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Automated antibody dispensing to improve high‐parameter flow cytometry throughput and analysis Cytom. Part A (IF 3.7) Pub Date : 2024-03-08 Victor Bosteels, Julie Van Duyse, Elien Ruyssinck, Katrien Van der Borght, Long Nguyen, Jannes Gavel, Sophie Janssens, Gert Van Isterdael
Over the past decade, the flow cytometry field has witnessed significant advancements in the number of fluorochromes that can be detected. This enables researchers to analyze more than 40 markers simultaneously on thousands of cells per second. However, with this increased complexity and multiplicity of markers, the manual dispensing of antibodies for flow cytometry experiments has become laborious
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Unveiling the epigenetic landscape of plants using flow cytometry approach Cytom. Part A (IF 3.7) Pub Date : 2024-03-04 Thakur Prava Jyoti, Shivani Chandel, Rajveer Singh
Plants are sessile creatures that have to adapt constantly changing environmental circumstances. Plants are subjected to a range of abiotic stressors as a result of unpredictable climate change. Understanding how stress‐responsive genes are regulated can help us better understand how plants can adapt to changing environmental conditions. Epigenetic markers that dynamically change in response to stimuli
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Investigation on lysosomal accumulation by a quantitative analysis of 2D phase‐maps in digital holography microscopy Cytom. Part A (IF 3.7) Pub Date : 2024-02-29 Giusy Giugliano, Michela Schiavo, Daniele Pirone, Jaromír Běhal, Vittorio Bianco, Sandro Montefusco, Pasquale Memmolo, Lisa Miccio, Pietro Ferraro, Diego L. Medina
Lysosomes are the terminal end of catabolic pathways in the cell, as well as signaling centers performing important functions such as the recycling of macromolecules, organelles, and nutrient adaptation. The importance of lysosomes in human health is supported by the fact that the deficiency of most lysosomal genes causes monogenic diseases called as a group Lysosomal Storage Diseases (LSDs). A common
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Segmentation, feature extraction and classification of leukocytes leveraging neural networks, a comparative study Cytom. Part A (IF 3.7) Pub Date : 2024-02-29 Tingxuan Fang, Xukun Huang, Xiao Chen, Deyong Chen, Junbo Wang, Jian Chen
The gold standard of leukocyte differentiation is a manual examination of blood smears, which is not only time and labor intensive but also susceptible to human error. As to automatic classification, there is still no comparative study of cell segmentation, feature extraction, and cell classification, where a variety of machine and deep learning models are compared with home‐developed approaches. In
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Modified cell trace violet proliferation assay preserves lymphocyte viability and allows spectral flow cytometry analysis Cytom. Part A (IF 3.7) Pub Date : 2024-02-29 Joanne E. Davis, Mandy Ludford‐Menting, Rachel Koldej, David S. Ritchie
In this study we describe three different methods for labeling T lymphocytes with cell trace violet (CTV), in order to track cell division in mouse and human cells, in both the in vitro and in vivo setting. We identified a modified method of CTV labeling that can be applied directly to either conventional or spectral flow cytometry, that maintained lymphocyte viability and function, yet minimized dye
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Development of a new assay for quantification of parasite load of intracellular Leishmania sp. in macrophages using flow cytometry Cytom. Part A (IF 3.7) Pub Date : 2024-02-27 Adriana C. Silva, Palloma P. Almeida, Juliana L. R. Fietto, Leandro L. Oliveira, Eduardo A. Marques‐da‐Silva
Finding novel methodologies that enhance the precision, agility, and standardization of drug discovery is crucial for studying leishmaniasis. The slide count is the technique most used to assess the leishmanicidal effect of a given drug in vitro. Despite being consolidated in the scientific environment, it presents several difficulties in its execution, assessment, and results. In addition to being
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Anti-HLA-B7/HLA-B44 strong cross immunoreactivity observed in flow cytometry HLA-B27 immunotyping Cytom. Part A (IF 3.7) Pub Date : 2024-02-20 Fabien Francois, Louis Waeckel, Anne-Emmanuelle Berger, Claude Lambert
Cross reactivities are known for human leukocyte antigen inside HLA-B7 related Cross-Reactive Group (B7CREG). Some CE-IVD flow-cytometry kits use double monoclonal antibodies (mAb) to distinguish HLA-B27 and HLA-B7 but practice reveals more complexes results. This study explores the performances of this test. Analysis of 466 consecutive cases using HLA-B27 IOTest™ kit on a Navios™ cytometer from Beckman-Coulter
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Quantitative image analysis pipeline for detecting circulating hybrid cells in immunofluorescence images with human‐level accuracy Cytom. Part A (IF 3.7) Pub Date : 2024-02-22 Robert T. Heussner, Riley M. Whalen, Ashley Anderson, Heather Theison, Joseph Baik, Summer Gibbs, Melissa H. Wong, Young Hwan Chang
