Cytometry Part A最新文献

{"title":"Issue Information - Editorial Policy","authors":"","doi":"10.1002/cyto.a.24653","DOIUrl":"https://doi.org/10.1002/cyto.a.24653","url":null,"abstract":"","PeriodicalId":11068,"journal":{"name":"Cytometry Part A","volume":"103 12","pages":"1020"},"PeriodicalIF":3.7,"publicationDate":"2023-12-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/cyto.a.24653","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138578217","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Volume 103A, Number 12, December 2023 Cover Image","authors":"","doi":"10.1002/cyto.a.24647","DOIUrl":"https://doi.org/10.1002/cyto.a.24647","url":null,"abstract":"","PeriodicalId":11068,"journal":{"name":"Cytometry Part A","volume":"103 12","pages":""},"PeriodicalIF":3.7,"publicationDate":"2023-12-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/cyto.a.24647","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138578215","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Issue Information - Instructions for contributors","authors":"","doi":"10.1002/cyto.a.24652","DOIUrl":"https://doi.org/10.1002/cyto.a.24652","url":null,"abstract":"","PeriodicalId":11068,"journal":{"name":"Cytometry Part A","volume":"103 12","pages":"1019"},"PeriodicalIF":3.7,"publicationDate":"2023-12-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/cyto.a.24652","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138578216","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Label-free cell detection of acute leukemia using ghost cytometry","authors":"Yoko Kawamura,&nbsp;Kayoko Nakanishi,&nbsp;Yuri Murata,&nbsp;Kazuki Teranishi,&nbsp;Ryusuke Miyazaki,&nbsp;Keisuke Toda,&nbsp;Toru Imai,&nbsp;Yasuhiro Kajiwara,&nbsp;Keiji Nakagawa,&nbsp;Hidemasa Matsuo,&nbsp;Souichi Adachi,&nbsp;Sadao Ota,&nbsp;Hidefumi Hiramatsu","doi":"10.1002/cyto.a.24821","DOIUrl":"10.1002/cyto.a.24821","url":null,"abstract":"<p>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 resource-intensive. Here, we present a novel diagnostic approach using ghost cytometry, a recently developed high-content flow cytometric approach, which enables machine vision-based, stain-free, high-speed analysis of cells, leveraging their detailed morphological information. We demonstrate that ghost cytometry can detect leukemic cells from the bone marrow cells of patients diagnosed with acute lymphoblastic leukemia and acute myeloid leukemia without relying on biological staining. The approach presented here holds promise as a precise, simple, swift, and cost-effective diagnostic method for acute leukemia in clinical practice.</p>","PeriodicalId":11068,"journal":{"name":"Cytometry Part A","volume":"105 3","pages":"196-202"},"PeriodicalIF":3.7,"publicationDate":"2023-12-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/cyto.a.24821","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138581045","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Issue Information - Publication Schedule","authors":"","doi":"10.1002/cyto.a.24650","DOIUrl":"https://doi.org/10.1002/cyto.a.24650","url":null,"abstract":"","PeriodicalId":11068,"journal":{"name":"Cytometry Part A","volume":"103 12","pages":"934"},"PeriodicalIF":3.7,"publicationDate":"2023-12-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/cyto.a.24650","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138578214","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Fluorescent characterization of differentiated myotubes using flow cytometry","authors":"Andy Nolan,&nbsp;Robert A. Heaton,&nbsp;Petra Adamova,&nbsp;Paige Cole,&nbsp;Nadia Turton,&nbsp;Scott H. Gillham,&nbsp;Daniel J. Owens,&nbsp;Darren W. Sexton","doi":"10.1002/cyto.a.24822","DOIUrl":"10.1002/cyto.a.24822","url":null,"abstract":"<p>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 allowing for their potential identification within heterogenous cell suspensions. To establish the utility of flow cytometry for determination of myoblasts and myotubes, C2C12 murine cell populations were assessed for cell morphology and metabolic reprogramming. Laser scatter, both forward (FSC; size) and side (SSC; granularity), measured cell morphology, while mitochondrial mass, reactive oxygen species (ROS) generation and DNA content were quantified using the fluorescent probes, MitoTracker green, CM-H₂DCFDA and Vybrant DyeCycle, respectively. Immunophenotyping for myosin heavy chain (MyHC) was utilized to confirm myotube differentiation. Cellular viability was determined using Annexin V/propidium iodide dual labelling. Fluorescent microscopy was employed to visualize fluorescence and morphology. Myotube and myoblast populations were resolvable through non-intuitive interpretation of laser scatter-based morphology assessment and mitochondrial mass and activity assessment. Myotubes appeared to have similar sizes to the myoblasts based on laser scatter but