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flow cytometry and compensation

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https://www.readbyqxmd.com/read/29735643/flow-cytometry-data-preparation-guidelines-for-improved-automated-phenotypic-analysis
#1
Daniel Jimenez-Carretero, José M Ligos, María Martínez-López, David Sancho, María C Montoya
Advances in flow cytometry (FCM) increasingly demand adoption of computational analysis tools to tackle the ever-growing data dimensionality. In this study, we tested different data input modes to evaluate how cytometry acquisition configuration and data compensation procedures affect the performance of unsupervised phenotyping tools. An analysis workflow was set up and tested for the detection of changes in reference bead subsets and in a rare subpopulation of murine lymph node CD103+ dendritic cells acquired by conventional or spectral cytometry...
May 15, 2018: Journal of Immunology: Official Journal of the American Association of Immunologists
https://www.readbyqxmd.com/read/29728510/rapid-turnover-and-high-production-rate-of-myeloid-cells-in-adult-rhesus-macaques-with-compensations-during-aging
#2
Ziyuan He, Carolina Allers, Chie Sugimoto, Nursarat Ahmed, Hideki Fujioka, Woong-Ki Kim, Elizabeth S Didier, Marcelo J Kuroda
Neutrophils, basophils, and monocytes are continuously produced in bone marrow via myelopoiesis, circulate in blood, and are eventually removed from circulation to maintain homeostasis. To quantitate the kinetics of myeloid cell movement during homeostasis, we applied 5-bromo-2'-deoxyuridine pulse labeling in healthy rhesus macaques ( Macaca mulatta ) followed by hematology and flow cytometry analyses. Results were applied to a mathematical model, and the blood circulating half-life and daily production, respectively, of each cell type from macaques aged 5-10 y old were calculated for neutrophils (1...
May 4, 2018: Journal of Immunology: Official Journal of the American Association of Immunologists
https://www.readbyqxmd.com/read/29629731/the-effect-of-mineralocorticoid-receptor-antagonists-on-recruitment-and-function-of-endothelial-progenitor-cells-in-patients-with-congestive-heart-failure
#3
Amos Levi, Dorit Leshem-Lev, Adaya Weissler-Snir, Tal Hasin, Israel Mats, Daniel Murninkas, Ran Kornowski, Eli I Lev, Tuvia Ben-Gal
BACKGROUND: Circulating endothelial progenitor cells have an important role in the process of vascular repair. Impaired recruitment and function of endothelial progenitor cells is related to the pathophysiology of congestive heart failure. Endothelial progenitor cells have been shown to express the mineralocorticoid receptor. OBJECTIVES: To investigate the effect of mineralocorticoid receptor antagonists on endothelial progenitor cells in patients with heart failure...
April 2018: Israel Medical Association Journal: IMAJ
https://www.readbyqxmd.com/read/29605184/compensation-of-signal-spillover-in-suspension-and-imaging-mass-cytometry
#4
Stéphane Chevrier, Helena L Crowell, Vito R T Zanotelli, Stefanie Engler, Mark D Robinson, Bernd Bodenmiller
The advent of mass cytometry increased the number of parameters measured at the single-cell level while decreasing the extent of crosstalk between channels relative to dye-based flow cytometry. Although reduced, spillover still exists in mass cytometry data, and minimizing its effect requires considerable expert knowledge and substantial experimental effort. Here, we describe a novel bead-based compensation workflow and R-based software that estimates and corrects for interference between channels. We performed an in-depth characterization of the spillover properties in mass cytometry, including limitations defined by the linear range of the mass cytometer and the reproducibility of the spillover over time and across machines...
March 23, 2018: Cell Systems
https://www.readbyqxmd.com/read/29600711/jq-1-inhibits-colon-cancer-proliferation-via-suppressing-wnt-%C3%AE-catenin-signaling-and-mir-21
#5
Yan Zhang, Suli Tian, Jidong Xiong, Yongxu Zhou, Hongyu Song, Chang Liu
Bromodoamin and extraterminal (BET) protein inhibitors are a novel class of cancer therapeutics. Here we aim to investigate the efficacy and mechanism of JQ-1, a potent BET inhibitor, in colon cancer therapy. JQ-1 was used to treat SW480 colon cancer mouse xenografts. Tumor size and mouse survival were recorded. Cell apoptosis was evaluated by Annex V-FIC/PI flow cytometry. ChIP-q-PCR analysis was used to assess the H3K27 trimethylation (H3K27m3) of the p16 promoter. Wnt signaling was evaluated by Nkd2 and β-catenin levels...