Circulating hybrid cells (CHCs) are a newly discovered, tumor‐derived cell population found in the peripheral blood of cancer patients and are thought to contribute to tumor metastasis. However, identifying CHCs by immunofluorescence (IF) imaging of patient peripheral blood mononuclear cells (PBMCs) is a time‐consuming and subjective process that currently relies on manual annotation by laboratory
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Convolutional neuronal network for identifying single-cell-platelet–platelet-aggregates in human whole blood using imaging flow cytometry Cytom. Part A (IF 3.7) Pub Date : 2024-02-15 Broder Poschkamp, Sander Bekeschus
Imaging flow cytometry is an attractive method to investigate individual cells by optical properties. However, imaging flow cytometry applications with clinical relevance are scarce so far. Platelet aggregation naturally occurs during blood coagulation to form a clot. However, aberrant platelet aggregation is associated with cardiovascular disease under steady-state conditions in the blood. Several
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OMIP-101: 27-color flow cytometry panel for immunophenotyping of major leukocyte populations in fixed whole blood Cytom. Part A (IF 3.7) Pub Date : 2024-02-11 Claire Imbratta, Timothy D. Reid, Asma Toefy, Thomas J. Scriba, Elisa Nemes
This 27-color flow cytometry antibody panel allows broad immune-profiling of major leukocyte subsets in human whole blood (WB). It includes lineage markers to identify myeloid and lymphoid cell populations including granulocytes, monocytes, myeloid dendritic cells (mDCs), natural killer (NK) cells, NKT-like cells, B cells, conventional CD4 and CD8 T cells, γδ T cells, mucosa-associated invariant T
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The challenge of standardizing CAR-T cell monitoring: A comparison of two flow-cytometry methods and correlation with qPCR technique Cytom. Part A (IF 3.7) Pub Date : 2024-02-07 Juan Luis Valdivieso Shephard, Elisabet Matas Pérez, Silvia García Bujalance, Isabel Mirones Aguilar, Berta González Martínez, Antonio Pérez Martínez, Eduardo López Granados, Ana Martínez Feito, Elena Sánchez Zapardiel
Chimeric antigen receptor (CAR) T-cell therapy is a breakthrough in hematologic malignancies, such as acute B lymphoblastic leukemia (B-ALL). Monitoring this treatment is recommended, although standardized protocols have not been developed yet. This work compares two flow cytometry monitoring strategies and correlates this technique with qPCR method. CAR-T cells were detected by two different flow-cytometry
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Combinatorial antibody titrations for high-parameter flow cytometry Cytom. Part A (IF 3.7) Pub Date : 2024-02-06 Olivia K. Burn, Florian Mair, Laura Ferrer-Font
The objective of titrating fluorochrome-labeled antibodies is to identify the optimal concentration for a given marker-fluorochrome pair that results in the best possible separation between the positive and negative cell populations, while minimizing the background within the negative population. Best practices in flow cytometry dictate that each new lot of antibody should be titrated on the sample
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OMIP-100: A flow cytometry panel to investigate human neutrophil subsets Cytom. Part A (IF 3.7) Pub Date : 2024-01-05 Craig J. Schofield, Rabindra Tirouvanziam, Luke W. Garratt
This 14-color, 13-antibody optimized multicolor immunofluorescence panel (OMIP) was designed for deep profiling of neutrophil subsets in various types of human samples to contextualize neutrophil plasticity in a range of healthy and diseased states. Markers present in the OMIP allow the profiling of neutrophil subsets associated with ontogeny, migration, phagocytosis capacity, granule release, and
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Studying antigen-specific T cells through a streamlined, whole blood-based extracellular approach Cytom. Part A (IF 3.7) Pub Date : 2023-12-27 Jacques Trauet, Penelope Bourgoin, Jana Schuldt, Guillaume Lefèvre, Myriam Labalette, Jean-Marc Busnel, Julie Demaret
Techniques currently used for the study of antigen-specific T-cell responses are either poorly informative or require a heavy workload. Consequently, many perspectives associated with the broader study of such approaches remain mostly unexplored in translational research. However, these could benefit many fields including but not limited to infectious diseases, oncology, and vaccination. Herein, the
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Deep ultraviolet 266 nm laser excitation for flow cytometry Cytom. Part A (IF 3.7) Pub Date : 2023-12-20 William Telford
High dimensional flow cytometry relies on multiple laser sources to excite the wide variety of fluorochromes now available for immunophenotyping. Ultraviolet lasers (usually solid state 355 nm) are a critical part of this as they excite the BD Horizon™ Brilliant Ultraviolet (BUV) series of polymer fluorochromes. The BUV dyes have increased the number of simultaneous fluorochromes available for practical
