Calibración de equipos de citometría para empleo de paneles EuroFlow
|
|
- Willis Eaton
- 6 years ago
- Views:
Transcription
1 Calibración de equipos de citometría para empleo de paneles EuroFlow Departamento de Medicina, Centro de Investigación del Cáncer y Servicio de Citometría. Universidad de Salamanca, Salamanca, España. EuroFlow TM Consortium Curso avanzado de Actualización en Onco Hematología. Buenos Aires, Mayo, 30 a Junio, 1, 2011
2 Standardization in diagnostic flow cytometry Standardization according to literature generally refers to: lists of CD codes and markers per disease category rarely a specific antibody is recommended and (almost) never a fluorochrome is proposed HOWEVER: Standardization according to GLP guidelines demands for much higher levels of standardization EuroFlow standardization aims at: usage of comparable flow cytometers (3 lasers and 8 colors) full standardization of instrument settings (e.g. based on standard beads) standardized laboratory protocols and immunostaining procedures (SOP s) careful selection of optimal antibody clones per marker/cd code selection of optimal 8-color antibody combinations and fluorochromes design of combinations of multiple 8-color tubes: estimation and APS view new software for fast and easy data analysis with automated pattern recognition recognition of normal and abnormal leukocyte subsets (complete differentiation pathways) with the same immunostaining protocols mapping of new patient samples against large data base of earlier collected patient samples, analyzed with the same immunostaining protocol
3 Standard operating procedures: What for? Get optimal, reproducible & comparable results
4 Standard operating procedures: What for? Get optimal, reproducible & comparable results
5 Standard operating procedures: What for? Get optimal, reproducible & comparable results
6 Standard operating procedures: How? Get optimal, reproducible & comparable results Instruments and set-up Fluorochromes selection & compensation issues Standardization Antibody reagents Sample preparation protocols
7 Standard operating procedures: How? Get optimal, reproducible & comparable results Instruments and set-up Fluorochromes selection & compensation issues Standardization Antibody reagents Sample preparation protocols
8 Instruments Optical configuration: Channel Laser Dichroic filter FACSCantoII (BD Bioscience) FACSAria (BD Bioscience) Emission filter LSRII (BD Bioscience) Cyan ADP (Dako/Beckman Coulter) 1 Violet 450/50 450/50 450/50 450/50 2 Violet /50 530/30 525/50 530/40 3 Blue /30 530/30 530/30 530/40 4 Blue /42 585/42 575/26 585/42 5 Blue 613/20 6 Blue LP 695/40 695/40 680/30 7 Blue /60 780/60 780/60 E750LP 8 Red 660/20 660/20 660/20 665/20 9 Red /60 780/60 780/60 E750LP
9 Instrument set up: Window of analysis SSC-A Exp-SSC Low CD8-APCH7 FSC-A LINEAR CD16-PECy7 Stable, reliable system reproduce, share and monitor Normal peripheral (PB) sample processed according to EuroFlow sample preparation protocol
10 Instrument set-up: FSC & SSC SSC-A Exp-SSC Low FSC-A LINEAR Mean FSC channel: 55,000 (± 5,000) Mean SSC channel: 13,000 (± 2,000) Fixed FSC threshold at 10,000 Normal peripheral PB sample processed according to EuroFlow sample preparation protocol
11 Instrument set up & monitoring: fluorescence APC H7-A: CD8 LOGICAL Reference value PE Cy7-A: CD16 LOGICAL - No bright positive population felt out of the window of analysis - Reference values in the plateau part of the curve in all instruments
12 Instrument set up & monitoring: fluorescence Target fluorescence channels for the 8 th Rainbow bead peak (lot: X02) Target values (lot: X02 of 8 th Rainbow bead peak) Target MFI PacB PacO FITC PE PerCPCy PECy APC APCH
13 Instrument set up & monitoring: fluorescence Reference values for the 8 th peak of Rainbow beads (lot: X02) Target MFI PacB PacO FITC PE PerCP Cy PECy APC APCH Range (MFI±1 5%) Population 1 MFI FITC PE PerCPCy PECy APC APCH PacB PacO Infinicyt TM automated bead gating and analysis tool
14 Fluorochrome selection Excitation & emission profile Instrument optical configuration APCCy7 Panel aim Relative brightness Alexa Fluor 647 APC PE Pacific Orange Availability AmCyan Type of sample/cells Spillover into other FL channels
15 Fluorochrome selection Optical configuration of the flow cytometer FL Channel Laser Dichroic mirror FACSCantoII (BD Biosciences) FACSAria (BD Biosciences) Emission filter LSRII (BD Biosciences) Cyan ADP (Dako/Beckman Coulter) 1 Violet 450/50 450/50 450/50 450/50 2 Violet /50 530/30 525/50 530/40 3 Blue /30 530/30 530/30 530/40 Commonly available fluorochromes Pacific Blue HORIZON V450 AmCyan Pacific Orange HORIZON V500 FITC Alexa Fluor Blue /42 585/42 575/26 585/42 PE 5 Blue 613/20 PETxRed 6 Blue LP 695/40 695/40 680/30 PerCPCy5.5 PerCP 7 Blue /60 780/60 780/60 E750LP PECy7 8 Red 660/20 660/20 660/20 665/20 9 Red /60 780/60 780/60 E750LP ** Alexa Fluor 700 requires a 710/50 emission filter APC Alexa Fluor 647 APCCy7 APCH7 Alexa Fluor 700**
