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Mark Hildebrand, Aubrey Davis, Raffaela Abbriano, Haley R Pugsley, Jesse C Traller, Sarah R Smith, Roshan P Shrestha, Orna Cook, Eva L Sánchez-Alvarez, Kalpana Manandhar-Shrestha, Benjamin Alderete
The ability to image large numbers of cells at high resolution enhances flow cytometric analysis of cells and cell populations. In particular, the ability to image intracellular features adds a unique aspect to analyses, and can enable correlation between molecular phenomena resulting in alterations in cellular phenotype. Unicellular microalgae are amenable to high-throughput analysis to capture the diversity of cell types in natural samples, or diverse cellular responses in clonal populations, especially using imaging cytometry...
2016: Methods in Molecular Biology
David A Basiji
Over the last decade imaging flow cytometry (IFC) has become an established technique, as evidenced by its use in over 500 peer-reviewed scientific articles. Nevertheless, it is still an emerging technique with an installed base of less than 5 % that of conventional flow cytometers. In parallel with its adoption, the technology has evolved rapidly, increasing in speed, sensitivity, and real-time data analysis capacity by over an order of magnitude since its introduction. This chapter summarizes IFC's basic principles of operation and describes the current state of the art...
2016: Methods in Molecular Biology
Aldo Nascimento, Joanne Lannigan, David Kashatus
Mitochondria are highly dynamic organelles whose fusion and fission play an increasingly important role in a number of both normal and pathological cellular functions. Despite the increased interest in mitochondrial dynamics, robust, and quantitative methods to analyze mitochondrial fusion and fission activity in intact cells have not been developed. The current state-of-the art method to measure mitochondrial fusion activity is the polyethylene glycol (PEG) fusion assay in which cells expressing distinct mitochondrially-targeted fluorescent proteins (FPs) are fused together and mitochondrial fusion activity is determined by the rate at which color mixing occurs...
August 2016: Cytometry. Part A: the Journal of the International Society for Analytical Cytology
I Tzoran, A Rebibo-Sabbah, B Brenner, A Aharon
INTRODUCTION: Acute myeloid leukemia (AML) is characterized by rapid growth of leukemic blast cells. Extracellular vesicles (EVs) are shed from normal and pathologic cells and express membrane proteins and antigens, reflecting their cellular origin. AIM: To explore whether bone marrow EVs of AML patients originate from blast cells and are capable of influencing hematopoietic stem cells (HSC) in a pseudo-natural microenvironment obtained by co-culture of HSC with mesenchymal stem cells (MSC)...
April 2016: Thrombosis Research
Feng Qian, Ruth R Montgomery
To demonstrate effects of aging visually requires a robust technique that can reproducibly detect small differences in efficiency or kinetics between groups. Investigators of aging will greatly appreciate the benefits of Amnis ImageStream technology (, which combines quantitative flow cytometry with simultaneous high-resolution digital imaging. Imagestream is quantitative, reproducible, feasible with limited samples, and it facilitates in-depth examination of cellular mechanisms between cohorts of samples...
2015: Methods in Molecular Biology
Christopher N Parris, Sheba Adam Zahir, Hussein Al-Ali, Emma C Bourton, Christina Plowman, Piers N Plowman
Accurate and rapid methods for the detection of DNA damage foci in eukaryotic cells are central to DNA repair studies, which identify differences in DNA repair capacity in cell lines. Such assays have been important in delineating mechanisms of DNA repair in human cells. Previously we were the first to demonstrate the use of imaging flow cytometry for the detection of γ-H2AX foci in cells exposed to ionizing radiation causing the induction of DNA strand breaks. In this report we extend these studies and show an enhancement of foci quantitation and image resolution using next generation imaging flow cytometry with the Amnis Imagestream(X) Mark II...
August 2015: Cytometry. Part A: the Journal of the International Society for Analytical Cytology
Vanessa Lagal, Márcia Dinis, Dominique Cannella, Daniel Bargieri, Virginie Gonzalez, Nicole Andenmatten, Markus Meissner, Isabelle Tardieux
The apical membrane antigen 1 (AMA1) protein was believed to be essential for the perpetuation of two Apicomplexa parasite genera, Plasmodium and Toxoplasma, until we genetically engineered viable parasites lacking AMA1. The reduction in invasiveness of the Toxoplasma gondii RH-AMA1 knockout (RH-AMA1(KO)) tachyzoite population, in vitro, raised key questions about the outcome associated with these tachyzoites once inoculated in the peritoneal cavity of mice. In this study, we used AMNIS technology to simultaneously quantify and image the parasitic process driven by AMA1(KO) tachyzoites...