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Leukocyte differential based on an imaging and impedance flow cytometry of microfluidics coupled with deep neural networks Cytom. Part A (IF 3.7) Pub Date : 2023-12-19 Xiao Chen, Xukun Huang, Jie Zhang, Minruihong Wang, Deyong Chen, Yueying Li, Xuzhen Qin, Junbo Wang, Jian Chen
The differential of leukocytes functions as the first indicator in clinical examinations. However, microscopic examinations suffered from key limitations of low throughputs in classifying leukocytes while commercially available hematology analyzers failed to provide quantitative accuracies in leukocyte differentials. A home-developed imaging and impedance flow cytometry of microfluidics was used to
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An approach of separating the overlapped cells or nuclei based on the outer Canny edges and morphological erosion Cytom. Part A (IF 3.7) Pub Date : 2023-12-18 Wenfei Zhang, Zhenzhou Wang
In biomedicine, the automatic processing of medical microscope images plays a key role in the subsequent analysis and diagnosis. Cell or nucleus segmentation is one of the most challenging tasks for microscope image processing. Due to the frequently occurred overlapping, few segmentation methods can achieve satisfactory segmentation accuracy yet. In this paper, we propose an approach to separate the
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Isolation of stage-specific spermatogenic cells by dynamic histone incorporation and removal in spermatogenesis Cytom. Part A (IF 3.7) Pub Date : 2023-12-12 Yasuhiro Fujiwara, Masashi Hada, Yuko Fukuda, Chizuko Koga, Erina Inoue, Yuki Okada
Due to the lack of an efficient in vitro spermatogenesis system, studies on mammalian spermatogenesis require the isolation of specific germ cell populations for further analyses. BSA gradient and elutriation have been used for several decades to purify testicular germ cells; more recently, flow cytometric cell sorting has become popular. Although each method has its advantages and disadvantages and
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Label-free cell detection of acute leukemia using ghost cytometry Cytom. Part A (IF 3.7) Pub Date : 2023-12-13 Yoko Kawamura, Kayoko Nakanishi, Yuri Murata, Kazuki Teranishi, Ryusuke Miyazaki, Keisuke Toda, Toru Imai, Yasuhiro Kajiwara, Keiji Nakagawa, Hidemasa Matsuo, Souichi Adachi, Sadao Ota, Hidefumi Hiramatsu
Early diagnosis and prompt initiation of appropriate treatment are critical for improving the prognosis of acute leukemia. Acute leukemia is diagnosed by microscopic morphological examination of bone marrow smears and flow cytometric immunophenotyping of bone marrow cells stained with fluorophore-conjugated antibodies. However, these diagnostic processes require trained professionals and are time and
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Fluorescent characterization of differentiated myotubes using flow cytometry Cytom. Part A (IF 3.7) Pub Date : 2023-12-13 Andy Nolan, Robert A. Heaton, Petra Adamova, Paige Cole, Nadia Turton, Scott H. Gillham, Daniel J. Owens, Darren W. Sexton
Flow cytometry is routinely used in the assessment of skeletal muscle progenitor cell (myoblast) populations. However, a full gating strategy, inclusive of difficult to interpret forward and side scatter data, which documents cytometric analysis of differentiated myoblasts (myotubes) has not been reported. Beyond changes in size and shape, there are substantial metabolic and protein changes in myotubes
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Correction to “A new computational approach, based on images trajectories, to identify the subjacent heterogeneity of sperm to the effects of ketanserin” Cytom. Part A (IF 3.7) Pub Date : 2023-12-11
Rodríguez-Martínez EA, Rivas CU, Ayala ME, Blanco-Rodríguez R, Juarez N, Hernandez-Vargas EA, et al. A new computational approach, based on images trajectories, to identify the subjacent heterogeneity of sperm to the effects of ketanserin. Cytometry. 2023; 103(8): 655–663. https://doi.org/10.1002/cyto.a.24732 In the originally-published article, in Figure 3, the “Time of incubation” should have been
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Combining CRISPR with Flow-FISH to study CRISPR-mediated genome perturbation Cytom. Part A (IF 3.7) Pub Date : 2023-12-06 Julian J. Freen-van Heeren
Since the advent of the clustered regularly interspaced short palindromic repeats (CRISPR)–CRISPR-associated (Cas) system as a genome editing tool, the ease of studying gene function and the impact thereof on cellular function has increased incrementally. Not surprisingly, the original describers of the CRISPR/Cas system received the 2020 Nobel Prize in Chemistry. Compared to conventional genome editing
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A workflow for the enrichment, the identification, and the isolation of non-apoptotic single circulating tumor cells for RNA sequencing analysis Cytom. Part A (IF 3.7) Pub Date : 2023-12-06 Anna Abramova, Mahdi Rivandi, Liwen Yang, Nadia Stamm, Jan-Philipp Cieslik, Ellen Honisch, Dieter Niederacher, Tanja Fehm, Hans Neubauer, André Franken