exhibited greater mitochondrial mass (159%, <i>p</i> &lt; 0.0001), ROS production (303%, <i>p</i> &lt; 0.0001), DNA content (18%, <i>p</i> &lt; 0.001) and expression of MyHC (147%, <i>p</i> &lt; 0.001) compared to myoblasts. Myotube sub-populations contained a larger viable cluster of cells which were unable to be fractionated from myoblast populations and a smaller population cluster which likely contains apoptotic bodies. Imaging of differentiated myoblasts that had transited through the flow cytometer revealed the presence of intact, ‘rolled-up’ myotubes, which would alter laser scatter properties and potential transit through the laser beam. Our results indicate that myotubes can be analyzed successfully using flow cytometry. Increased mitochondrial mass, ROS and DNA content are key features that correlate with MyHC expression but due to myotubes ‘rolling up’ during flow cytometric analysis, laser scatter determination of size is not positively correlated; a phenomenon observed with some size determination particles and related to surface properties of said particles. We also note a greater heterogeneity of myotubes compared to myoblasts as evidenced by the 2 distinct sub-populations. We suggest that acoustic focussing may prove effective in identifying myotube sub populations compared to traditional hydrodynamic focussing.</p>","PeriodicalId":11068,"journal":{"name":"Cytometry Part A","volume":"105 5","pages":"332-344"},"PeriodicalIF":3.7,"publicationDate":"2023-12-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/cyto.a.24822","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138581390","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Isolation of stage-specific spermatogenic cells by dynamic histone incorporation and removal in spermatogenesis","authors":"Yasuhiro Fujiwara,&nbsp;Masashi Hada,&nbsp;Yuko Fukuda,&nbsp;Chizuko Koga,&nbsp;Erina Inoue,&nbsp;Yuki Okada","doi":"10.1002/cyto.a.24812","DOIUrl":"10.1002/cyto.a.24812","url":null,"abstract":"<p>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 is used depending on the purpose of the experiment, reliance on flow cytometric cell sorting is expected to be more prevalent because fewer cells can be managed. However, the currently used flow cytometric cell sorting method for testicular germ cells relies on karyotypic differences via DNA staining. Thus, it remains challenging to separate post-meiotic haploid cells (spermatids) according to their differentiation stage despite significant variations in morphology and chromatin state. In this study, we developed a method for finely separating testicular germ cells using VC mice carrying fluorescently tagged histones. This method enables the separation of spermatogonia, spermatocytes, and spermatids based on the intensity of histone fluorescence and cell size. Combined with a DNA staining dye, this method separates spermatids after elongation according to each spermiogenic stage. Although the necessity for a specific transgenic mouse line is less versatile, this method is expected to be helpful for the isolation of testicular germ cell populations because it is highly reproducible and independent of complex cell sorter settings and staining conditions.</p>","PeriodicalId":11068,"journal":{"name":"Cytometry Part A","volume":"105 4","pages":"297-307"},"PeriodicalIF":3.7,"publicationDate":"2023-12-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/cyto.a.24812","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138581601","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Correction to “A new computational approach, based on images trajectories, to identify the subjacent heterogeneity of sperm to the effects of ketanserin”","authors":"","doi":"10.1002/cyto.a.24817","DOIUrl":"10.1002/cyto.a.24817","url":null,"abstract":"<p>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</p><p>We apologize for this error.</p>","PeriodicalId":11068,"journal":{"name":"Cytometry Part A","volume":"105 2","pages":"157"},"PeriodicalIF":3.7,"publicationDate":"2023-12-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/cyto.a.24817","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138576224","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A workflow for the enrichment, the identification, and the isolation of non-apoptotic single circulating tumor cells for RNA sequencing analysis","authors":"Anna Abramova,&nbsp;Mahdi Rivandi,&nbsp;Liwen Yang,&nbsp;Nadia Stamm,&nbsp;Jan-Philipp Cieslik,&nbsp;Ellen Honisch,&nbsp;Dieter Niederacher,&nbsp;Tanja Fehm,&nbsp;Hans Neubauer,&nbsp;André Franken","doi":"10.1002/cyto.a.24816","DOIUrl":"10.1002/cyto.a.24816","url":null,"abstract":"<p>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 and non-apoptotic CTCs for further single CTC transcriptome analysis. Viable tumor cells—we first used cells from breast cancer cell lines followed by CTCs from