Hotlink to MACSQuant Photomultiplier Tube Calibration QuickStart
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1. Miltenyi Biotec Photomultiplier tube calibration Excite and inspire MACSQuant Analyzer quick guide Description In flow cytometry fluorescence intensity is used to distinguish between positive and negative populations of particles The reproducibility and stability of the fluorescence signal is of vital importance especially when performing comparable experiments over time As a quality control the MACSQuant Analyzer automatically adjusts voltage gains when performing photomultiplier tube PMT calibration with MACSQuant Calibrarion Beads to ensure that known fluorescent intensities are always set to the same channel This process is recommended to be performed every other day Required materials e MACSQuant Calibration Beads 130 093 607 e 12x75 mm 5 mL tube or microcentrifuge tube e MACSQuant Running Buffer 130 092 747 T ie ol E MEA Eg W Ww E e FA irei Dm ri y Ho oe ri rg 4 vi bid ei ll bos in a TE m muaf RA Th 7 MM a pull Aa An a rI s EHHH Fd ey Lj n s arel au Ti ii el li rl BH MED a4 fay A mgt af jarra EE cael a IE F 2H a3 orth a at wf aan Ion pon Taian EP Han ae he Fae iF aly iz a pat Figure 1 Calibration analysis template report Automated PMT calibration 1 Prime the MACSQuant Analyzer and wait a minimum of 30 minutes for warm up of optics Thoroughly vortex the MACSQuant Calibration Beads to break up any aggregates Click the ju icon a2. 0 c Check that the trigger is not set too high Move trigger down save over default setting and rerun calibration d Call technical service or initiate a MACSQuant Live Support session for assistance Samples Experiment Tools Channels Expenment Rack Single tube rack LI File adm2009 09 07 o fa oO Project A Sample ID F Description Flow rate Low Mediumi High Pickup and measure Liquid sensor C Mix sample Mode Standard w Uptake volume 150 pl LI Sample volume me 200 TARI E Annotations Autolabel settings C Custom Express Type Setup vw Mode Calibration v Figure 2 Experiment tab showing selection of calibration process in the settings tab Are you in need of additional assistance Visit www miltenyibiotec com local to find your nearest Miltenyi Biotec contact Miltenyi Biotec provides products and services worldwide Visit www miltenyibiotec com local to find your nearest Miltenyi Biotec contact MACS MACSQuant and MACSQuantify are registered trademarks or trademarks of Miltenyi Biotec GmbH Unless otherwise specifically indicated Miltenyi Biotec products and services are for research use only and not for therapeutic or diagnostic use Copyright 2010 Miltenyi Biotec GmbH All rights reserved 140 003 144
3. nd present the vial barcode when the reader begins to flash Note The reader will take a few seconds to recognize the vial When the barcode is recognized a dialog box will appear Select Yes to proceed with calibration process Follow the on screen instructions i e place an empty tube into the attached single tube holder and place one drop of the MACSQuant Calibration Beads into it Click OK to start the calibration process The MACSQuant will automatically dilute the Calibration Beads and start measurement Upon completion an analysis template will indicate that the calibration has passed Voltage gain staining index and fluorescence histogram plots figure 1 are displayed The MACSQuant Analyzer status bar should report Acquisition mode Calibration OK Manual PMT calibration 1 Prime the MACSQuant Analyzer and wait a minimum of 30 minutes for warm up of optics Ensure single tube holder is correctly attached 2 Thoroughly vortex the MACSQuant Calibration Beads to break up any aggregates 3 In custom mode select the Settings tab within the Experiment tab figure 2 4 In the Settings tab click the Express radio button 5 Select Setup from the Type drop down menu and Calibration from the Mode drop down menu 6 Dilute one drop of the MACSQuant Calibration Beads in 500 uL of the MACSQuant Running Buffer in a 12x75 mm tube Place the tube in
4. the single tube holder 7 Click the start measurement J icon or push the orange button on the front of the single tube holder This will start the calibration process 8 Upon completion an analysis template will indicate that the calibration has passed Voltage gain staining index and fluorescence histogram plots figure 1 are displayed 9 The MACSQuant Analyzer status bar should report Acquisition mode Calibration OK Optional The MACSQuant Analyzer can perform pre dilution and mixing of the MACSQuant Calibration Beads by setting preferences in the Autolabel tab For details refer to section 3 6 2 of the user manual Troubleshooting PMT calibration There could be multiple reasons why a calibration may not pass or be incomplete Listed below are some common reasons Calibration failed 1 High CV for fluorescence channels a Confirm optical bench has warmed up for at least 30 minutes b Laser alignment may have drifted Call technical service or initiate a MACSQuant Live Support session for assistance 2 High noise a Check for air in Pallfilter Rerun calibration b Run Clean Rerun calibration Miltenyi Biotec Calibration incomplete 1 No events acquired a Check calibration of needle arm to washing station For detailed information refer to section 3 5 of the user manual b Check that live events in Edit gt Options gt Software gt Acquire is set to at least 5 00
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