March 30, 2018: Chemical Research in Toxicology
https://www.readbyqxmd.com/read/29370405/oviductal-extracellular-vesicles-oviductosomes-ovs-are-conserved-in-humans-murine-ovs-play-a-pivotal-role-in-sperm-capacitation-and-fertility
#6
Pradeepthi Bathala, Zeinab Fereshteh, Kun Li, Amal A Al-Dossary, Deni S Galileo, Patricia A Martin-DeLeon
STUDY QUESTIONS: Are extracellular vesicles (EVs) in the murine oviduct (oviductosomes, OVS) conserved in humans and do they play a role in the fertility of Pmca4-/- females? SUMMARY ANSWER: OVS and their fertility-modulating proteins are conserved in humans, arise via the apocrine pathway, and mediate a compensatory upregulation of PMCA1 (plasma membrane Ca2+-ATPase 1) in Pmca4-/- female mice during proestrus/estrus, to account for their fertility. WHAT IS KNOWN ALREADY: Recently murine OVS were identified and shown during proestrus/estrus to express elevated levels of PMCA4 which they can deliver to sperm...
March 1, 2018: Molecular Human Reproduction
https://www.readbyqxmd.com/read/29289758/comparison-of-autologous-stem-cell-transplantation-versus-haploidentical-donor-stem-cell-transplantation-for-favorable-and-intermediate-risk-acute-myeloid-leukemia-patients-in-first-complete-remission
#7
Jia Chen, Lingyi Yang, Yi Fan, Yang Xu, Yue Han, Xiaowen Tang, Huiying Qiu, Chengcheng Fu, Miao Miao, Feng Chen, Depei Wu
Stem cell transplantation (SCT) is an attractive postremission treatment option for patients with intermediate-risk acute myeloid leukemia (AML) and for some favorable-risk AML patients with additional nongenetic risk factors. Autologous SCT (auto-SCT) and haploidentical donor SCT (haplo-SCT) are the widely used alternatives in cases of a lack of a HLA-matched donor. However, limited data have been published on the direct comparison between these 2 transplant types. Based on the transplant database in our center, we conducted a retrospective study involving patients with favorable- and intermediate-risk AML in first complete remission (CR1), according to the National Comprehensive Cancer Network guideline...
April 2018: Biology of Blood and Marrow Transplantation
https://www.readbyqxmd.com/read/29282803/hair-follicle-stem-cell-faith-is-dependent-on-chromatin-remodeling-capacity-following-low-dose-radiation
#8
Nadine Schuler, Sara Timm, Claudia E Rübe
The main function of the skin, to protect against the environment, is supported by the activity of different stem cell populations. The main focus of this study was elucidating the coping mechanisms of stem cells against the stimulation of constant exposure to genotoxic stresses, both endogenous and exogenous, to ensure long-term function. Investigation of various mouse strains, differing in their DNA repair capacity, enables us to clarify fractionated low-dose irradiation (LDR)-induced consequences for different stem cell populations of the murine hair follicle (HF) in their physiological stem cell niche...
April 2018: Stem Cells
https://www.readbyqxmd.com/read/29128078/applications-of-mass-cytometry-in-clinical-medicine-the-promise-and-perils-of-clinical-cytof
#9
REVIEW
Gregory K Behbehani
Mass cytometry is a novel technology similar to flow cytometry in which antibodies are tagged with heavy metal molecules rather than fluorophores and then detected with time-of-flight mass spectrometry. This enables measurement of up to 50 simultaneous parameters with no autofluorescent background and little or no spillover or required compensation. Mass cytometry has tremendous potential for the analysis of highly complex clinical samples for the diagnosis and monitoring of malignant and autoimmune disorders...
December 2017: Clinics in Laboratory Medicine
https://www.readbyqxmd.com/read/29071677/quantitative-fluorescence-measurements-with-multicolor-flow-cytometry
#10
Lili Wang, Adolfas K Gaigalas, James Wood
Multicolor flow cytometer assays are routinely used in clinical laboratories for immunophenotyping, monitoring disease and treatment, and determining prognostic factors. However, existing methods for quantitative measurements have not yet produced satisfactory results independent of flow cytometers used. This chapter details a procedure for quantifying surface and intracellular protein biomarkers by calibrating the output of a multicolor flow cytometer in units of antibodies bound per cell (ABC). The procedure includes the following critical steps: (a) quality control (QC) and performance characterization of the multicolor flow cytometer, (b) fluorescence calibration using hard dyed microspheres assigned with fluorescence intensity values in equivalent number of reference fluorophores (ERF), (c) compensation for correction of fluorescence spillover, and (d) application of a biological reference standard for translating the ERF scale to the ABC scale...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28797111/angiogenic-t-cell-expansion-correlates-with-severity-of-peripheral-vascular-damage-in-systemic-sclerosis
#11
Mirko Manetti, Sara Pratesi, Eloisa Romano, Silvia Bellando-Randone, Irene Rosa, Serena Guiducci, Bianca Saveria Fioretto, Lidia Ibba-Manneschi, Enrico Maggi, Marco Matucci-Cerinic
The mechanisms underlying endothelial cell injury and defective vascular repair in systemic sclerosis (SSc) remain unclear. Since the recently discovered angiogenic T cells (Tang) may have an important role in the repair of damaged endothelium, this study aimed to analyze the Tang population in relation to disease-related peripheral vascular features in SSc patients. Tang (CD3+CD31+CXCR4+) were quantified by flow cytometry in peripheral blood samples from 39 SSc patients and 18 healthy controls (HC). Circulating levels of the CXCR4 ligand stromal cell-derived factor (SDF)-1α and proangiogenic factors were assessed in paired serum samples by immunoassay...