16 Fluorochrome selection Initial selection Further comparisons FL Channel Laser Commonly available fluorochromes FL Channel Laser Commonly available fluorochromes 1 Violet 2 Violet 3 Blue Pacific Blue HORIZON V450 AmCyan Pacific Orange HORIZON V500 FITC Alexa Fluor Blue PE 5 Blue PETxRed 6 Blue PerCPCy5.5 PerCP 7 Blue PECy7 8 Red 9 Red APC Alexa Fluor 647 APCCy7 APCH7 Alexa Fluor Violet 2 Violet 3 Blue Pacific Blue HORIZON V450 AmCyan Pacific Orange HORIZON V500 FITC Alexa Fluor Blue PE 5 Blue PETxRed 6 Blue PerCPCy5.5 PerCP 7 Blue PECy7 8 Red 9 Red APC Alexa Fluor 647 APCCy7 APCH7 Alexa Fluor 700
17 Fluorochrome selection: further comparisons FL Channel Fluorochrome conjugate MFI* SI* Main Overlap Availability Violet 1 CD4 Pacific Blue (BD Biosciences) CD4 HORIZON V450 (BD Biosciences) 9, , Violet 2 (AmCyan/Pacific Orange/ Horizon V500) Violet 2 (AmCyan/Pacific Orange/ Horizon V500) * Intensity of staining and staining index (SI) obtained for reagents evaluated in normal PB samples (n=5). Overlay histograms of normal PBL stained with both reagents Blue line: Pacific Blue. Green line: HORIZON V450 % FL compensation requirements in other fluorescence channels Channel PB HV ± ± Violet 1-A Logical
18 Fluorochrome selection: further comparisons FL Channel Fluorochrome conjugate MFI* SI* Main Overlaps Availability Violet 2 CD45 Pacific Orange (Invitrogen) CD45 HORIZON V500 (BD Biosciences) CD45 AmCyan (BD Biosciences) 5, , , Violet 1 (Pacific Blue/Horizon V450) Blue 1 (FITC) Violet 1 (Pacific Blue/Horizon V450) Blue 1 (FITC) Violet 1 (Pacific Blue/Horizon V450) Blue 1 (FITC) * Intensity of staining and staining index (SI) obtained for reagents evaluated in normal PB samples (n=5). Overlay histograms of normal PBL stained with both reagents Blue line: Pacific Orange Green line: HORIZON V500 Yellow line: AmCyan Violet 2-A Logical % FL of compensation requirements in other fluorescence channels Channel Pacific HORIZON Orange V500 AmCyan 1 2.2± ± ±1.5 2 NA NA NA 3 0.8± ± ± ± ± ± ± ± ± ±0.1 NR NR 7 0.3±0.4 NR NR 8 0.1±0.2 NR NR
19 Fluorochrome selection: further comparisons FL Channel Fluorochrome conjugate MFI* SI* Main Overlaps Availability CD4 APCCy7 (BD Bioscience) 13, Blue 4 (PECy7) Red 1 (APC) Red 2 CD4 APCH7 (BD Bioscience) 9, Blue 4 (PECy7) Red 1 (APC) CD4 Alexa Fluor 700º (BD Bioscience) 4, Blue 3 (PerCPCy5.5) Blue 4 (PECy7) *Intensity of staining and staining index (SI) obtained for reagents evaluated in normal PB samples (n=5). ºInvolves modification in the optical configuration of the instrument Overlay histograms of normal PBL stained with both reagents Blue line: APC Cy7 Green line: APC H7 Yellow line: Alexa Fluor 700 Red 2-A Logical % FL of compensation requirements in other fluorescence channels Channel APC Cy7 APC H7 Alexa Fluor ± ± ± ± 0.4 NR 0.1 ± ± ± ± ± ± 0.2 NR ± ± ± ± ± ± ± ± ± NA NA NA
20 Final fluorochrome selection FL Channel Laser Fluorochromes 1 Violet Pacific Blue 2 Violet Pacific Orange 3 Blue FITC 4 Blue PE 6 Blue PerCPCy5.5 7 Blue PECy7 8 Red APC 9 Red APCH7
21 Fluorescence overlaps: Violet laser line Blue laser line Red laser line Spectrum Viewer at
22 Fluorescence compensation Single fluorochrome dyes Tandem dyes Pacific Blue Pacific Orange FITC PE APC PerCPCy5.5 PECy7 APCH7 Generic (reagent independent) FL compensation Reagent conjugate & lot specific needs
23 Fluorescence compensation Generic and reagent/lot-especific compensation tubes Generic PECy7 conjugates APCH7 conjugates CD20 PacB CD2 CD3 CD45 PacO CD8 CD4 CD8 FITC CD1 0 CD8 CD8 PE CD16 CD9 CD5 PerCPCy5-5* CD19 CD1 0 CD8 APC CD45RA CD14 CD45RO CD19 CD56 CD24 *Tandem dye CD117 HLADR CD38 CD43 CD49d CD71 CD81 Anti- Igλ. Matched negative and positive reference populations. Use of beads when reference cell population is not possible
24 Fluorescence compensation Primary Fluorescence Channel Summary of % FL compensation requirements obtained from data files (n=14) generated in 7 different flow cytometry instruments Secondary Fluorescence Channel PB PO FITC PE PerCP- Cy5.5 PE-Cy7 APC APC-H7 MIN 24.3% 0.0% 0.0% 0.0% PB MED 27.7% 0.0% NR 0.0% 0.0% NR NR MAX 31.0% 0.2% 0.6% 0.1% MIN 1.9% 0.2% 0.0% 0.0% 0.0% 0.0% 0.0% PO MED 2.4% 0.4% 0.2% 0.3% 0.0% 0.0% 0.0% MAX 2.9% 0.5% 0.3% 0.5% 0.1% 0.3% 0.5% MIN 0.0% 4.8% 10.0% 3.0% 0.2% 0.0% 0.0% FITC MED 0.0% 5.6% 12.0% 3.5% 0.3% 0.0% 0.0% MAX 0.1% 6.4% 16.0% 4.0% 0.5% 0.2% 0.2% MIN 0.0% 0.0% 0.2% 30.1% 2.2% 0.0% PE MED 0.0% 0.1% 1.3% 32.9% 2.5% 0.1% NR PerCP- Cy5.5 MAX 0.1% 0.3% 1.7% 38.9% 2.8% 0.1% MIN 0.0% 0.0% 0.0% 0.0% 12.5% 1.6% 1.0% MED 0.0% 0.0% 0.0% 0.0% 16.5% 2.4% 5.5% MAX 0.9% 0.9% 0.2% 0.1% 18.8% 3.7% 8.0% MIN 0.0% 0.0% 0.0% 0.2% 0.6% 0.0% 3.2% PE-Cy7 MED 0.0% 0.0% 0.1% 0.7% 2.9% 0.0% 6.8% MAX 0.4% 0.6% 0.5% 13.1% 5.7% 0.9% 9.1% MIN 1.0% 0.1% 8.5% APC MED NR NR NR NR 1.2% 0.1% 9.6% MAX 1.4% 0.2% 11.6% MIN 0.0% 0.0% 0.0% 0.0% 0.0% 1.3% 1.3% APC-H7 MED 0.0% 0.0% 0.0% 0.0% 0.0% 1.5% 1.8% MAX 0.2% 0.1% 0.2% 0.1% 0.2% 2.0% 3.9% NR: never required