June 2015: Infection and Immunity
Sarah E Headland, Hefin R Jones, Adelina S V D'Sa, Mauro Perretti, Lucy V Norling
Interest in extracellular vesicle biology has exploded in the past decade, since these microstructures seem endowed with multiple roles, from blood coagulation to inter-cellular communication in pathophysiology. In order for microparticle research to evolve as a preclinical and clinical tool, accurate quantification of microparticle levels is a fundamental requirement, but their size and the complexity of sample fluids present major technical challenges. Flow cytometry is commonly used, but suffers from low sensitivity and accuracy...
June 10, 2014: Scientific Reports
Giuseppina Bologna, Paola Lanuti, Primiano D'Ambrosio, Lucia Tonucci, Laura Pierdomenico, Carlo D'Emilio, Nicola Celli, Marco Marchisio, Nicola d'Alessandro, Eugenio Santavenere, Mario Bressan, Sebastiano Miscia
Breast cancer represents the second cause of death in the European female population. The lack of specific therapies together with its high invasive potential are the major problems associated to such a tumor. In the last three decades platinum-based drugs have been considered essential constituents of many therapeutic strategies, even though with side effects and frequent generation of drug resistance. These drugs have been the guide for the research, in last years, of novel platinum and ruthenium based compounds, able to overcome these limitations...
June 2014: Biometals: An International Journal on the Role of Metal Ions in Biology, Biochemistry, and Medicine
Carrie A Franzen, Patricia E Simms, Adam F Van Huis, Kimberly E Foreman, Paul C Kuo, Gopal N Gupta
Bladder tumors represent a special therapeutic challenge as they have a high recurrence rate requiring repeated interventions and may progress to invasive or metastatic disease. Exosomes carry proteins implicated in bladder cancer progression and have been implicated in bladder cancer cell survival. Here, we characterized exosome uptake and internalization by human bladder cancer cells using Amnis ImageStreamX, an image cytometer. Exosomes were isolated by ultracentrifugation from bladder cancer culture conditioned supernatant, labeled with PKH-26, and analyzed on the ImageStreamX with an internal standard added to determine concentration...
2014: BioMed Research International
Amny N Mendenhall, Susan Frauenholtz
No abstract text is available yet for this article.
October 2013: Social Work
Iris Marangon, Nicole Boggetto, Cécilia Ménard-Moyon, Nathalie Luciani, Claire Wilhelm, Alberto Bianco, Florence Gazeau
Carbon-based nanomaterials, like carbon nanotubes (CNTs), belong to this type of nanoparticles which are very difficult to discriminate from carbon-rich cell structures and de facto there is still no quantitative method to assess their distribution at cell and tissue levels. What we propose here is an innovative method allowing the detection and quantification of CNTs in cells using a multispectral imaging flow cytometer (ImageStream, Amnis). This newly developed device integrates both a high-throughput of cells and high resolution imaging, providing thus images for each cell directly in flow and therefore statistically relevant image analysis...
2013: Journal of Visualized Experiments: JoVE
Jonathan D King, Peter Odermatt, Jürg Utzinger, Jeremiah Ngondi, Sanoussi Bamani, Yaya Kamissoko, Kadri Boubicar, Adamou Sabo Hassan, Benjamin C Nwobi, Nimzing Jip, Asrat Amnie, Tesfaye Teferi, Aryc W Mosher, Aisha E P Stewart, Elizabeth A Cromwell, Paul M Emerson
BACKGROUND: School surveys provide a convenient platform to obtain large child cohorts from multiple communities and are widely used as a proxy to determine community prevalence of neglected tropical diseases. The purpose of this study was to compare trachoma prevalence between preschool- and school-aged children and children who attend and do not attend school. METHODS: We analysed data from community-based trachoma surveys conducted from 2008-2011 in Ethiopia, Mali, Niger and Nigeria...
December 2013: International Health
Matthew C Freeman, Stephanie Ogden, Julie Jacobson, Daniel Abbott, David G Addiss, Asrat G Amnie, Colin Beckwith, Sandy Cairncross, Rafael Callejas, Jack M Colford, Paul M Emerson, Alan Fenwick, Rebecca Fishman, Kerry Gallo, Jack Grimes, Gagik Karapetyan, Brooks Keene, Patrick J Lammie, Chad Macarthur, Peter Lochery, Helen Petach, Jennifer Platt, Sarina Prabasi, Jan Willem Rosenboom, Sharon Roy, Darren Saywell, Lisa Schechtman, Anupama Tantri, Yael Velleman, Jürg Utzinger
Improvements of water, sanitation, and hygiene (WASH) infrastructure and appropriate health-seeking behavior are necessary for achieving sustained control, elimination, or eradication of many neglected tropical diseases (NTDs). Indeed, the global strategies to fight NTDs include provision of WASH, but few programs have specific WASH targets and approaches. Collaboration between disease control programs and stakeholders in WASH is a critical next step. A group of stakeholders from the NTD control, child health, and WASH sectors convened in late 2012 to discuss opportunities for, and barriers to, collaboration...