Circulating tumor cells (CTCs) are constantly shed by tumor tissue and can serve as a valuable analyte for a gene expression analysis from a liquid biopsy. However, a high proportion of CTCs can be apoptotic leading to rapid mRNA decay and challenging the analysis of their transcriptome. We established a workflow to enrich, to identify, and to isolate single CTCs including the discrimination of apoptotic
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Imaging flow cytometry of tumoroids: A new method for studying GPCR expression Cytom. Part A (IF 3.7) Pub Date : 2023-11-28 V. Gratio, S. Dayot, S. Benadda, P. Nicole, L. Saveanu, T. Voisin, A. Couvineau
Fluorescence confocal microscopy is commonly used to analyze the regulation membrane proteins expression such as G protein-coupled receptors (GPCRs). With this approach, the internal movement of GPCRs within the cell can be observed with a high degree of resolution. However, these microscopy techniques led to complex and time-consuming analysis and did not allow a large population of events to be sampled
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Human mesenchymal stem cells increase LLC metastasis and stimulate or decelerate tumor development depending on injection method and cell amount Cytom. Part A (IF 3.7) Pub Date : 2023-12-01 Yurii V. Stepanov, Iuliia Golovynska, Galyna Ostrovska, Larysa Pylyp, Taisa Dovbynchuk, Liudmyla I. Stepanova, Oleksandr Gorbach, Volodymyr Shablii, Hao Xu, Liudmyla V. Garmanchuk, Tymish Y. Ohulchanskyy, Junle Qu, Galina I. Solyanik
Mesenchymal stem cells (MSCs) being injected into the body can stimulate or decelerate carcinogenesis. Here, the direction of influence of human placenta-derived MSCs (P-MSCs) on the Lewis lung carcinoma (LLC) tumor development and metastatic potential is investigated in C57BL/6 mice depending on the injection method. After intramuscular co-inoculation of LLC and P-MSCs (LLC + P-MSCs), the growth of
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A 19-color single-tube full spectrum flow cytometry assay for the detection of measurable residual disease in acute myeloid leukemia Cytom. Part A (IF 3.7) Pub Date : 2023-11-20 Hendrik Fokken, Julian Waclawski, Nadine Kattre, Arnold Kloos, Sebastian Müller, Max Ettinger, Tim Kacprowski, Michael Heuser, Tobias Maetzig, Adrian Schwarzer
Multiparameter flow cytometry (MFC) has emerged as a standard method for quantifying measurable residual disease (MRD) in acute myeloid leukemia. However, the limited number of available channels on conventional flow cytometers requires the division of a diagnostic sample into several tubes, restricting the number of cells and the complexity of immunophenotypes that can be analyzed. Full spectrum flow
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Assessment of inter-operator variability in peripheral monocyte subset gating strategy using flow cytometry in patients with suspected acute stroke Cytom. Part A (IF 3.7) Pub Date : 2023-11-16 Evelyne Heng, Marie Neuwirth, Floriane Mas, Geneviève Contant, Mikaël Mazighi, Joffrey Feriel, Bertrand Montpellier, Caren Brumpt, Georges Jourdi, Emmanuel Curis, Virginie Siguret
Innovative tools to reliably identify patients with acute stroke are needed. Peripheral monocyte subsets, that is, classical-Mon1, intermediate-Mon2, and non-classical-Mon3, with their activation marker expression analyzed using flow-cytometry (FCM) could be interesting cell biomarker candidates.
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Guidelines for establishing a cytometry laboratory Cytom. Part A (IF 3.7) Pub Date : 2023-11-08 Anna C. Belkina, Caroline E. Roe, Vera A. Tang, Jessica B. Back, Claudia Bispo, Alexis Conway, Uttara Chakraborty, Kathleen T. Daniels, Gelo de la Cruz, Laura Ferrer-Font, Andrew Filby, David M. Gravano, Michael D. Gregory, Christopher Hall, Christian Kukat, André Mozes, Diana Ordoñez-Rueda, Eva Orlowski-Oliver, Isabella Pesce, Ziv Porat, Nicole J. Poulton, Kristen M. Reifel, Aja M. Rieger, Rachael
The purpose of this document is to provide guidance for establishing and maintaining growth and development of flow cytometry shared resource laboratories. While the best practices offered in this manuscript are not intended to be universal or exhaustive, they do outline key goals that should be prioritized to achieve operational excellence and meet the needs of the scientific community. Additionally
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CD72 is a pan-tumor antigen associated to pediatric acute leukemia Cytom. Part A (IF 3.7) Pub Date : 2023-10-24 Barbara Buldini, Giovanni Faggin, Elena Porcù, Pamela Scarparo, Katia Polato, Claudia Tregnago, Elena Varotto, Paolo Rizzardi, Carmelo Rizzari, Franco Locatelli, Alessandra Biffi, Martina Pigazzi
In the development of novel immunotherapeutic approaches, the step of target identification is a challenging process, because it aims at identifying robust tumor-associated antigens (TAAs) specific for the pathological population and causing no off-target effects. Here we propose CD72 as a novel and robust TAA for pediatric acute leukemias. We provided an outline of CD72 expression assessed by flow