25 Reagent s evaluations CD19 PECy7 example Population CD19 PC7 MFI (Beckman/Coulter) CD19 PECy7 MFI (BD Bioscience) Monocytes B-cells Non B-cells Stain Index Intensity of staining and staining index (SI) obtained for reagents evaluated in normal PB samples (n=7). Ilustrative bivariate dot plots CD19 PC7 (Beckman/Coulter) CD19 PECy7 (BD Bioscience)
26 Sample preparation protocol
27 Sample preparation protocol Red cell lysis Amonium Chloride Versalyse (Beckman-Coulter) Quicklysis (Cytognos) FACS Lyse (BD Biosciences) Protocol sequence: Stain-lyse-non-wash Stain-lyse-wash Stain-lyse-wash-fix Time for acquisition Immediate 1 hour 3 hours 24 hours (SLW) (SLNW) (SLWF)
28 Sample preparation protocol GOALS: * Adequate scatter resolution of major cell populations * Minimal cell loss * Preservation of fluorochrome brightness * Low impact on stability of fluorochrome tandems * Easy and fast to use * Reduced data acquisition time
29 Sample preparation protocol Scatter (FSC vs SSC) characteristics of PBL
30 Impact of the sample preparation protocol Cell loss Fluorescence intensity (MFI)
31 Sample preparation protocol & lysing reagent Fluorescence intensity & stability
32 Sample preparation protocol & lysing reagent NH 4 CL FACS Lyse MFI of CD19 + PECy7 cells in PE SLNW SLW SLWF 0h 1h 3h 24h 0h 1h 3h 24h Fluorescence intensity & stability
33 Sample preparation protocol Red cell lysis Amonium Chloride Versalyse (Beckman-Coulter) Quicklysis (Cytognos) FACS Lyse (BD Biosciences) Protocol sequence: Stain-lyse-non-wash Stain-lyse-wash Stain-lyse-wash-fix Time for acquisition Immediate 1 hour 3 hours 24 hours (SLW) (SLNW) (SLWF)
34 Sample preparation protocol 10 µl 7 µl Titration of reagents: - Final volume of 100 µl 5 µl 2,5 µl 1,25 µl - Saturation conditions for variable volumes (e.g. 2x) 0,625 µl 0 µl
35 Sample preparation protocol
36 Sample preparation protocol Initial steps (B-CLPD): - Wash twice (SmIgs) - Premix & stain BB markers first
37 Thanks!! J. Flores-Montero 1,T. Kalina 2, Q. Lecrevisse 1, J.J Pérez 3,M. Cullen 4,L. Lhermitte 5, L. Sedek 6, A. Mendonça 7, S. Bötcher 8, J. te Marvelde 9, E. Mejstříková 2, O. Hrušák 2, J.J.M. van Dongen 9 and A.Orfao 1 1, Department of Medicine, Cancer Research Centre and Cytometry Service, University of Salamanca, Salamanca, ES; 2, Department of Pediatric Hematology and Oncology, Charles University, Prague, Czech Republic; 3, Department of Hematology, University Hospital of Salamanca, Salamanca, ES; 4,St. James University Hospital, Leeds, UK; 5, Department of Hematology, Hôpital Necker, Paris, FR; 6, Department of Pediatric Hematology and Oncology, Medical University of Silesia, Zabrze, PL; 7, Department of Hematology, Instituto Portugues de Oncologia, Lisbon, PT; 8, 2nd Department of Medicine, University Klinik Schleswig- Holstein, Kiel, DE; 9, Department of Immunology, Erasmus MC, Rotterdam, NL
Visualization of digital data Interpretation of the visual
Technical aspects of 8-color flow cytometry in the diagnosis and classification of hematopoietic malignancies Tomas Kalina!"#$%&'()*+,&$'+-./(0 *1 (2#34%-.(56(7&1+3+*&/( 8$#94&/(!:&3"(;&
More informationEuroFlow standardization of flow cytometer instrument settings and immunophenotyping protocols
Leukemia (2012) 26, 1986 2010 & 2012 Macmillan Publishers Limited All rights reserved 0887-6924/12 www.nature.com/leu SPECIAL REPORT EuroFlow standardization of flow cytometer instrument settings and immunophenotyping
More informationQuality Control in polychromatic Flow Cytometry - EuroFlow interlaboratory standardization
IV. International Symposium Flow Cytometry/Molecular Biology HOSPITAL ISRAELITA ALBERT EINSTEIN São Paulo 2012 Quality Control in polychromatic Flow Cytometry - EuroFlow interlaboratory standardization
More informationDesigning and Implementing a High-Level Multicolor Flow Cytometry Assay. Brent Wood MD PhD Department of Laboratory Medicine University of Washington
Designing and Implementing a High-Level Multicolor Flow Cytometry Assay Brent Wood MD PhD Department of Laboratory Medicine University of Washington Define Purpose of Assay Most important question What
More informationIncorporating New, Bright Fluorochromes into Multicolor Panel Design
Incorporating New, Bright Fluorochromes into Multicolor Panel Design Maria C. Jaimes, MD Senior Staff Scientist BD Biosciences 23-14684-00 Overview Multicolor flow: successful application prerequisites
More informationBringing the EuroFlow Concept
Bringing the EuroFlow Concept Cytognos - EuroFlow Supporting Company Company Overview Cytognos provides through worldwide distribution a broad range of reagents and software for flow cytometry applications
More informationAnatomy of a flow cytometer
Anatomy of a flow cytometer Fluidics Optics Electronics Cells in suspension flow in single-file through an illuminated volume where they scatter light and emit fluorescence that is collected, filtered,
More informationEach question may have MULTIPLE correct answers. Select all that are correct.
