CyFlow ML CyFlow space

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1 CyFlow ML CyFlow space Immunology Haematology Pathology Microbiology Industrial Applications Cell Biology Cell Sorting Multilaser 16 9 Parameters Desktop FCM Systems

2 01 COMPANY Flow Cytometry made in Germany New sophisticated applications and increasing requirements for reliable results in research and routine within the shortest possible time. The challenge for Flow Cytometry instrumen tation, automation and software. Partec. The pioneer of Flow Cytometry. More than 40 Years of Experience and Professional Expertise Partec (established 1967) pioneer in Flow Cytometry since more than 40 years responds to these requirements with the newest generation of CyFlow analyzers and CyFlow sorters featuring innovative computer controlled fluidic systems, modular optical bench systems with advanced PMTs for all optical channels, most modern computer and digital electronic technologies as well as realtime data acquisition and realtime data display. In more than 100 countries worldwide 2

3 02 INTRODUCTION Excellence in Flow Cytometry Ultracompact desktop high-end instruments CyFlow ML Multilaser 16 Parameters 13 Colors Desktop Flow Cytometer CyFlow space 3-Laser 9 Parameters 7 Colors Desktop Flow Cytometer Dimensions (cm): L 56 x W 65 x H 30 (stand alone) _ 16 optical parameters: FSC1, FSC2, SSC, FL1 FL13 _ Up to 5 light sources _ 488 nm blue solid state laser (200 mw, 50 mw or 20 mw) _ 638 nm red diode laser (25 mw or 40 mw) _ 407 nm violet diode laser (100 mw) _ Choice of UV, green and yellow lasers _ High power UV LED for highest resolution DNA analysis _ Flexible and modular system configurations Dimensions (cm): L 56 x W 65 x H 30 (stand alone) _ 9 optical parameters 7 colors: FSC, SSC, FL1 FL7 _ Up to 3 laser light sources _ 488 nm blue solid state laser (200 mw, 50 mw or 20 mw) _ 638 nm red diode laser (25 mw or 40 mw) _ 407 nm violet diode laser (100 mw) _ 375 nm UV laser (16 mw) _ Other lasers optionally available (green, yellow, etc.) _ Cell sorter upgrade option Both feature: _ Parallel 16 bit digital pulse processing _ High fluorescence sensitivity: < 100 MESF (FITC) < 50 MESF (PE) _ Submicron particle detection < 0,2 µm for scatter _ RobbyWell Autoloader option _ True volumetric absolute counting _ Windows FloMax software for realtime data aquisition, data display and data evaluation _ Automated software compensation online and offline 3

4 Partec Quartz Flow Cuvette FORWARD SCATTER HIGH NA OBJECTIVES UV OR VIOLET DIODE LASER RED DIODE LASER BLUE SOLID STATE LASER SHEATH FLUID (HYDRODYNAMIC FOCUSING) SAMPLE The unique Partec quartz flow cuvette is the heart of the CyFlow ML/space, ensuring that particles and cells cross the excitation light with the best possible precision. The design of the flow cuvette incorporates more than 40 years of experience in handling fluids with sub-micrometer and nanoliter precision. Thanks to the optical and mechanical precision of the flow cuvette, superior results are guaranteed for all parameters, e.g. coefficients of variation (CVs) of about 2% on all fluorescence channels. The sample is transported with the help of a computer controlled syringe pump, part of a virtually cross-contamination-free fluid system. CyFlow Sorter as built-in module for safe closed piezo sorting offering highest purity and sorting stability. The Partec CyFlow Sorter is the result of more than 40 years of experience in the field of Flow Cytometry for research and routine applications in cell and particle characterization. The sorter is used as an add-on to the Partec CyFlow space and serves as an optimized module for simplified and precise cell sorting. In order to overcome the problems of droplet sorters (biohazard, contamination) Partec has developed this closed sorting system. The sorting device can be sterilized safely. Any environmental contamination is completely avoided under normal operation conditions. In addition, the sorting action takes place smoothly. Fragile cells or large particles are sorted without any distortion and other forces acting on the cells during deflection. Long sorting runs (for several hundred milliliters of sample volume) are possible. The Partec Sorter has a Flow Chamber with a piezo element and an electronic device for piezoelectric activation. The diamond piezo deflects the sorted cells into a second branch of the flow cell (see sorting scheme opposite page above). 4