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PACMan: A software package for automated single-cell chlorophyll fluorometry Cytom. Part A (IF 3.7) Pub Date : 2023-10-20 Olle Pontén, Linhong Xiao, Jeanne Kutter, Yuan Cui, Carolina Wählby, Lars Behrendt
Microalgae, small photosynthetic unicells, are of great interest to ecology, ecotoxicology and biotechnology and there is a growing need to investigate the ability of cells to photosynthesize under variable conditions. Current strategies involve hand-operated pulse-amplitude-modulated (PAM) chlorophyll fluorimeters, which can provide detailed insights into the photophysiology of entire populations-
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Advance of microfluidic flow cytometry enabling high-throughput characterization of single-cell electrical and structural properties Cytom. Part A (IF 3.7) Pub Date : 2023-10-10 Xukun Huang, Xiao Chen, Huiwen Tan, Minruihong Wang, Yimin Li, Yuanchen Wei, Jie Zhang, Deyong Chen, Junbo Wang, Yueying Li, Jian Chen
This paper reported a micro flow cytometer capable of high-throughput characterization of single-cell electrical and structural features based on constrictional microchannels and deep neural networks. When single cells traveled through microchannels with constricted cross-sectional areas, they effectively blocked concentrated electric field lines, producing large impedance variations. Meanwhile, the
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OMIP-099: 31-color spectral flow cytometry panel to investigate the steady-state phenotype of human T cells Cytom. Part A (IF 3.7) Pub Date : 2023-10-09 Zeb R. Zacharias, Jon C. D. Houtman
We have developed a 31-color panel to define the steady-state phenotype of T cells in human peripheral blood (Table 1). The panel presented here was optimized using cryopreserved peripheral blood mononuclear cells (PBMC). The markers included in this panel were chosen in order to characterize the steady-state phenotype of T cells and includes markers (CD45RA, CD45RO, CCR7, CD95) to distinguish the
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Hematopoietic stem cells and extramedullary hematopoiesis in the lungs Cytom. Part A (IF 3.7) Pub Date : 2023-10-09 Andrew Reichard, Nicholas Wanner, Samar Farha, Kewal Asosingh
Hematopoietic stem cells are key players in hematopoiesis as the body maintains a physiologic steady state, and the signaling pathways and control mechanisms of these dynamic cells are implicated in processes from inflammation to cancer. Although the bone marrow is commonly regarded as the site of hematopoiesis and hematopoietic stem cell residence, these cells also circulate in the blood and reside
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OMIP-098: A 26 parameter, 24 color flow cytometry panel for human memory NK cell phenotyping Cytom. Part A (IF 3.7) Pub Date : 2023-10-08 Matthew Creegan, Justin Degler, Dominic Paquin-Proulx, Michael A. Eller, Kawthar Machmach
1 PURPOSE AND APPROPRIATE SAMPLES This 26-parameter flow cytometry panel has been developed and optimized to analyze NK cell phenotype, using cryopreserved peripheral blood mononuclear cells (PBMCs) from people living with and without human immunodeficiency virus (PLWH, PWOH). Our panel is designed for the analysis of several parameters of total NK cells and memory NK cell subsets including markers
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Best practices for instrument settings and raw data analysis in plant flow cytometry Cytom. Part A (IF 3.7) Pub Date : 2023-10-08 Petr Koutecký, Tyler Smith, João Loureiro, Paul Kron
Flow cytometry (FCM) is now the most widely used method to determine ploidy levels and genome size of plants. To get reliable estimates and allow reproducibility of measurements, the methodology should be standardized and follow the best practices in the field. In this article, we discuss instrument calibration and quality control and various instrument and acquisition settings (parameters, flow rate
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Shared resource lab (SRL) strategies for supporting high-dimensional cytometry data analysis Cytom. Part A (IF 3.7) Pub Date : 2023-10-06 David M. Gravano, Aja M. Rieger, Lauren Nettenstrom, Christopher Hall, Laura Ferrer-Font
With the increase in the number of parameters that can be detected at the single-cell level using flow and mass cytometry, there has been a paradigm shift when handling and analyzing data sets. Cytometry Shared Resource Laboratories (SRLs) already take on the responsibility of ensuring users have resources and training in experimental design and operation of instruments to promote high-quality data
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OMIP-097: High-parameter phenotyping of human platelets by spectral flow cytometry Cytom. Part A (IF 3.7) Pub Date : 2023-10-03 Benjamin E. J. Spurgeon, Andrew L. Frelinger
Using spectral flow cytometry, we developed a 16-color panel for analysis of platelet phenotype and function in human whole blood. The panel contains markers of clinical relevance and follows an optimized protocol for the high-parameter phenotyping of (phosphatidylserine positive) procoagulant platelets. Inclusion of established markers, such as CD62P and PAC-1, allows the subsetting of classic (proinflammatory