Knowledge Assessment Flow Cytometry Workshop, Part 1 April 20, 2015 Each question may have MULTIPLE correct answers. Select all that are correct. 1. Tandem dyes are a. highly stable fluorophores after
More informationWhat you need to know before designing a panel
Design What you need to know before designing a panel For Research Use Only. Not for use in diagnostic or therapeutic procedures. Alexa Fluor is a registered trademark of Life Technologies Corporation.
More informationApplication Note. Assay Portability on the BD FACSVerse System. Summary. Maria Jaimes, Yibing Wang, Catherine McIntyre, and Dev Mittar
September Assay Portability on the BD FACSVerse System Maria Jaimes, Yibing Wang, Catherine McIntyre, and Dev Mittar Contents Summary Introduction 3 Objective 4 Methods 6 Results Discussion Conclusions
More informationUnderstanding Flow Cytometry
Understanding Flow Cytometry The Basic Concepts Maree Bagnara Products Sales Specialist/Account Manager Flow Cytometry PN775136 1 Successful Flow Cytometry data is driven by.. Understanding the Biology
More informationa Beckman Coulter Life Sciences: White Paper
a Beckman Coulter Life Sciences: White Paper Long Term Stabilization of Tandem Dyes for Use in High Content, Multi Variant Flow Cytometry Authors: Snehita Sattiraju 1, Tewfik Miloud 2, Neha Girish 1, Murthy
More informationPrinciples of flow cytometry: overview of flow cytometry and its uses for cell analysis and sorting. Shoreline Community College BIOL 288
Principles of flow cytometry: overview of flow cytometry and its uses for cell analysis and sorting Shoreline Community College BIOL 288 Flow Cytometry What is Flow Cytometry? Measurement of cells or particles
More informationSpherotech, Inc. 1. SPHERO TM Technical Note STN-8 Rev C
SPHERO TM Technical Note STN- Rev C. 0070 CALIBRATION AND PERFORMANCE TRACKING OF FLOW CYTOMETERS USING SPHERO TM CALIBRATION PARTICLES Introduction The SPHERO TM Calibration Particles are versatile, stable,
More informationUsing Fluorescence Spillover to Advantage
Using Fluorescence Spillover to Advantage Boston June 9, 2011 Compensation: Why and When is it Necessary? Clare Rogers, Marketing and Applications Accuri Cytometers, Inc. Ann Arbor/St.Ives 1 June 11 Summary
More informationCongreso Nacional del Laboratorio Clínico 2016
Técnicas de alta sensibilidad en la detección de enfermedad mínima residual en Mieloma Múltiple J. Flores-Montero, MD, PhD Departamento de Medicina, Servicio Central de Citometría, Centro de Investigación
More informationFlow Cytometry SOP: Monocytes from Frozen Cells
Flow Cytometry SOP: Monocytes from Frozen Cells Purpose This SOP standardizes the procedure for measuring immune cells using flow cytometry in ACTG Immunology Laboratories. Materials 1. 12x75mm flow tubes
More informationExpanding Multicolor Options in Flow Cytometry with Novel Brilliant Violet TM Fluorophores. Carsten Wiethe Scientific Application Manager BioLegend
Expanding Multicolor Options in Flow Cytometry with Novel Brilliant Violet TM Fluorophores Carsten Wiethe Scientific Application Manager BioLegend Seminar Outline BrilliantViolet TM (BV)fluorophores Instrument
More informationSelected Topics in Electrical Engineering: Flow Cytometry Data Analysis
Selected Topics in Electrical Engineering: Flow Cytometry Data Analysis Bilge Karaçalı, PhD Department of Electrical and Electronics Engineering Izmir Institute of Technology Outline Experimental design
More informationFlow Cytometry Immune Activation SOP
Flow Cytometry Immune Activation SOP Purpose This SOP standardizes the procedure for measuring immune activation of T cells using flow cytometry in ACTG Immunology Laboratories. Materials 1. 12x75mm flow
More informationFlow Cytometry - The Essentials
Flow Cytometry - The Essentials Pocket Guide to Flow Cytometry: 1. Know your Cytometer 2. Understanding Fluorescence and Fluorophores 3. Gating Process 4. Controls 5. Optimization 6. Panel Building 7.
More informationPrinciples of Multicolor Panel Design BD. BD, the BD Logo and all other trademarks are property of Becton, Dickinson and Company.
1 Principles of Multicolor Panel Design 2 Common Multicolor Applications Intracellular cytokine staining Regulatory T cells (Tregs) Protein phosphorylation (BD Phosflow) Leukemia and lymphoma phenotyping
More informationa Beckman Coulter Life Sciences: White Paper
a Beckman Coulter Life Sciences: White Paper DuraClone improves standardization of compensation workflow for high content multicolor applications by flow cytometry Authors: Neha Girish 2, Sudharsan Sathyamurthy
More informationYour Research, Revolutionized
Your Research, Revolutionized Drive Your Research Forward Your research needs are evolving and with the CytoFLEX flow cytometer you ll see just how far your data can take you. CytoFLEX has the advanced
More informationBD Multicolor CompBeads
4/2015 23-9955-01 IVD BD Multicolor CompBeads 100 compensation setups Catalog No. 644204 BD, BD Logo and all other trademarks are property of Becton, Dickinson and Company. 2015 BD Becton, Dickinson and
More informationBeyond Colorful Science. A Practical Approach to Setting Up Multicolor Panels
Beyond Colorful Science A Practical Approach to Setting Up Multicolor Panels ISCT Paris April 2014 Layout: Multicolor Panel Design Identify the pitfalls Have a look at issues with spectral overlap How
More informationFlow Cytometry SOP: 14 color flow for immune activation, senescence, and exhaustion
Flow Cytometry SOP: 14 color flow for immune activation, senescence, and exhaustion Purpose This SOP standardizes the procedure for measuring immune cells using flow cytometry in ACTG Immunology Laboratories.
