
• reziduální nádor metabolismus   patologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• dopisy MeSH
• práce podpořená grantem MeSH
• Názvy látek
• CCND1 protein, human MeSH Prohlížeč
• cyklin D1 MeSH

To confirm a diagnosis of malignant lymphomas it is imperative to distinguish between reactive and neoplastic proliferation. The PCR (polymerase chain reaction) is a method that can be used for detection of clonal rearrangements of the immunoglobulin heavy chain (IgH) and T-cell receptor (TCR) genes. This study summarizes the outcomes of PCR analysis of IgH and TCR gene rearrangements in 91 bioptic cases of lymphoproliferative disorders. In the class of B lymphomas we detected clonal IgH rearrangement in nearly 83% of cases and in class of T lymphomas in 81% of cases. We can affirm that PCR analysis of B and T cell clonality on DNA extracted from the whole section of formalin-fixed, paraffin-embedded tissue is very suitable for routinely elaborate this. Its influence on the diagnostics of morphological unclear cases in particular, is crucial and is useful in establishing a diagnosis of lymphoid neoplasias in specimens in which histological and immunophenotypic studies are inconclusive.

• MeSH
• fixativa MeSH
• formaldehyd MeSH
• genová přestavba T-lymfocytů * MeSH
• lidé MeSH
• lymfoproliferativní nemoci diagnóza   genetika   MeSH
• polymerázová řetězová reakce MeSH
• přestavba genů pro těžké řetězce B-lymfocytů * MeSH
• zalévání tkání do parafínu MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH
• práce podpořená grantem MeSH
• Názvy látek
• fixativa MeSH
• formaldehyd MeSH

Microsatellite instability (MSI) is present in 15-20% of primary colorectal cancers. MSI status is assessed to detect Lynch syndrome, guide adjuvant chemotherapy, determine prognosis, and use as a companion test for checkpoint blockade inhibitors. Traditionally, MSI status is determined by immunohistochemistry or molecular methods. The Idylla™ MSI Assay is a fully automated molecular method (including automated result interpretation), using seven novel MSI biomarkers (ACVR2A, BTBD7, DIDO1, MRE11, RYR3, SEC31A, SULF2) and not requiring matched normal tissue. In this real-world global study, 44 clinical centers performed Idylla™ testing on a total of 1301 archived colorectal cancer formalin-fixed, paraffin-embedded (FFPE) tissue sections and compared Idylla™ results against available results from routine diagnostic testing in those sites. MSI mutations detected with the Idylla™ MSI Assay were equally distributed over the seven biomarkers, and 84.48% of the MSI-high samples had ≥ 5 mutated biomarkers, while 98.25% of the microsatellite-stable samples had zero mutated biomarkers. The concordance level between the Idylla™ MSI Assay and immunohistochemistry was 96.39% (988/1025); 17/37 discordant samples were found to be concordant when a third method was used. Compared with routine molecular methods, the concordance level was 98.01% (789/805); third-method analysis found concordance for 8/16 discordant samples. The failure rate of the Idylla™ MSI Assay (0.23%; 3/1301) was lower than that of referenced immunohistochemistry (4.37%; 47/1075) or molecular assays (0.86%; 7/812). In conclusion, lower failure rates and high concordance levels were found between the Idylla™ MSI Assay and routine tests.

• Klíčová slova
• Colorectal cancer, FFPE clinical tissue samples, Idylla™ MSI assay, Microsatellite instability, Multi-center study,
• MeSH
• fixace tkání * MeSH
• fixativa MeSH
• formaldehyd MeSH
• imunohistochemie * MeSH
• kolorektální nádory chemie   genetika   patologie   MeSH
• laboratorní automatizace MeSH
• lidé MeSH
• mikrosatelitní nestabilita * MeSH
• mutace * MeSH
• mutační analýza DNA * MeSH
• nádorové biomarkery * analýza   genetika   MeSH
• prediktivní hodnota testů MeSH
• reprodukovatelnost výsledků MeSH
• zalévání tkání do parafínu * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• multicentrická studie MeSH
• srovnávací studie MeSH
• Názvy látek
• fixativa MeSH
• formaldehyd MeSH
• nádorové biomarkery * MeSH