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A flow cytometry method for safe detection of bacterial viability Cytom. Part A (IF 3.7) Pub Date : 2023-10-03 S. Servain-Viel, M.-L. Aknin, S. Domenichini, G. Perlemuter, A.-M. Cassard, G. Schlecht-Louf, V. Lievin-Le Moal
Flow cytometry is a relevant tool to meet the requirements of academic and industrial research projects aimed at estimating the features of a bacterial population (e.g., quantity, viability, activity). One of the remaining challenges is now the safe assessment of bacterial viability while minimizing the risks inherent to existing protocols. In our core facility at the Paris-Saclay University, we have
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Assessing the performance of the Cell Painting assay across different imaging systems Cytom. Part A (IF 3.7) Pub Date : 2023-10-03 Callum Tromans-Coia, Nasim Jamali, Hamdah Shafqat Abbasi, Kenneth A. Giuliano, Mai Hagimoto, Kevin Jan, Erika Kaneko, Stefan Letzsch, Alexander Schreiner, Jonathan Z. Sexton, Mahomi Suzuki, O. Joseph Trask, Mitsunari Yamaguchi, Fumiki Yanagawa, Michael Yang, Anne E. Carpenter, Beth A. Cimini
Quantitative microscopy is a powerful method for performing phenotypic screens from which image-based profiling can extract a wealth of information, termed profiles. These profiles can be used to elucidate the changes in cellular phenotypes across cell populations from different patient samples or following genetic or chemical perturbations. One such image-based profiling method is the Cell Painting
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FCM marker importance for MRD assessment in T-cell acute lymphoblastic leukemia: An AIEOP-BFM-ALL-FLOW study group report Cytom. Part A (IF 3.7) Pub Date : 2023-09-30 Florian Kowarsch, Margarita Maurer-Granofszky, Lisa Weijler, Matthias Wödlinger, Michael Reiter, Angela Schumich, Tamar Feuerstein, Simona Sala, Michaela Nováková, Giovanni Faggin, Giuseppe Gaipa, Ondrej Hrusak, Barbara Buldini, Michael N. Dworzak
T-lineage acute lymphoblastic leukemia (T-ALL) accounts for about 15% of pediatric and about 25% of adult ALL cases. Minimal/measurable residual disease (MRD) assessed by flow cytometry (FCM) is an important prognostic indicator for risk stratification. In order to assess the MRD a limited number of antibodies directed against the most discriminative antigens must be selected. We propose a pipeline
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OMIP-096: A 24-color flow cytometry panel to identify and characterize CD4+ and CD8+ tissue-resident T cells in human skin, intestinal, and type II mucosal tissue Cytom. Part A (IF 3.7) Pub Date : 2023-09-29 Thomas R. O'Neil, Andrew N. Harman, Anthony L. Cunningham, Najla Nasr, Kirstie M. Bertram
There is a great need to understand human immune cells within tissue, where disease manifests and infection occurs. Tissue-resident memory T cells (TRMs) were discovered over a decade ago, there is a great need to understand their role in human disease. We developed a 24-color flow cytometry panel to comprehensively interrogate CD4+ and CD8+ TRMs isolated from human tissues. When interrogating cells
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Flow cytometry analysis of protein expression using antibody-derived tags followed by CITE-Seq Cytom. Part A (IF 3.7) Pub Date : 2023-09-29 Xiaoshan Shi, Wei Fan, Majid Mehrpouyan, Yu Chen, Louise M. D'Cruz, Stephanie J. Widmann, Aaron J. Tyznik
Cellular Indexing of Transcriptomes and Epitopes by Sequencing (CITE-Seq) is a single-cell phenotyping method that uses antibody-derived tags (ADTs) to quantitatively detect cell surface protein expression and generate transcriptomic data at the single-cell level. Despite the increased popularity of this technique to study cellular heterogeneity and dynamics, detailed methods on how to choose ADT markers
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A novel room concept for shared resource laboratories Cytom. Part A (IF 3.7) Pub Date : 2023-09-29 Christian Kukat, Eckhard Neumann, Eugenio Fava, Hans Fried
Shared resource laboratories/core facilities (SRLs) are centralized platforms that house and provide access to complex and expensive research equipment. Due to the highly complex nature of the instrumentation they support, SRLs have special environmental requirements for their laboratory space. Here, we describe the planning and establishment of a large light microscopy SRL, with a special focus on
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OMIP-095: 40-Color spectral flow cytometry delineates all major leukocyte populations in murine lymphoid tissues Cytom. Part A (IF 3.7) Pub Date : 2023-09-28 Aris J. Kare, Lisa Nichols, Ricardo Zermeno, Marina N. Raie, Spencer K. Tumbale, Katherine W. Ferrara
High-dimensional immunoprofiling is essential for studying host response to immunotherapy, infection, and disease in murine model systems. However, the difficulty of multiparameter panel design combined with a lack of existing murine tools has prevented the comprehensive study of all major leukocyte phenotypes in a single assay. Herein, we present a 40-color flow cytometry panel for deep immunophenotyping