More informationYour Research, Revolutionized
Your Research, Revolutionized Drive Your Research Forward Your research needs are evolving and with the CytoFLEX flow cytometer you ll see just how far your data can take you. CytoFLEX has the advanced
More information11/19/2013. Janine Zankl FACS Core Facility 13. November Cellular Parameters. Cellular Parameters. Monocytes. Granulocytes.
DEPARTEMENT BIOZENTRUM Janine Zankl FACS Core Facility 13. November 2013 Cellular Parameters Granulocytes Monocytes Basophils Neutrophils Lymphocytes Eosinophils Cellular Parameters 1 What Is Flow Cytometry?
More informationA previous ICCS Module entitled Instrument optimization - Adjusting PMT voltages and compensation 1 should be read as a prerequisite to this module.
Sponsored and reviewed by ICCS Quality and Standards Committee Title: Compensation Tips for Beckman Coulter 10-Color Navios Platform Written by: Salima Janmohamed-Anastasakis Ph.D., Applications Scientist,
More informationBest practices in panel design to optimize the isolation of cells of interest
Sort Best practices in panel design to optimize the isolation of cells of interest For Research Use Only. Not for use in diagnostic or therapeutic procedures. Alexa Fluor is a registered trademark of Life
More informationSelecting Reagents for Multicolor Flow Cytometry
HotLines Platinum Edition f a l l 0 0 6 Selecting Reagents for Multicolor Flow Cytometry By Holden Maecker and Joe Trotter The availability of flow cytometers capable of detecting 6, 8, and more colors
More informationCancer inflammation research applications and products
Cancer inflammation research applications and products Flow cytometry Immunoassays Cell imaging Instrumentation Invitrogen Attune NxT Flow Cytometer Antibodies RNA flow Conjugated antibodies for flow cytometry
More informationQdot nanocrystal. wide range of biological investigations, Qdot nanocrystals are powerful complements
Feature nanocrystal conjugates for flow cytometry Take the easy route to multicolor flow cytometry. With applications across a wide range of biological investigations, nanocrystals are powerful complements
More informationBD FACSCanto II. A proven research platform for maximum reliability and the highest quality results
BD FACSCanto II A proven research platform for maximum reliability and the highest quality results A proven platform for maximum reliability and the highest quality results Built on more than 25 years
More informationa Beckman Coulter Life Sciences: White Paper
a Beckman Coulter Life Sciences: White Paper CytoFLEX Instrument Evaluation Using Biological Specimens Authors: James Tung 1, Dan Condello 3, Albert Donnenberg 4, Erika Duggan 3, Jesus Lemus 1, John Nolan
More informationHigh-throughput automation with the Attune NxT Autosampler: consistent results across all wells and across plates
APPLICATION NOTE Attune NxT Flow Cytometer with Autosampler High-throughput automation with the Attune NxT Autosampler: consistent results across all wells and across plates Introduction The emerging field
More informationNovoCyte Flow Cytometer
NovoCyte Flow Cytometer The Flow Cytometer for Everyone 2 Experience the NovoCyte Advantage Focus on advancing your research. Let the flow cytometer do the rest. NovoCyte Flow Cytometer High Performance
More informationPowERFUL. INTUITIvE. customizable. ThE NEw STAR of. BENchToP FLow cytometry
PowERFUL. INTUITIvE. customizable. ThE NEw STAR of BENchToP FLow cytometry ABoUT AcEA Established in 2002, ACEA Biosciences, Inc. develops cutting-edge cell analysis platforms for life science research.
More informationDetecting human circulating endothelial cells using the Attune Acoustic Focusing Cytometer
APPLICATION NOTE Attune Acoustic Focusing Cytometer Detecting human circulating endothelial cells using the Attune Acoustic Focusing Cytometer Circulating endothelial cells (CECs) are mature cells shed
More informationFLOW CYTOMETRY. CyAn ADP. Analyzer
FLOW CYTOMETRY CyAn ADP Analyzer Experience the Power of the CyAn ADP and its optimal performance The Power of Detection The Power of Speed The Power of Ease The CyAn ADP Analyzer is the next step in Advanced
More informationApplication Information Bulletin: DuraClone IM Tubes Compensation setup for high content DuraClone reagents
Application Information Bulletin: DuraClone IM Tubes Compensation setup for high content DuraClone reagents Compensation setup for high content DuraClone reagents INTRODUCTION High content flow cytometry
More informationApplication Information Bulletin: Set-Up of the CytoFLEX Set-Up of the CytoFLEX* for Extracellular Vesicle Measurement
Application Information Bulletin: Set-Up of the CytoFLEX Set-Up of the CytoFLEX* for Extracellular Vesicle Measurement Andreas Spittler, MD, Associate Professor for Pathophysiology, Medical University
More informationAMREP FLOW CYTOMETRY CORE FACILITY. Flow Cytometry Data Analysis Workshop Friday 24 th October
AMREP FLOW CYTOMETRY CORE FACILITY Flow Cytometry Data Analysis Workshop Friday 24 th October Polychromatic Flow Cytometry Data Analysis: Immunophenotyping Lymphocyte sub-populations and targeting their
More informationThe NIAID Flow Cytometry Advisory Committee; the Guidelines Subcommittee
January, 1999 From: To: Subject: The NIAID Flow Cytometry Advisory Committee; the Guidelines Subcommittee NIAID DAIDS Flow Cytometry Laboratories Comparison study information for labs wishing to switch
More informationSupplementary Figure 1 (Page1)
Supplementary Figure 1 (Page1) A B FSC-A 45 Fitc-A 133 APC-A 19 APC-Cy7-A 34 PE-A 45 Fitc-A FSC-A 45 Fitcs-A C 38 PE-Cy7-A 45 Fitc-A 45 Fitc-A 7AAD FSC-A Supplementary Figure 1 (Page2) D (1) (6) (2) (3)
More informationBoundary-breaking acoustic focusing cytometry
Boundary-breaking acoustic focusing cytometry Introducing the Attune NxT Acoustic Focusing Cytometer a high-performance system that s flexible enough for any lab One of the main projects in my laboratory
More informationNovel Developments in Cytometry. Martin Adelmann Marketing Manager Cellular Analysis & Life Science Beckman Coulter EMEAI
Novel Developments in Cytometry Martin Adelmann Marketing Manager Cellular Analysis & Life Science Beckman Coulter EMEAI 1 Innovation in Flow Cytometry Cellios EPICS 1V 2 lasers, 3 colors EPICS Profile
More informationAttune NxT Acoustic Focusing Cytometer The next generation in acoustic cytometry
Attune NxT Acoustic Focusing Cytometer The next generation in acoustic cytometry Maybelline Giam Field Application Scientist The world leader in serving science Attune NxT Flow Cytometer Attune NxT Acoustic
More informationReview of techniques in flow cytometry. Peter Meeus Onze-Lieve-Vrouwziekenhuis, Aalst 5 may 2011, SCK/CEN Mol
Review of techniques in flow cytometry Peter Meeus Onze-Lieve-Vrouwziekenhuis, Aalst 5 may 2011, SCK/CEN Mol Cytometry definitions The counting and measuring of cells,... Mosby's Medical Dictionary, 8th
More informationFlexible, Intuitive and Affordable
Data Sheet guava easycyte Flow Cytometry Systems Flexible, Intuitive and Affordable The guava flow cytometry systems are easy to use and deliver complete and comprehensive cell analysis right on your benchtop.