5 CyFlow Sorter Module DIAMOND PIEZO FOR CLOSED CELL SORTING During piezoelectric activation, a pressure wave is generated and, as a consequence, a small volume of sheath fluid is deflected together with the cell or particle of interest. The piezo activation of the sorting process is repeated each time there is a cell to be sorted, according to the acquisition signals. In the sample flow each cell is analyzed and all the signals reach the computer, where the decision is made which cell should be sorted, according to any software gating. Special feature of the CyFlow Sorter is the correct sorting of ALL the cells in a region. The high yield is a result of the practically zero dead time data acquisition system. No event is lost because all Partec hard- and software acquisition work without any interrups. In addition, in this way no wrong cell can be sorted by mistake, achieving directly highest purity. The Partec sorting principle is utmost stable, lasting, constant, and without vibrations, because the whole system is closed, non-elastic, and without air. The delay time setting can be controlled easily on the CCD camera screen, no microsopic check is needed. The Partec sorter works with low pressure differences in the capillary system. The morphology and vitality of cells (for e.g. most fragile particles like plant protoplasts) are not influenced by sorting. As far as the control of the sorting process is concerned, Partec instruments have the advantage of an extremely precise and stable system control of the time delay between measurement and piezo activation. Another unique feature of Partec instruments is the built-in CCD camera, which allows an easy check of the whole process directly in the monitor of the instrument, this is also maintained while sorting (See the photograph above: CCD camera shot of the sorting process. Sorted cells are deflected to the upper right). Sorting flow cell specifications Completely closed quartz type flow cell with 200 microns sorting channel for particle sizes up to 60 microns diameter (other sorting channel dimensions optional). Sorting action Piezo crystal for controlled particle deviation. Adjustable delay between point of analysis and deflection. Sorting Control Unit Setting of sorting parameters for piezo crystal energy, delay, slope, and hold time. Direct connection to processor. Sorting speed Input up to 10,000 cells per second, for highest purity (99%) of sorted fraction up to 100 cells per second (300 cells per second at 95% puritiy). 5

6 Laboratories demand highest flexibility for upcoming applications. The CyFlow ML/space open system architecture is designed for today and the future. Taking advantage of most recent computer and electronic technologies Each optical channel is equipped with an independent processor controlled digital pulse analyzer for parallel pulse processing in order to minimize signal losses due to coincidence and deadtime effects. 16 bit (65536 channels) analogue-to-digital converters for each parameter are the fundament for software-based color crosstalk compensation algorithms and other numerical operations, e.g. fluorescence ratio measurements, with out artifacts. The computing power of the CyFlow ML/space system allows signal analysis, processing and display of each event generated by a particle in realtime, which is a prerequisite for precise high speed analysis and accurate absolut counting. All-in-one design and modular extension for future cytometric analysis The CyFlow ML/space in its ultracompact design incorporates all modules and components within smallest dimensions of L 56 x W 65 x H 30 [cm]. In contrast to other available FCM systems, the CyFlow line of instruments does not require any external equipment. Power supply, pressure regulators and the fluidic system are completely built-in. Additional space beside or under your laboratory desk is not any longer required. In order to be prepared for future applications, the CyFlow ML/space offers flexible and modular configurations in a most compact system architecture. 6

7 The True Volumetric Absolute Counting (TVAC) is a unique feature of all Partec Flow Cytometers, offering highest absolute counting precision and accuracy. The CyFlow ML/space features True Volumetric Absolute Counting for precise cell and particle enumeration. This unique method is based on the fundamental definition of absolute counts respectively the particle concentration (c = N/V). In the CyFlow ML/space, the sample volume is measured directly by mechanical means, rather than by calibration with oftenly expensive beads with a given nominal concentration. Thus, the volume measurement is eliminating any error influence related to eventually varying bead concentrations or bead aggregations. The CyFlow ML/ space allows the analysis of a fixed volume as defined by the distance between two platinum electrodes. Alternatively, a well defined volume of free choice involving the digital sample speed control can be used. Benefits of True Volumetric Absolute Counting: _ digital volumetric precision by mechanical design: CV < 2% _ no errors related to calibration _ no additional time and preparation steps _ no expenses for calibration beads _ no separate haematology counter required Selection of Fluorochromes for the CyFlow ML/space Light Source System Excitation (nm) Detector Exemplary Dyes Exemplary Dyes Exemplary Dyes Exemplary Dyes Exemplary Dyes Exemplary Dyes Exemplary Dyes Exemplary Dyes Blue 488 Green FITC GFP Alexa Fluor 488 Syto 9-24 Oregon Green JC-1 (monomers) DiOC6(3) H2-DCF-DA Orange PE YFP Hydroethidine Orange Red PE-Texas Red PI ECD EB JC-1 (aggregates) Red I PE-Cy5 PerCP PE-Dy647 Acridine Orange 7-AAD Red II PE-Cy5.5 PerCP-Cy5.5 Far Red PE-Cy7 Red 638 Red I APC APC-Cy5 Syto Dy647 TO-PRO3 Alexa Fluor 647 Draq5 Cy5 Red II APC-Cy5.5 Cy5.5 Far Red APC-Cy7 APC-H7 Alexa Fluor 750 Cy7 Violet 407 Blue Pacific Blue Alexa Fluor 405 Monobromobimane Hoechst Green AmCyan CFP Qdot 525 Lucifer Yellow Orange Cascade Yellow Pacific Orange Qdot 585 UV LED 365 UV Laser Blue DAPI Hoechst Alexa Fluor 350 Cascade Blue BFP AMCA Indo-1 (Ca ++ ) Green Qdot 525 Orange Qdot 585 Indo-1 (no Ca++) Green 532 Orange mstrawberry DsRed DY590 morange Red mcherry LDS 751 Yellow 561 Orange PE Dy590 Philippin Cy3 mbanana Red PE-Cy5 PI LDS 751 Orange 594 Orange Red Texas Red Alexa Fluor 594 Red APC mcherry Cell Tracker Red Far Red APC-Cy7 mplum mraspberry mkate Katushka HCRed HCRed 7