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Standardized high-dimensional spectral cytometry protocol and panels for whole blood immune phenotyping in clinical and translational studies Cytom. Part A (IF 3.7) Pub Date : 2023-09-26 Tom Dott, Slobodan Culina, Rene Chemali, Cedric Ait Mansour, Florian Dubois, Bernd Jagla, Jean Marc Doisne, Lars Rogge, François Huetz, Friederike Jönsson, Pierre-Henri Commere, James Di Santo, Benjamin Terrier, Lluis Quintana-Murci, Darragh Duffy, Milena Hasan
Flow cytometry is the method of choice for immunophenotyping in the context of clinical, translational, and systems immunology studies. Among the latter, the Milieu Intérieur (MI) project aims at defining the boundaries of a healthy immune response to identify determinants of immune response variation. MI used immunophenotyping of a 1000 healthy donor cohort by flow cytometry as a principal outcome
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OPTIMAL: An OPTimized Imaging Mass cytometry AnaLysis framework for benchmarking segmentation and data exploration Cytom. Part A (IF 3.7) Pub Date : 2023-09-26 Bethany Hunter, Ioana Nicorescu, Emma Foster, David McDonald, Gillian Hulme, Andrew Fuller, Amanda Thomson, Thibaut Goldsborough, Catharien M. U. Hilkens, Joaquim Majo, Luke Milross, Andrew Fisher, Peter Bankhead, John Wills, Paul Rees, Andrew Filby, George Merces
Analysis of imaging mass cytometry (IMC) data and other low-resolution multiplexed tissue imaging technologies is often confounded by poor single-cell segmentation and suboptimal approaches for data visualization and exploration. This can lead to inaccurate identification of cell phenotypes, states, or spatial relationships compared to reference data from single-cell suspension technologies. To this
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A novel process for H&E, immunofluorescence, and imaging mass cytometry on a single slide with a concise analytics pipeline Cytom. Part A (IF 3.7) Pub Date : 2023-09-19 M. Caleb Marlin, Tayte Stephens, Christian Wright, Miles Smith, Kyle Wright, Joel M. Guthridge
Imaging mass cytometry (IMC) is a powerful spatial technology that utilizes cytometry time of flight to acquire multiplexed image datasets with up to 40 markers, via metal-tagged antibodies. Recent advances in IMC have led to the inclusion of RNAScope probes and multiple new analysis pipelines have led to faster analyses and better results. However, IMC still suffers from lower resolution (1 μm2 pixels)
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Electrical micro flow cytometry with LSTM and its application in leukocyte differential Cytom. Part A (IF 3.7) Pub Date : 2023-09-15 Huiwen Tan, Xiao Chen, Xukun Huang, Deyong Chen, Xuzhen Qin, Junbo Wang, Jian Chen
This paper developed an electrical micro flow cytometry to realize leukocyte differentials leveraging a constrictional microchannel and a deep neural network. Firstly, purified granulocytes, lymphocytes or monocytes traveled through the constrictional microchannel with a cross-sectional area marginally larger than individual cells and produced large impedance variations by blocking focused electric
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Optimization and validation of in vivo flow cytometry chimeric antigen receptor T cell detection method using CD19his indirect staining Cytom. Part A (IF 3.7) Pub Date : 2023-09-14 Silvia Zaninelli, Cristian Meli, Gianmaria Borleri, Michele Quaroni, Chiara Pavoni, Giuseppe Gaipa, Andrea Biondi, Martino Introna, Josée Golay, Alessandro Rambaldi, Benedetta Rambaldi
CD19-targeted chimeric antigen receptor T (CAR-T) cell therapy has shown unprecedented results in patients with B cell relapsed/refractory acute lymphoblastic leukemia (R/R-ALL) and B cell non-Hodgkin lymphomas where no other curative options are available. In vivo monitoring of CAR-T cell kinetics is fundamental to understand the correlation between CAR-T cells expansion and persistence with treatment
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Qualification of the differential leukocyte count and immunophenotyping in cryopreserved ex vivo whole blood assay Cytom. Part A (IF 3.7) Pub Date : 2023-09-07 Claire Imbratta, Anele Gela, Nicole Bilek, Simbarashe Mabwe, Yolundi Cloete, Rasmus Mortensen, Álvaro H. Borges, Pholo Maenetje, Mandla Mlotshwa, Gavin Churchyard, Lwitiho Sudi, Issa Sabi, Peter Meewes, Carole L. Wallis, Mark Hatherill, Thomas J. Scriba, Elisa Nemes
We developed a flow cytometry-based assay, termed Differential Leukocyte Counting and Immunophenotyping in Cryopreserved Ex vivo whole blood (DLC-ICE), that allows quantification of absolute counts and frequencies of leukocyte subsets and measures expression of activation, phenotypic and functional markers. We evaluated the performance of the DLC-ICE assay by determining inter-operator variability
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Counting rare earth metals Cytom. Part A (IF 3.7) Pub Date : 2023-08-21 Attila Tárnok