More informationDURACLONE IM ACCELERATE YOUR PACE IN IMMUNE SYSTEM RESEARCH. For Reseach Use Only - Not for use in Diagnostic procedures
DURACLONE IM ACCELERATE YOUR PACE IN IMMUNE SYSTEM RESEARCH Your clinical research trial companion For Reseach Use Only - Not for use in Diagnostic procedures ACCELERATE YOUR PACE IN IMMUNE SYSTEM RESEARCH
More informationINTRODUCTION TO FLOW CYTOMETRY
DEPARTEMENT BIOZENTRUM INTRODUCTION TO FLOW CYTOMETRY F ACS C ore F acility Janine Zankl FACS Core Facility 3. Dezember 2015, 4pm Cellular Parameters Granulocytes Monocytes Basophils Lymphocytes Neutrophils
More informationBD FACSMelody. Cell Sorter. The simple solution for consistent, quality results
BD FACSMelody Cell Sorter The simple solution for consistent, quality results The right instrument is needed for the best results Simplicity, affordability and quality are key Cell sorting is fast becoming
More informationIdentification of red and white blood cells from whole blood samples using the Agilent 2100 bioanalyzer. Application Note
Identification of red and white blood cells from whole blood samples using the Agilent 2100 bioanalyzer Application Note Sylvie Veriac Valérie Perrone Madeleine Avon Abstract Agilent Equipment: 2100 bioanalyzer
More informationBASICS OF FLOW CYTOMETRY
BASICS OF FLOW CYTOMETRY AUTHOR: Ana Isabel Vieira APPROVAL: Henrique Veiga Fernandes Ana Sílvia Gonçalves SOP.UCF.002 03-09-2015 Pag. 1/9 Overview Flow: Fluid Cyto: Cell Metry: Measurement Flow cytometry
More information7-amino actinomycin D (7ADD) was added to all samples 10 minutes prior to analysis on the flow cytometer in order to gate 7AAD viable cells.
Antibody staining for Ho uptake analyses For HSC staining, 10 7 BM cells from Ho perfused mice were stained with biotinylated lineage antibodies (CD3, CD5, B220, CD11b, Gr-1, CD41, Ter119), anti Sca-1-PECY7,
More informationSpherotech, Inc Irma Lee Circle, Lake Forest, IL 60045
Spherotech, Inc. 36 7845 Irma Lee Circle, Lake Forest, IL 60045 SPHERO TM Ultra Rainbow Fluorescent Particles Consists of a single peak for optical alignment of any flow cytometer in all channels from
More informationSCIENTIFIC INSTITUTE OF PUBLIC HEALTH QUALITY OF MEDICAL LABORATORIES CLINICAL BIOLOGY COMMISSION COMMITTEE OF EXPERTS GLOBAL REPORT
SCIENTIFIC INSTITUTE OF PUBLIC HEALTH QUALITY OF MEDICAL LABORATORIES CLINICAL BIOLOGY COMMISSION COMMITTEE OF EXPERTS GLOBAL REPORT EXTERNAL QUALITY ASSESSMENT IN CLINICAL BIOLOGY FLOW CYTOMETRY CD34+
More informationInterest in any of the products, request or order them at Bio-Connect Diagnostics.
CE IVD Minicatalog Interest in any of the products, request or order them at Bio-Connect Diagnostics. Bio-Connect Diagnostics B.V. T NL +31 (0)26 326 44 60 T BE +32 (0)2 502 12 53 Begonialaan 3a F NL +31
More informationFlowcytometry Dirk Pacholsky
Flowcytometry Dirk Pacholsky Flowcytometry: Overview 1 2 Overview Flow Cyto Metry Fluid Cell Measurement measuring cell properties of cells in suspension Most Flow Cytometer have Spatially separated Lasers
More informationReview of techniques in imaging and cytometry. Peter Meeus Onze Lieve Vrouw Ziekenhuis, Aalst 7 may 2009, SCK/CEN Mol
Review of techniques in imaging and cytometry Peter Meeus Onze Lieve Vrouw Ziekenhuis, Aalst 7 may 2009, SCK/CEN Mol Cytometry: The counting and measuring of cells,... Mosby's Medical Dictionary, 8th edition.
More informationIntroduction to. BD FACSAria TM Cell Sorter. Flow = Fluid Cyto = Cell Metry = Measurement
What is Flow Cytometry? Introduction to BD FACSAria TM Cell Sorter Flow = Fluid Cyto = Cell Metry = Measurement BD Biosciences Application Specialist 產品應用專員 Daisy Kuo 郭正佼 A variety of measurements are
More informationFEDERAL PUBLIC SERVICE, HEALTH, FOOD CHAIN SECURITY AND ENVIRONMENT CLINICAL BIOLOGY COMMISSION COMMITTEE OF EXPERTS GLOBAL REPORT
FEDERAL PUBLIC SERVICE, HEALTH, FOOD CHAIN SECURITY AND ENVIRONMENT CLINICAL BIOLOGY COMMISSION COMMITTEE OF EXPERTS GLOBAL REPORT EXTERNAL QUALITY ASSESSMENT IN CLINICAL BIOLOGY FLOW CYTOMETRY CD34+ STEM
More informationGUAVA EASYCYTE SYSTEMS
GUAVA EASYCYTE SYSTEMS Expanding the potential of flow cytometry The life science business of Merck KGaA, Darmstadt, Germany operates as MilliporeSigma in the U.S. and Canada. Guava easycyte Systems Expanding
More informationACTG Laboratory Technologist Committee Revised Version 2.0 ACTG Lab Man CD38 Quantitation 07 May 2004
1. Principle Flow Cytometric quantitation of CD38 expression on CD8+ T lymphocytes The purpose of this assay is to quantitate the surface expression of the CD38 molecule on CD8 positive T lymphocytes.