8 03 OVERVIEW Fields of Applications Overview of applications and upgrade options for CyFlow ML and CyFlow space CyFlow ML CyFlow space Healthcare Immunology, Hematology, Pathology, Cancer Research, DNA Analysis Microbiology Cell Counting & Live/Dead Analysis in Biotechnology and Cell Culture, Toxicology, Biomonitoring, Virus Detection, Research Industrial Applications Quality Control in Food and Beverage Industry, Yeast Analysis, Fermentation Control, Process Optimization, Particle Counting, Particle Size Distribution Agrosciences Ploidy Analysis, Plant Genome Size, Apomixis Cell Counting eukaryotic cell cultures (mammalian cell cultures plant cell cultures), prokaryotic cell cultures, leukocyte counting in depleted blood products, counting of immunolabelled subtypes, sperm cell counting Functional Assays cell cycle and cell proliferation, viability Live/Dead cell counting, apoptosis Wine Industry Rapid and Automatic Detection, Quantification and Live/Dead Analysis of Yeasts in Wine & Champagne HIV Monitoring and AIDS Patient Follow-up Diagnostics Dedicated, Accurate and Affordable CD4 and CD4% Testing for Adults and Children Cell Sorting Options & Upgrades RobbyWell Autoloader RobbyWell Autoloader CyFlow Sorter for closed and non-hazardous, non-destructive cell and particle sorting Notes High-End FCM system for a wide range of applications High-End FCM system for a wide range of applications and cell sorting 8

9 04 ANALYZE CyFlow ML/space FCM System The Windows FloMax software integrates instrument control including acquisition, on- and offline data analysis, onand offline compensation into a complete software package. Ready prepared and freely adaptable instrument settings and panels facilitate switching between different applications. FloMax is optimized for immunophenotyping, microbiology analysis, cell cycle, DNA ploidy, and scientific flow cytometric analysis. Data are stored in FCS Flow Cytometry standard file format for easy exchange with other analysis software. One of the unique features is the digital on- and offline color crosstalk compensation of the spectral overlap of fluorescence from simultaneously analyzed dyes. The N-color compensation algorithm allows a correction of the crosstalk between any parameters without the need of rerunning a sample. FloMax optimally supports the True Volumetric Absolute Counting feature of the CyFlow ML/space, displaying particle concentrations for any subsets of cells, even if defined by a gate at a later time after the acquisition. Full flexibility and automation by the multi-tube FloMax panel system and the FloMax Report system within the FloMax software package. The flexible FloMax Panel system allows automated analysis of repeating sample series employing different dyes or instrument settings. The FloMax Report system generates easy-to-adapt automated single or multi-tube panel results. Reports are based on Microsoft Word or Excel written in Visual Basic. 9