2017: PloS One
https://www.readbyqxmd.com/read/28760670/standardization-of-8-color-flow-cytometry-across-different-flow-cytometer-instruments-a-feasibility-study-in-clinical-laboratories-in-switzerland
#12
Glier Hana, Heijnen Ingmar, Hauwel Mathieu, Dirks Jan, Quarroz Stéphane, Lehmann Thomas, Rovo Alicia, Arn Kornelius, Matthes Thomas, Hogan Cassandra, Keller Peter, Dudkiewicz Ewa, Stüssi Georg, Fernandez Paula
The EuroFlow Consortium developed a fully standardized flow cytometric approach from instrument settings, through antibody panel, reagents and sample preparation protocols, to data acquisition and analysis. The Swiss Cytometry Society (SCS) promoted a study to evaluate the feasibility of using such standardized measurements of 8-color data across two different flow cytometry platforms - Becton Dickinson (BD) FACSCanto II and Beckman Coulter (BC) Navios, aiming at increasing reproducibility and inter-laboratory comparability of immunophenotypic data in clinical laboratories in Switzerland...
July 28, 2017: Journal of Immunological Methods
https://www.readbyqxmd.com/read/28565739/-research-on-the-negative-immune-regulation-of-nk-cells-in-patients-with-primary-immune-thrombocytopenia
#13
Y J Zhang, W Qu, H Liu, Y H Wang, C Y Liu, L J Li, H Q Wang, R Fu, L M Xing, Z H Shao
Objective: To investigate the levels of NK cells and their relevant cytokines (IL-10, TGF-β and IFN-γ) in patients with primary immune thrombocytopenia (ITP) . Methods: All samples were obtained from 42 patients (22 newly diagnosed and 20 in remission) and 20 healthy volunteers. The levels of IL-10 and IFN-γ in blood serum were detected by enzyme-linked immunosorbent assay (ELISA) . The percentage of CD3(-) CD56(+) NK cell, CD3(-) CD56(bright) CD16(-) NK cell, CD3(-) CD56(dim) CD16(+) NK cell in peripheral blood lymphocyte were detected by flow cytometry...
May 14, 2017: Zhonghua Xue Ye Xue za Zhi, Zhonghua Xueyexue Zazhi
https://www.readbyqxmd.com/read/28518119/analysis-of-cell-suspensions-isolated-from-solid-tissues-by-spectral-flow-cytometry
#14
Sandrine Schmutz, Mariana Valente, Ana Cumano, Sophie Novault
Flow cytometry has been used for the past 40 years to define and analyze the phenotype of lymphoid and other hematopoietic cells. Initially restricted to the analysis of a few fluorochromes, currently there are dozens of different fluorescent dyes, and up to 14-18 different dyes can be combined at a time. However, several limitations still impair the analytical capabilities. Because of the multiplicity of fluorescent probes, data analysis has become increasingly complex due to the need of large, multi-parametric compensation matrices...
May 5, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/28493330/improved-panels-for-clinical-immune-phenotyping-utilization-of-the-violet-laser
#15
Mark Ryherd, Matthew Plassmeyer, Connor Alexander, Ines Eugenio, Yuliya Kleschenko, Ariel Badger, Raavi Gupta, Oral Alpan, Søren Ulrik Sønder
BACKGROUND: Clinical diagnostic laboratories are subject to numerous regulations imposed by government agencies. Laboratory developed tests for flow cytometry panels are essentially restricted to the use of analyte-specific reagents (ASR) antibodies. With the advances in clinical flow cytometry systems, there is a trend toward the utilization of blue/red/violet laser flow systems and 8 to 10-color panels. Currently, the selection of commercially available ASR antibodies for the violet laser is very limited...