Ewing's sarcoma is relatively uncommon tumor representing 6-8 percent of malignant bone tumors with variable morphology. Cytogenetically, Ewing's sarcomas are characterized by a specific reciprocal chromosomal translocation t(11;22)(q24;q12). The presence of this chromosomal translocation has been detected in approximately 85 percent of the cases. The translocation results in the fusion of EWS gene from chromosome 22 to FLI1 gene at 11q24 which is a member of ETS family of transcription factors. In this study we performed a comparison of two molecular diagnostic strategies, namely RT-PCR and FISH, in fresh, frozen and formalin-fixed paraffin-embedded tissues. We conclude that FISH is a more sensitive technique than RT-PCR for the diagnosis of Ewing's tumors in formalin-fixed paraffin-embedded tissue. In conclusion, molecular pathology techniques, using reverse transcription-polymerase chain reaction (RT-PCR) and/or fluorescence in situ hybridization (FISH) are valuable diagnostic tools for evaluation of undifferentiated small round-cell tumors like Ewing's sarcoma.

• MeSH
• Ewingův sarkom diagnóza   genetika   MeSH
• fúzní onkogenní proteiny analýza   MeSH
• genetické markery * MeSH
• hybridizace in situ fluorescenční * MeSH
• lidé MeSH
• lidské chromozomy, pár 22 MeSH
• nádory měkkých tkání diagnóza   genetika   MeSH
• polymerázová řetězová reakce s reverzní transkripcí * MeSH
• protein EWS vázající RNA genetika   MeSH
• protoonkogenní protein c-fli-1 genetika   MeSH
• Retroviridae - proteiny onkogenní genetika   MeSH
• translokace genetická MeSH
• zalévání tkání do parafínu MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• FLI1 protein, human MeSH Prohlížeč
• fúzní onkogenní proteiny MeSH
• genetické markery * MeSH
• oncogene proteins v-ets MeSH Prohlížeč
• protein EWS vázající RNA MeSH
• protoonkogenní protein c-fli-1 MeSH
• Retroviridae - proteiny onkogenní MeSH

The currently valid molecular genetic subclassification of patients with diffuse large B-cell lymphoma (DLBCL) into three prognostic subgroups based on expression profiling has been the objective of numerous genetic studies. In routine clinical practice, however, expression profiling technology remains unavailable for the most of centers. Apart from the technology, in some cases molecular genetic laboratories have problems obtaining high-quality material, i.e. fresh tissues, for RNA isolation to determine gene expression. One possibility is to determine the gene expression from RNA obtained by isolation from formalin-fixed, paraffin-embedded (FFPE) tissue. This pilot study aimed at isolating RNA from FFPE in patients diagnosed with DLBCL and verifying the potential use of such RNA for the expression analysis of 7 selected genes. Although the study showed that it is possible to isolate RNA and determine the expression of the selected genes from archival material, the values of relative expression of some genes in the set were too variable to be used for unambiguous prognostic classification. It was confirmed that retrospective analyses of selected genes may be performed with sufficient material obtained, and that properly archived blocks may be used for molecular biology analyses even after 8 years.