For this editorial, I planned an experiment to write an augmented intelligence editorial with the help of ChatGPT. This worked quite nicely and yielded well formulated text. After several cycles, I had a 10 lines compendium of one of the articles from this issue of Cytometry Part A. But in the end, it took much longer than writing by myself. So, I went back to the old fashioned hand written editorial
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Novel phenotypical and functional sub-classification of liver macrophages highlights changes in population dynamics in experimental mouse models Cytom. Part A (IF 3.7) Pub Date : 2023-08-22 Hiroyuki Nakashima, Bradley M. Kearney, Azusa Kato, Hiromi Miyazaki, Seigo Ito, Masahiro Nakashima, Manabu Kinoshita
Liver macrophages are critical components of systemic immune system defense mechanisms. F4/80high Kupffer cells (KCs) are the predominant liver-resident macrophages and the first immune cells to contact pathogens entering the liver. F4/80low monocyte-derived macrophages (MoMφs) are essential macrophages that modulate liver immune functions. Here we report a novel method of identifying subpopulations
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HLA DQ protein changes the cell surface distribution pattern of HLA proteins as monitored by Förster resonance energy transfer and high-resolution electron microscopy Cytom. Part A (IF 3.7) Pub Date : 2023-08-21 József Kormos, Adrienn J. Veres, László Imre, László Mátyus, Szilvia Benkő, János Szöllősi, Attila Jenei
Peptide presentation by MHC class I and MHC class II molecules plays important roles in the regulation of the immune response. One factor in these displays is the density of antigen, which must exceed a critical threshold for the effective activation of T cells. Nonrandom distribution of MHC class I and class II has already been detected at the nanometer level and at higher hierarchical levels. It
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Intra-nucleus mosaic pattern (InMop) and whole-cell Haralick combined-descriptor for identifying and characterizing acute leukemia blasts on single cell peripheral blood images Cytom. Part A (IF 3.7) Pub Date : 2023-08-11 Jonathan Tarquino, Sara Arabyarmohammadi, Rafael Enrique Tejada, Anant Madabhushi, Eduardo Romero
Acute leukemia is usually diagnosed when a test of peripheral blood shows at least 20% of abnormal immature cells (blasts), a figure even lower in case of recurrent cytogenetic abnormalities. Blast identification is crucial for white blood cell (WBC) counting, which depends on both identifying the cell type and characterizing the cellular morphology, processes susceptible of inter- and intraobserver
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FRET causing misleading signal from fluorescein excited by the violet laser in flow cytometry Cytom. Part A (IF 3.7) Pub Date : 2023-08-08 Louis Waeckel, Hana Khenine, Anne-Emmanuelle Berger, Claude Lambert
Multiple immunolabeling introduces high risks of interferences between fluorescences. As an example, in analyzing T cell clonality, we recently reported a fluorescence resonance energy transfer (FRET) effect providing an unexpected signal on B770 (PE-Cy7) detector, on the Vβ-PE positive CD3 APC-Alexa750+ T cell subsets. Here, we report another FRET effect produced by the violet laser in Vβ-FITC positive
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flowSim: Near duplicate detection for flow cytometry data Cytom. Part A (IF 3.7) Pub Date : 2023-08-02 Sebastiano Montante, Yixuan Chen, Ryan R. Brinkman
The analysis of large amounts of data is important for the development of machine learning (ML) models. flowSim is the first algorithm designed to visualize, detect and remove highly redundant information in flow cytometry (FCM) training sets to decrease the computational time for training and increase the performance of ML algorithms by reducing overfitting. flowSim performs near duplicate image detection
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Betrayal of the Markers Cytom. Part A (IF 3.7) Pub Date : 2023-07-28 Tulieca Jeyaseelan, Marina Romero-Ramos, Charlotte Christie Petersen
1 INTRODUCTION Two well-established multicolor flow cytometry panels to stain peripheral blood mononuclear cells (PBMCs) had been used with success for a long period in my laboratory. However, suddenly some very unusual results were noticed. In some samples, several populations were very positive for specific markers. This was true only for some of the samples, nevertheless, the patient group (healthy
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Improved enrichment of circulating tumor cells from diagnostic leukapheresis product Cytom. Part A (IF 3.7) Pub Date : 2023-07-17 Michiel Stevens, Anouk Mentink, Afroditi Nanou, Frank A. W. Coumans, Khrystany T. Isebia, Jaco Kraan, Paul Hamberg, John W. M. Martens, Leon W. M. M. Terstappen
The median number of circulating tumor cells (CTCs) detected in 7.5 mL of peripheral blood by CellSearch (PB-CS) in patients with metastatic prostate cancer is in the order of 1–10, which means many samples have insufficient tumor cells for comprehensive characterization. A significant increase is obtained through diagnostic leukapheresis (DLA), however, only 2%–3% of the DLA product can be processed