May 10, 2017: Cytometry. Part B, Clinical Cytometry
https://www.readbyqxmd.com/read/28440417/increased-cd19-cd24-cd27-b-regulatory-cells-are-associated-with-insulin-resistance-in-patients-with-type-i-hashimoto-s-thyroiditis
#16
Min Yang, Changji Du, Yinping Wang, Jun Liu
Hashimoto's thyroiditis (HT) is characterized by dysregulated immune responses and is commonly associated with insulin resistance. However, the mechanism of insulin resistance in HT remains to be fully elucidated. The aim of the present study was to investigate the correlation between the percentage of B regulatory lymphocytes (Bregs) and insulin resistance in patients with HT but with normal thyroid function (type I). A total of 59 patients with type I HT and 38 healthy volunteers were enrolled in the study...
June 2017: Molecular Medicine Reports
https://www.readbyqxmd.com/read/28369683/basic-multicolor-flow-cytometry
#17
Zofia Maciorowski, Pratip K Chattopadhyay, Paresh Jain
Multicolor flow cytometry is a rapidly evolving technology that uses multiple fluorescent markers to identify and characterize cellular subpopulations of interest, allowing rapid analysis on tens of thousands of cells per second, with the possibility of isolating pure, viable populations by cell sorting for further experimentation. This unit covers the tools needed by the beginning immunologist to plan and run multicolor experiments, with information on fluorochromes and their characteristics, spectral spillover, compensation and spread, instrument and reagent variables, and the basic elements of multicolor panel design...
April 3, 2017: Current Protocols in Immunology
https://www.readbyqxmd.com/read/28160404/evaluating-flow-cytometer-performance-with-weighted-quadratic-least-squares-analysis-of-led-and-multi-level-bead-data
#18
David R Parks, Faysal El Khettabi, Eric Chase, Robert A Hoffman, Stephen P Perfetto, Josef Spidlen, James C S Wood, Wayne A Moore, Ryan R Brinkman
We developed a fully automated procedure for analyzing data from LED pulses and multilevel bead sets to evaluate backgrounds and photoelectron scales of cytometer fluorescence channels. The method improves on previous formulations by fitting a full quadratic model with appropriate weighting and by providing standard errors and peak residuals as well as the fitted parameters themselves. Here we describe the details of the methods and procedures involved and present a set of illustrations and test cases that demonstrate the consistency and reliability of the results...
March 2017: Cytometry. Part A: the Journal of the International Society for Analytical Cytology
https://www.readbyqxmd.com/read/28133433/bioconjugatable-pegylated-hydroporphyrins-for-photochemistry-and-photomedicine-narrow-band-near-infrared-emitting-bacteriochlorins
#19
Nuonuo Zhang, Jianbing Jiang, Mengran Liu, Masahiko Taniguchi, Amit Kumar Mandal, Rosemary B Evans-Storms, J Bruce Pitner, David F Bocian, Dewey Holten, Jonathan S Lindsey
Synthetic bacteriochlorins absorb in the near-infrared (NIR) region and are versatile analogues of natural bacteriochlorophylls. The utilization of these chromophores in energy sciences and photomedicine requires the ability to tailor their physicochemical properties, including the incorporation of units to impart water solubility. Herein, we report the synthesis, from two common bacteriochlorin building blocks, of five wavelength-tunable, bioconjugatable and water-soluble bacteriochlorins along with two non-bioconjugatable benchmarks...
September 1, 2016: New Journal of Chemistry, Nouveau Journal de Chimie
https://www.readbyqxmd.com/read/28058860/liposome-mediated-rna-interference-delivery-against-erk1-and-erk2-does-not-equally-promote-chemosensitivity-in-human-hepatocellular-carcinoma-cell-line-hepg2
#20
Amir Mehdizadeh, Mohammad Hossein Somi, Masoud Darabi, Safar Farajnia, Abolfazl Akbarzadeh, Soheila Montazersaheb, Mehdi Yousefi, Mortaza Bonyadi
BACKGROUND: Extracellular signal-regulated kinase (Erk)1 and Erk2 are central mediators of mitogen-activated protein kinase signaling pathway, which plays a key role in proliferation and chemoresistance of cancer cells. However, the effect of Erk1 and Erk2 in these processes may not be the same. The aim of this study was to investigate differential effect of Erk1 and Erk2 down-regulation on chemoresistance in human hepatocellular carcinoma (HCC) cells. Expression level and relative expression analysis in HepG2 cells were performed using RT-PCR and qRT-PCR, respectively...
December 2017: Artificial Cells, Nanomedicine, and Biotechnology
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