2013: PLoS Neglected Tropical Diseases
Brian K McFarlin, Randall R Williams, Adam S Venable, Karen C Dwyer, David L Haviland
Granulocytes play a key role in innate immunity and the most common functional assays are phagocytosis and oxidative burst. The purpose of this technical note is to use image-based flow cytometry to divide activated granulocytes into unique subsets based on their degree of phagocytosis and oxidative burst in response to different experimental incubations. Prior to the experiments, all reagents were titered to determine the lowest dose that resulted in an acceptable signal to noise ratio. Heparinized, whole blood (100 µl) was mixed with one of two bioparticles (E...
August 2013: Cytometry. Part A: the Journal of the International Society for Analytical Cytology
Leigh Samsel, Pradeep K Dagur, Nalini Raghavachari, Catherine Seamon, Gregory J Kato, J Philip McCoy
Endothelial cells in the peripheral circulation are rare events that require technically rigorous approaches for detection by flow cytometry. Visualization of these cells has been even more demanding, as this has historically required extensive enrichment and processing prior to attempting imaging. As a result, few, if any, examples exist on images of peripheral blood circulating endothelial cells (CECs) that include verification of the cell lineage both phenotypically and genomically. In this study, we have devised a method whereby CECs can be directly visualized after lysis of red blood cells and staining, without pre-enrichment or additional processing...
November 2013: Cytometry. Part B, Clinical Cytometry
Natividad López-Riquelme, Alfredo Minguela, Flori Villar-Permuy, Daniel Ciprian, Adela Castillejo, María-Rocio Álvarez-López, José-Luis Soto
Circulating tumor cell (CTC) enumeration is important clinically for identifying prognostic and predictive factors in patients with solid cancers. The CellSearch device (Veridex) is an immunomagnetic CTC selection and enumeration system used in clinical practice. The ImageStream (Amnis) combines the strengths of flow cytometry and fluorescent microscopy in a single platform and has potential application for CTC counting. The performance in CTC enumeration was compared between the ImageStream and CellSearch systems...
December 2013: APMIS: Acta Pathologica, Microbiologica, et Immunologica Scandinavica
Lizz Grimwade, Emma Gudgin, David Bloxham, Graham Bottley, George Vassiliou, Brian Huntly, Mike A Scott, Wendy N Erber
Mutations within the nucleophosmin NPM1 gene occur in approximately one-third of cases of acute myeloid leukemia (AML). These mutations result in cytoplasmic accumulation of the mutant NPM protein. NPM1 mutations are currently detected by molecular methods. Using samples from 37 AML patients, we investigated whether imaging flow cytometry could be a viable alternative to this current technique. Bone marrow/peripheral blood cells were stained with anti-NPM antibody and DRAQ5 nuclear stain, and data were acquired on an ImageStream imaging flow cytometer (Amnis Corp...
October 2012: Cytometry. Part A: the Journal of the International Society for Analytical Cytology
G Warnes, S Martins
The standard method of distinguishing apoptotic and oncotic cells has been by microscopic analysis of nuclei and cell membrane morphology. Thus a rapid test for analyzing large numbers of cells in the study of cell necrobiology has not been possible until the recent advent of the Amnis Image-stream and real-time Lab-on-a-Chip technologies. An interesting difference between apoptosis and oncosis is that they are ATP dependent and independent processes, respectively. Here we describe an assay measuring real-time kinetic changes in the potential differences of the inner mitochondrial membrane (mmp) and the plasma membrane (pmp) in cells immediately before and after the addition of the inducing agent...
March 2011: Cytometry. Part A: the Journal of the International Society for Analytical Cytology
Lizz Grimwade, Emma Gudgin, David Bloxham, Mike A Scott, Wendy N Erber
Acute promyelocytic leukaemia (APML) can be promptly diagnosed by detecting abnormal diffuse staining patterns of PML bodies in abnormal promyelocytes using immunofluorescence microscopy. However, this technique is subjective, with low sensitivity. Using samples from 18 patients with acute myeloid leukaemia (AML) (including four with APML), the authors investigated whether imaging flow cytometry could be a viable alternative to this current technique and improve sensitivity levels. Bone marrow/peripheral blood cells were stained with an antibody to PML, and data were acquired on an ImageStream (Amnis Corporation, Seattle, Washington, USA)...
May 2011: Journal of Clinical Pathology
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