• MeSH
• difúzní velkobuněčný B-lymfom genetika   MeSH
• formaldehyd MeSH
• geny nádorové * MeSH
• lidé MeSH
• odběr biologického vzorku MeSH
• parafín MeSH
• pilotní projekty MeSH
• prognóza MeSH
• retrospektivní studie MeSH
• RNA izolace a purifikace   MeSH
• stanovení celkové genové exprese * MeSH
• zalévání tkání do parafínu MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• formaldehyd MeSH
• parafín MeSH
• RNA MeSH

Detection of various epitopes of the p53 and MDM-2 proteins, using new antibodies was performed on formalin-fixed and paraffin-embedded tissue samples from breast cancer and compared with results obtained using well-characterized antibodies. The results show that the distribution of positive nuclei and intensity of staining varies significantly depending on the antibody used, as well as on the microwaving procedure. Antibodies DO-14, DO-13 and SMP-14 have very good characteristics and are available for immunohistochemical analysis of p53 and MDM-2. Our results indicate, that immunohistochemical expression of p53 and MDM-2 is a not stable and unitary phenomenon and from this point of view a single antibody is not sufficient for its determination. Since, characteristics of p53 and MDM-2 molecules can vary from one sample to the next, panel antibodies capable of determining a wide range of wide type and mutant conformations must be used.

• MeSH
• geny p53 imunologie   MeSH
• imunita * MeSH
• imunoadherenční reakce MeSH
• imunohistochemie MeSH
• jaderné proteiny * MeSH
• lidé MeSH
• nádorový supresorový protein p53 imunologie   MeSH
• nádory prsu imunologie   MeSH
• protilátky imunologie   MeSH
• protoonkogenní proteiny c-mdm2 MeSH
• protoonkogenní proteiny imunologie   MeSH
• techniky in vitro MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• jaderné proteiny * MeSH
• MDM2 protein, human MeSH Prohlížeč
• nádorový supresorový protein p53 MeSH
• protilátky MeSH
• protoonkogenní proteiny c-mdm2 MeSH
• protoonkogenní proteiny MeSH

BACKGROUND: Synovial sarcomas comprise up to 10 percent of malignant soft tissue tumors, and most are characterized by the chromosomal translocation t(X;18) (pl 1.2;q11.2), which results in the expression of SYT-SSX fusion transcripts. These tumors include two major histological subtypes, biphasic and monophasic. Diagnosing biphasic synovial sarcomas does not usually pose a problem, whereas the monophasic spindle-cell form can be difficult to distinguish from other spindle-cell neoplasms using histological and immunohistochemical profiles only. MATERIAL/METHODS: We investigated the presence of SYT-SSX1/2 chimeric RNA in tumors from 7 patients. We applied amplification of the specific fusion transcripts by reverse transcriptase-polymerase chain reaction (RT-PCR) in fresh, frozen tumors. We also developed a method useful for RT-PCR SYT-SSX fusion transcript detection in formalin-fixed, paraffin-embedded tissue. RESULTS: We found that both histological subtypes of synovial sarcoma were SYT-SSX positive. Moreover, we observed a correlation between histological subtype and type of SYT-SSX fusion transcript. Biphasic synovial sarcoma expressed the SYT-SSX1 fusion transcript, whereas the monophasic subtype expressed the SYT-SSX2 fusion transcript. CONCLUSIONS: The detection of SYT-SSX1/2 fusion transcripts by RT-PCR is a valuable diagnostic marker of synovial sarcoma which can be used for the reclassification of cases whose diagnosis is difficult by routine methods.

• MeSH
• dospělí MeSH
• fúzní onkogenní proteiny genetika   MeSH
• genetická transkripce MeSH
• lidé středního věku MeSH
• lidé MeSH
• lidské chromozomy X MeSH
• lidské chromozomy, pár 18 MeSH
• nádorové biomarkery genetika   MeSH
• nádorové proteiny genetika   MeSH
• polymerázová řetězová reakce s reverzní transkripcí * MeSH
• retrospektivní studie MeSH
• senioři MeSH
• synoviom diagnóza   genetika   patologie   chirurgie   MeSH
• translokace genetická MeSH
• zalévání tkání do parafínu MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Geografické názvy
• Česká republika MeSH
• Názvy látek
• fúzní onkogenní proteiny MeSH
• nádorové biomarkery MeSH
• nádorové proteiny MeSH
• SYT-SSX fusion protein MeSH Prohlížeč