More informationFEDERAL PUBLIC SERVICE, HEALTH, FOOD CHAIN SECURITY AND ENVIRONMENT CLINICAL BIOLOGY COMMISSION COMMITTEE OF EXPERTS GLOBAL REPORT
FEDERAL PUBLIC SERVICE, HEALTH, FOOD CHAIN SECURITY AND ENVIRONMENT CLINICAL BIOLOGY COMMISSION COMMITTEE OF EXPERTS GLOBAL REPORT EXTERNAL QUALITY ASSESSMENT IN CLINICAL BIOLOGY FLOW CYTOMETRY CD34+ STEM
More informationNavios EX FLOW CYTOMETER POWERFUL, DEPENDABLE CLINICAL FLOW CYTOMETRY
Navios EX FLOW CYTOMETER POWERFUL, DEPENDABLE CLINICAL FLOW CYTOMETRY BECAUSE EVERY EVENT MATTERS The Navios EX flow cytometer offers a solution for advanced cytometry applications optimized for the clinical
More informationBD LSRFortessa. Performance without peer, choice without compromise
BD LSRFortessa Performance without peer, choice without compromise Performance without peer, choice without compromise The BD LSRFortessa cell analyzer offers the ultimate in choice for flow cytometry,
More informationNavios EX FLOW CYTOMETER POWERFUL, DEPENDABLE FLOW CYTOMETRY
Navios EX FLOW CYTOMETER POWERFUL, DEPENDABLE FLOW CYTOMETRY BECAUSE EVERY EVENT MATTERS The Navios EX flow cytometer offers a solution for advanced cytometry applications with optimized workflows for
More informationLower spectral overlap. Optimized stain index. Innovative antibodies. Superior multicolor flow cytometry. Violet Laser Reagents: The efluor Solution
Lower spectral overlap Optimized stain index Innovative antibodies Superior multicolor flow cytometry Violet Laser Reagents: The efluor Solution www.ebioscience.com ebioscience provides innovative high
More informationWelcome to More Choice. Mouse Panels
Welcome to More Choice Mouse Panels Choose from our extensive portfolio of high-quality fluorescent-conjugated reagents to build your multicolor flow cytometry panels. Welcome to a More Colorful World
More informationAVANÇOS NO DIAGNÓSTICO E CLASSIFICAÇÃO IMUNOFENOTIPICA DE LEUCEMIAS LINFOCITICAS CRÓNICAS B
AVANÇOS NO DIAGNÓSTICO E CLASSIFICAÇÃO IMUNOFENOTIPICA DE LEUCEMIAS LINFOCITICAS CRÓNICAS B CANCER RESEARCH CENTER IBSAL-UNIVERSITY OF SALAMANCA/CSIC HEMO 2016 Congreso Brasileiro de Hematologia, Hemoterapia
More informationGuava easycyte Systems Expanding the potential of flow cytometry
Guava easycyte Systems Expanding the potential of flow cytometry Guava easycyte Systems Flexible Intuitive Affordable Merck Millipore is a business of Unleash what s possible Fifteen years ago, Guava Technologies
More informationRef: CYT-MM-MRD. For Research Use Only. Not for use in diagnostic procedures
Ref: CYT-MM-MRD For Research Use Only. Not for use in diagnostic procedures MM MRD VIALS AND COMPENSATION TUBES ARE LYOPHILIZED PRODUCTS. READ CAREFULLY THE FOLLOWING INSTRUCTIONS FOR RECONSTITUTION: The
More informationGuava EasyCyte Systems Expanding the potential of flow cytometry
Guava EasyCyte Systems Expanding the potential of flow cytometry Guava easycyte Systems Flexible Intuitive Affordable Unleash what s possible Fifteen years ago, Guava Technologies introduced the first
More informationdetermine optimum instrument settings for their own instruments and establish their own daily values.