metastatic breast cancer patients—were enriched with the CellSearch system from diagnostic leukapheresis products, identified by immunofluorescence analysis for neoplastic markers, and isolated by micromanipulation. Then, their cDNA was generated, amplified, and sequenced. In order to exclude early apoptotic tumor cells, staining with Annexin V coupled to a fluorescent dye was used. Annexin V staining intensity was associated with decreased RNA integrity as well as lower numbers of total reads, exon reads, and detected genes in cell line cells and CTCs. A comparative RNA analysis of single cells from MDA-MB-231 and MCF7 cell lines revealed the expected differential transcriptome profiles. Enrichment and staining procedures of cell line cells that were spiked into blood had only little effect on the obtained RNA sequencing data compared to processing of naïve cells. Further, the detection of transcripts of housekeeping genes such as GAPDH was associated with a significantly higher quality of expression data from CTCs. This workflow enables the enrichment, detection, and isolation of single CTCs for individual transcriptome analyses. The discrimination of apoptotic and non-apoptotic cells allows to focus on CTCs with a high RNA integrity to ensure a successful transcriptome analysis.</p>","PeriodicalId":11068,"journal":{"name":"Cytometry Part A","volume":"105 4","pages":"242-251"},"PeriodicalIF":3.7,"publicationDate":"2023-12-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/cyto.a.24816","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138486952","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Combining CRISPR with Flow-FISH to study CRISPR-mediated genome perturbation","authors":"Julian J. Freen-van Heeren","doi":"10.1002/cyto.a.24815","DOIUrl":"10.1002/cyto.a.24815","url":null,"abstract":"&lt;p&gt;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 tools such as Transcription Activator-Like Effector Nucleases (TALENs) or Zinc Finger Nucleases (ZFNs), CRISPR is a more versatile platform that can be easily adjusted to target new genes of interest.&lt;/p&gt;&lt;p&gt;The mechanism behind genome editing by the CRISPR/Cas9 system has been recently thoroughly reviewed elsewhere [&lt;span&gt;1&lt;/span&gt;]. Briefly, CRISPR-mediated genome editing is dependent on at least two components: (1) a Cas protein that possesses endonuclease activity and (2) a variable ~20 base pair nucleic-acid based targeting crisprRNA (crRNA) that defines the target of interest. Depending on the type of Cas protein employed, also a trans-activating RNA (tracrRNA) is required in order to activate nuclease activity. Together, the gRNA and tracrRNA are often referred to as the single guide RNA, or sgRNA. Additionally, nuclease activity only occurs in the context of a protospacer adjacent motive (PAM)—a specific 3–5 nucleotide sequence that is specific to the Cas-molecule employed, further enhancing on-target specificity. Indeed, the PAM and the gene-specific targeting gRNA together define the genomic locus of interest that is targeted for double-stranded cleavage. Subsequent inefficient DNA repair machinery introduces mutations, often disabling the gene of interest in the process [&lt;span&gt;2&lt;/span&gt;]. To ensure knock-out, the gene of interest can also be targeted with two specific gRNAs, resulting in deletion of a specific piece of (non-)coding genomic information [&lt;span&gt;3&lt;/span&gt;]. CRISPR/Cas-mediated genome editing can also be used to introduce a specific mutation of interest or partial gene replacement by making use of a donor repair template [&lt;span&gt;4&lt;/span&gt;], often referred to as a homology-directed repair template after the cellular process that is exploited to facilitate this.&lt;/p&gt;&lt;p&gt;Together, these characteristics have made the CRISPR/Cas system the genome editing tool of choice for many (molecular) biologists. However, in order to validate gene knock-out or the effect thereof, researchers still often rely on (genome) sequencing data, after which knock-out cells are no longer viable nor usable in experimentation, providing only information on a genomic or RNA level. When knock-out efficiency is suboptimal, the resulting data set may be confounded unless single cell RNA sequencing has been performed. However, these types of data are both often costly and require a high level of expertise to analyze. Therefore, expanding the CRISPR toolbox with other tools that allow for visualization of gene alterations, or their influence on other genes, is hig","PeriodicalId":11068,"journal":{"name":"Cytometry Part A","volume":"105 1","pages":"7-9"},"PeriodicalIF":3.7,"publicationDate":"2023-12-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/cyto.a.24815","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138486953","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}