10 05 SPECIFICATIONS CyFlow ML/space FCM System GENERAL _ compact flow cytometer for automated sequential analysis of single cells and microscopic particles _ scatter particle size range: 0.2 µm 200 µm _ fluorescence sensitivity: <100 MESF (FITC) < 50 MESF (PE) _ fluorescence resolution: CV < 2% _ configurations with up to 16/9 optical parameters + time parameter LIGHT SOURCES _ up to 5/3 light sources _ blue solid state laser: 200 mw@488 nm _ red diode laser: 40 mw@635 nm _ violet diode laser: 100 mw@407 nm _ green DPSS laser 30/100 mw@532 nm _ yellow solid state laser 100 mw@561 nm _ UV LED 365 nm or ultra-violet diode laser 16 mw@375 nm _ other laser power and laser wavelengths available OPTICS _ modular optical system with up to 16/9 optical parameters with selected PMTs with integrated electronic preamplifier for CyFlow ML: FSC1, FSC2, SSC, FL1-FL13 CyFlow space: FSC1, SSC, FL1-FL7 _ standard setup and filters _ color CCD camera for video flow monitor _ standard objective mount with high numerical aperture _ immersion gel coupling, e.g. for detection of weak cytokines (option) _ separated intermediate image planes for optimized spatial filtering by diaphragms FLOW SYSTEM _ synthetic quartz flow cuvette for laminar sample transport with sheath fluid _ sample port with computer controlled BioSafety cleaning system, avoids sample droplets and minimizes cross contamination _ True Volumetric Absolute Counting based on mechanical volume measurement, no need for reference particles _ contamination-free computer controlled precision syringe pump for sample transport and True Volumetric Absolute Counting, pump speed continuously adjustable from 0-50 µl/s, sheath fluid pressure continuously adjustable from mbar _ easily accessible sheath fluid and waste reservoirs with fluid level sensors ELECTRONICS _ parallel signal processing for each of the optical channels with selectable linear, 3- or 4-decade logarithmic scale _ pulse height, area, and width analysis for doublet discrimination _ 16 bit analog-to-digital converters, trigger on any parameter or parameter combinations COMPUTER _ latest industry standard computer _ 19" TFT LCD display _ CCD video camera _ dual screen setup (optional) _ DVD-RW _ keyboard, mouse, barcode reader (optional) _ 100 MB/s and 1000 MB/s Ethernet connection _ DeskJet color printer, b&w or color laser printer (optional), printing via network SOFTWARE _ Microsoft Windows operating system with full network support _ Apple Macintosh network connection (optional) _ 32 bit Windows FloMax software for routine and research applications _ 64 parameter realtime data acquisition and analysis _ 3-spot acquisition from spatially separated light sources with time-window delay system _ internal floating point numerics with double precision _ one and two parameter histograms and dotplots _ channels resolution for 1P histograms _ 32/ /1024 channels for 2P dotplots _ multiparameter N-colors crosstalk compensation, settings may be corrected at any time _ multiparameter ColorGating _ ratio measurement _ doublet discrimination based on pulse height, area, and width analysis _ peak and cluster analysis and statistics _ connection to 96 well plate automate _ DNA cell cycle analysis _ DNA peak analysis _ multitube panel system with automated acquisition _ Flow Cytometry standard data format (FCS) for storage of original and evaluated data _ storage of gates and screen layout _ automated data transfer of graphics, statistics and instrument settings to Word, Excel, PowerPoint and other desktop publishing systems or laboratory information systems (LIS) Length 560 mm Width 650 mm Length 560 mm Width 650 mm Height 300 mm Height 300 mm 10 For in vitro diagnostic use.

11 High Quality Made in Germany Quality, performance, precision and affordability of Partec products profit from a unique production depth realized at Partec for manufacturing modules and components in optics, electronics, laser technology, fluidics, and mechanics. 11

12 Contact Partec GmbH Otto-Hahn-Straße 32 D Münster Germany Phone +49 (0) Fax +49 (0) Partec GmbH Am Flugplatz 13 D Görlitz Germany Phone +49 (0) Fax +49 (0) Partec North America, Inc. 603 Heron Dr., Unit 9 Swedesboro, NJ USA Phone US toll-free Fax partecna@partec.com Partec Japan, Inc Kandatsu Tsuchiura Japan Phone Fax partecjapan@partec.com Partec S.A.R.L. 14/16 rue Gallieni Sainte Geneviève des Bois France Phone Fax partecfrance@partec.com Partec Italia S.r.l. Via. G. Mascherpa Carate Brianza Italia Phone Fax partecitalia@partec.com Partec UK Ltd Unit 44 Canterbury Innovation Centre University Road Canterbury Kent CT2 7FG United Kingdom Phone Fax partecuk@partec.com Partec Nederland BV Middenweg 591 B 1704 BH Heerhugowaard Nederland Phone Fax partecnederland@partec.com Partec India E-135, Ground Floor Lajpat Nagar I New Delhi India Phone Fax partecindia@partec.com Partec West Africa SARL 01 BP 1323 Ouagadougou Burkina Faso Phone Fax pwa@partec.com Partec Afrique Centrale SARL Rue Bebey Elame AKWA Douala B.P Douala Cameroon Partec East Africa Ltd N.H.I.F. Building, 2 nd Floor Ragati Road Code Uhuru Gardens Nairobi Kenya Partec Southern Africa (Pty) Ltd Unit IF Plot 119 Millenium Park Gaborone Botswana Complete list of subsidiaries and distributors: Phone Phone Fax pac@partec.com Phone Fax pea@partec.com Phone Fax psa@partec.com Medical Diagnostics Diagnostics for HIV/AIDS, TB, Malaria Microbiology/ Industrial Applications Agrosciences/Animal Research/Aquaculture