PC7 (770/488) SETUP KIT 6607121 PN 4299504-C FLOW CYTOMETER ALIGNMENT VERIFICATION FLUOROSPHERES FLOW CYTOMETER DETECTOR STANDARDIZATION FLUOROSPHERES INTENDED USE For Research Use Only. Not for use in
More informationab pdc Subset Phenotyping Kit
ab177772 pdc Subset Phenotyping Kit Instructions for Use A set of antibodies designed to identify pdcs and subsets of pdcs by multi-color flow cytometry. This product is for research use only and is not
More informationDesigning and executing a successful flow cytometry experiment
Designing and executing a successful flow cytometry experiment 1x1 0 5 Orange Fluorescent Protein 10000 1000 100 10 0 1000 2000 3000 4000 Forward Scatter This presentation is based on true events, however,
More informationSOPVII-7. Panel X: NK-characterization
Created by judith.eckl Page 1 of 8 09/06/2011 SOPVII-7 Panel X: NK-characterization Date: Author: Petra Prinz, Judith Eckl Experimenter: Date: 08/06/2011 Experiment description: Version: 1.0 Start: End:
More informationFlow Cytometry Facility The Panel Design Tool
The Automated design of multicolor flow cytometry panels. 01.06.2017 Contents The antibody fluorophore panel Parameters & desired properties The What it can do The input needed Input processing Panel creation
More informationAntibody Titrations. Institut für HIV Forschung SOP #06-03 (Nov BS) Background. Reagents
Institut für HIV Forschung SOP #06-03 (Nov 2017 - BS) Antibody Titrations Reagents Reagent Vendor Catalogue # Stock Conc. FACS Tubes BD Falcon 352054 - Anti-CD28/CD49d Antibody BD Fastimmune 347690 - GolgiPlug
More informationRuud Hulspas, Mike Keeney, Ben Hedley and Andrea Illingworth
Sponsored and reviewed by ICCS Quality and Standards Committee Title: Quality of Reagents Monoclonal Antibodies Written by: Date: March 6, 2018 Ruud Hulspas, Mike Keeney, Ben Hedley and Andrea Illingworth
More informationCompensation: Fundamental Principles
Flow Cytometry Seminar Series 2017 : Fundamental Principles Spillover correction in multicolor flow cytometry 28.02.2017 http://www.cytometry.uzh.ch Contents Fluorescence and its detection Absorption and
More informationIntroduction to Flow Cytometry. -- BD FACSCanto II TM. Daisy Kuo Application Specialist BDBiosciences
Introduction to Flow Cytometry -- BD FACSCanto II TM Daisy Kuo Application Specialist E-mail: daisy_kuo@bd.com BDBiosciences Outline Basic Concept of Flow Cytometry FACSCanto II System Introduction Application
More informationCLEARLLAB LS LYMPHOID SCREEN REAGENT
CLEARLLAB LS LYMPHOID SCREEN REAGENT CE MARKED ANTIBODY COMBINATION FOR LEUKEMIA / LYMPHOMA ANALYSIS Because Your Patient is Her Everything BECAUSE YOUR PATIENT IS HER EVERYTHING ClearLLab LS Lymphoid
More informationFlow Cytometry For New PhDs
Flow Cytometry For New PhDs 2012 Simon Monard SCRM and CIR smonard@staffmail.ed.ac.uk Derek Davies CRUK derek.davies@cancer.org.uk Monday 20th and Tues 21st Feb 2012 Program Day 1: Day 2: Basics of flow
More informationFlow Cytometry. Flow Cytometry Basics Guide
Flow Cytometry Flow Cytometry Basics Guide Table of Contents Chapter 1 Chapter 2 Chapter 3 Chapter 4 Chapter 5 Principles of the Flow Cytometer Fluidics System.... 3 Optics and Detection.... 4 Signal and
More informationFACS Core Facility User Group Meeting. 25. April 2018 Janine Bögli
FACS Core Facility User Group Meeting 25. April 2018 Janine Bögli Overview Basis: Forensic Flow Cytometry by Jennifer Wilshire, PhD, Stemcell Technologies You will suggest the crime in each case 8 Practical
More informationStandardized flow cytometry for highly sensitive MRD measurements in B-cell acute lymphoblastic leukemia
Supplemental Methods & Figures Standardized flow cytometry for highly sensitive MRD measurements in B-cell acute lymphoblastic leukemia Prisca Theunissen 1#, Ester Mejstrikova 2#, Lukasz Sedek 3, Alita
More informationSupplementary Figure. S1
Supplementary Figure. S1 Supplementary Figure S1. Correlation of phagocytic ability measured with YG and YO beads. Fresh human monocytes (2 10 6 /ml) were labelled with APC conjugated anti CD14 mab alone
More informationMinimum Information about a Flow Cytometry Experiment (MIFlowCyt) Annotation
Minimum Information about a Flow Cytometry Experiment (MIFlowCyt) Annotation 1. Experiment Overview 1.1 Purpose The purpose of these sets of experiments is to develop a methodology of identifying and quantifying
More informationApplication Note. Yeast Analysis with a life - dead staining kit (Yeast control - viability)
Application Note Yeast Analysis with a life - dead staining kit (Yeast control - viability) (CyFlow Cube 8 and CyFlow Cube 6 with CyView TM version 1.5 CyFlow Space) Introduction The Partec Yeast control
More informationCyFlow Cube series Appealing from every angle
CyFlow Cube series Appealing from every angle www.sysmex-partec.com CyFlow Cube 6 and Cube 8: compact, economic flow cytometers with a great performance Panta rhei a flexible solution for demands in flow
More informationImmunophenotyping of Peripheral Blood and Bone Marrow Cells by Flow Cytometry *Akanni EO and # Palini A.
Immunophenotyping of Peripheral Blood and Bone Marrow Cells by Flow Cytometry *Akanni EO and # Palini A. * Department of Haematology & Blood Transfusion,College of Health Science, Ladoke Akintola University
More informationInhibiScreen Kinase Inhibitor Assay
InhibiScreen Kinase Inhibitor Assay Flow Cytometric assessment of efficacy of PI3Kγ, PI3Kδ, BTK and SYK pathway inhibitors using the measurement of basophil activation in human EDTA and Heparin Whole Blood.
More informationIntroduction to Flow Cytometry. -- BD FACSCanto II TM. Daisy Kuo Assistant Product Manager BDBiosciences
Introduction to Flow Cytometry -- BD FACSCanto II TM Daisy Kuo Assistant Product Manager E-mail: daisy_kuo@bd.com BDBiosciences Outline Basic Concept of Flow Cytometry FACSCanto II System Introduction
More informationTECHNICAL BULLETIN. QUANTUM SIMPLY CELLULAR KIT Product Numbers QSC-20 AND QSC-100 Storage Temperature 2-8 C. Do Not Freeze
QUANTUM SIMPLY CELLULAR KIT Product Numbers QSC-20 AND QSC-100 Storage Temperature 2-8 C. Do Not Freeze TECHNICAL BULLETIN Product Description The Quantum Simply Cellular Kit provides a convenient method
More informationFor Reference Use Only
Exclusively Distributed in the US by: 26-G Keeywadin Drive, Salem NH, 03079 800-592-5726 www.alpco.com Small Sample Tube (CD20 -Pacific Blue TM / CD45-0C515 / CD8 + smig -FITC / CD56 + smig PE / CD4 PerCP-Cyanine5.5
More informationCyFlow Space. Your Flexible Flow Cytometer.
CyFlow Space Your Flexible Flow Cytometer www.sysmex-partec.com Ultimate Flexibility Modular System The CyFlow Space flow cytometer is a modular system with ultimate flexibility: from a basic configuration
More information