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Dyskeratosis Congenita
Published in Dongyou Liu, Handbook of Tumor Syndromes, 2020
Patients with any of the following combinations of features are defined as having DC [31]: All three classic mucocutaneous findings of abnormal skin pigmentation, nail dystrophy, and leukoplakia.One of three mucocutaneous features plus bone marrow failure and at least two other somatic features known to occur in DC (see Table 69.2).Four or more features of HHS (such as IUGR, developmental delay, severe immune deficiency, bone marrow failure, cerebellar hypoplasia).Aplastic anemia, MDS, or pulmonary fibrosis in the setting of a known pathogenic genetic variant affecting telomere function.Two or more features of DC and laboratory evidence of short telomeres (i.e., less than first percentile for age by multicolor flow cytometry with fluorescence in situ hybridization (flow-FISH) in several subsets of lymphocytes.
Forensic Application of Telomere Shortening in Age-at-Death Estimation
Published in Sara C. Zapico, Mechanisms Linking Aging, Diseases and Biological Age Estimation, 2017
Flow FISH: The combination of Q-FISH with flow cytometry (Flow-FISH) provides more sensitivity, accuracy and speed but has a few major disadvantages, namely complexity and cost (Baerlocher and Lansdorp 2004). Flow FISH has become the method of choice for the measurement of telomere length in peripheral blood cells from human samples and has allowed the determination of the normal range of telomere lengths for specific cell subsets (Baerlocher and Lansdorp 2004, Aubert and Lansdorp 2008) that are being used both for research and for clinical investigations (Alter et al. 2007, Brummendorf et al. 2001, Yamaguchi et al. 2005).
What is the future of telomere length testing in telomere biology disorders?
Published in Expert Review of Hematology, 2023
The diagnosis of TBDs was revolutionized with the advent of leukocyte telomere length measurement by flow cytometry with in situ hybridization (flow FISH) [11]. Lymphocyte telomere length less than the first percentile for age has proven to be highly sensitive and specific in differentiating patients with TBDs from their unaffected relatives and healthy controls and has facilitated discovery of causative genes [12]. Flow FISH can provide additional information about telomere length in hematopoiesis by measuring granulocytes, total lymphocytes, naïve T-cell, memory T-cell, B-cell, and natural killer (NK) cell telomeres. However, very few laboratories have developed expertise in flow FISH telomere length testing because it is expensive, labor intensive, and requires a fresh blood sample. US CLIA-certified laboratories currently measuring lymphocyte telomeres include Repeat Diagnostics (Vancouver, BC, Canada) and Johns Hopkins Hospital (Baltimore, MD, U.S.A) [13]. There are also centers at the University of Bern (Bern, Switzerland), Aachen University (Aachen, NRW, Germany), and the Children’s Hospital at Westmead (Westmead, NSW, Australia).
Deducing the cellular mechanisms associated with the potential genotoxic impact of gold and silver engineered nanoparticles upon different lung epithelial cell lines in vitro
Published in Nanotoxicology, 2022
Samantha V. Llewellyn, Wolfgang J. Parak, Jonas Hühn, Michael J. Burgum, Stephen J. Evans, Katherine E. Chapman, Gareth J. S. Jenkins, Shareen H. Doak, Martin J. D. Clift
Flow FISH was conducted as previously described in Section 2.6, except following centrifugation the samples were re-suspended in pre-warmed (37 °C) hybridization buffer (350.0 μL of Formamide, 100.0 μL of 5% BSA, 47.0 μL of 106 mM Tris-HCl and 3.0 μL of TelC-FITC PNA probe) and incubated for 10 min, on a heat block, at 82 °C. Samples were then mixed thoroughly and incubated in a dark, humid chamber for 16-h at 37 °C and 5% CO2. Following hybridization, samples were centrifuged and re-suspended in 50% Formamide in (2X) SSC for a 10-min incubation at 40 °C twice. After the final centrifugation, supernatants were discarded, and the cells were re-suspended in pre-prepared FACS buffer (1% BSA and 0.1% Sodium Azide in (1X) PBS). Using the flow cytometer (FACSAriaTM, BD Biosciences, USA) and supporting FlowJo software, 10 000 events per biological replicate were analyzed for each ENP respectively.
Modelling the role of microbial p-cresol in colorectal genotoxicity
Published in Gut Microbes, 2019
Eiman Abdulla Al Hinai, Piyarach Kullamethee, Ian R. Rowland, Jonathan Swann, Gemma E. Walton, Daniel M. Commane
Bacterial populations from the batch culture samples were enumerated using fluorescence in situ hybridisation and flow cytometry (Flow-FISH), with oligonucleotide probes targeting specific regions of 16S rRNA as described previously.14 Briefly, commercially synthesised probes were coated with the fluorescent dye, Cy3 (Table 1). Bacteria from fermentation samples were isolated via centrifugation, washed in PBS and fixed in 4% (v/v) paraformaldehyde, they were then washed and stored at −20◦C in PBS and 99% ethanol (1:1 v/v). For analysis, bacteria were prepared in TE-FISH buffer and incubated with lysozyme (1 mg/mL of 50 000 U/mg protein) for 10 min. The cells were then washed and hybridisation completed by incubating the cells in 150 μL of hybridisation buffer (5 M NaCl, 1 M Tris/HCl pH 8, 30% formamide, ddH2O, 10% SDS and 4.55 ng ml−1 probe) for 4 hours.