autoMACSPro Presentation
Contents
1. Daily Maintenance Priming Program Rinse Shut down Program Sleep for overnight storage fill with Ethanol A 5 Periodic maintenance Periodic Maintenance Exchange Columns Program Option Special Col ex Every 14 days or after 100 separations whichever comes first Clean pump syringe Every 1 2 month Long term storage Program Store substitute columns Cleaning of Tubing system also recommended for decontamination Program Safe cleaning procedure with hypochlorite MACS Bleach solution A 6 Cleaning exchange of Wear parts e Cleaning of pump syringe 7 3 2 e Cleaning of washing station 7 3 3 e Exchange of valves 7 4 1 are explained in details inthe user manual 7 Rescue procedure Described in User Manuel under 7 7 7 7 1 Rescue procedure A 1 Restart the instrument by switching it OFF and ON again 2 Undo the tubing connector at the negative port and place intoa 50 mL tube 3 Take out the uptake port needle fromthe needle holder and place it into a50 mL tube 4 Undo the tubing connector of the waste tube at the waste bottle and place it into a50 mL tube place a second 50mL tube beside this one 5 Run the program Qrinse This will rinse the complete fluidic system with autoMACS Pro Running Buffer eluting the cells into the 50 mL tubes Depending on which step of the separation program that the interruption occurred the cells will be found in any one of th2. 15 12 5 mL 15 mL Chill 3 50 50 mL 50 mL Reagent Rack 4 S Holds up to 4 vials for automated sample labeling cells 5x 108 2 5x 10 4x 10 Cooling of sample and sorted fractions to Sustain cell viability MiniS ampler lid for Sample protection Automatic recognition of different tube racks 2 MACS Buffer e Connects directly to the instrument e Controlled for cell separation performance e High quality ingredients Three Solutions are used to Running Buffer operate and maintain autoMACS Instruments A 2 Re usable columns e For 14 days or up to 100 separations Special cell friendly matrix coating Design no air contact e High capacity cell sorting within a few minutes 10 up to 4 x 10 total nucleated cells upto 2 x 10 magnetically labeled cells upto 15 mL of whole blood 2 autoMACSTM Technology Key benefits Based on MACS Technology utilizes 50nm MACS MicroBeads Proven quality worldwide since 1999 classic autoMACSTM 43 countries more than 1800 Instruments sold e Pioneer in automated cell sorting several hundred publications in cellular research e Renowned for time ef fective and easy sample sorting Minimal operator training required lower the workload for busy laboratories A 3 Before you start e Prepare single cell suspension Avoid cell clumps use pre f ilters Avoid excess of dead cells e Fo
3. Rinse from Menu Wash only Qrinse Quick Rinse 2 min rinse and refill with Running buffer Standard short rinse abundant cells _ Rinse 5 min rinse with W ashing solution rinse and refill with Running buffer Mandatory after sticky samples e g whole blood or tissues Recommended before rare cell isolations A 4 Easy operation with intuitive software Reagent menu load protocol information by scanning reagents Separation menu program separation sequence and start process Status menu monitor instrument status and progress of separation maasi 1a auae A ao Reagent Separation Status Log list Option 4 Bottle illumination display instrument status Code Green Blue Yellow Red Purple Blinking Status Ready for separation Instrument operating Not ready for separation Error Program Sleep is completed Action required User action No action required No action required Run wash program Rinse or Orinse Check screen for error correction Switch off autoMACS Pro Separator Check screen for required action 5 Routine maintenance Good sample preparation Avoid cell clumps use Pre Sep filters and excess of dead cells Choose corect separation amp wash program combination for best results QRINSE vs RINSE Rinse recommended before rare cell isolations between species sticky material e g whole blood bone marrow tissues
4. multiparameter sorting For virtually any cell type more than 250 separation reagents 2 AutoMACS is Based on MACS Technology Automated sensor controlled multisample labeling with MACS MicroBeads and subsequent cell sorting en i i e 2 Automated multisample cell sorting MACS MiniS ampler amp robotic arm Automated labeling option Cooling racks for 5 15 50 mL sample tubes Sensor controlled process Choice of 12 separation programs Automated sensor controlled sample loading and fraction elution Automated mandatory washing between samples Sensor controlled buffer supply 2 Key components I Touchscreen Robotic arm Barcode Reader Fluid container 4 Rack detection U PUNE 2 Key components Il 1 Robotic arm Uptake port outlet port positive fraction Outlet port negative fraction MiniS ampler guiding Memory card slot Power switch NO HD VI PP W 2 Key components Ill MACS MiniSampler Position of Cooling Tube Rack Position of Reagent Rack 4 MiniS ampler socket MiniS ampler lid guiding Plug connecting at the rear of the autoMACSTM Pro Separator U B WN Move Racks in x direction left right S Direct connection to the instrument S Lid for sample protection 2 Cooling Tube Racks IB Cooling Tube Racks Rack Max Max vol Max type no of per sample no of tubes Chill 5 6 5 mL 2 5 mL Chill 5
5. AutoMACS Pro Presentation Lausanne EPFL SV PTECH PTCF 9 6 2010 Alain Hirschy Application Specialist DACH Alainn miltenyibiotec de Agenda Introduction to MACS Technology Key components of the AutoMACS Pro How to work with the instrument Software and sensors Maintenance Wear parts cleaning exchange Rescue procedure Troubleshooting 1 Introduction to MACS Technology MACS MicroBeads are super paramagnetic particles of BE ey 50 nanometers in diameter being comparable to the size of a virus MicroBeads do not change the scatter properties of the cell in the flow cytometer or influence the light microscopic appearance of the cell They form a stable colloidal suspension and do not precipitate or aggregate in magnetic fields MACS MicroBeads are composed of a biodegradable matrix made of iron oxide and polysaccharide Hence it is not necessary to detach cell bound beads after the separation process saving hands on time Usually MACS MicroBeads do not alter structure function or activity status of labeled cells and they are not known to interfere with Subsequent experiments a isolated cells can be used directly for subsequent studies or A culture 1 MACS Technology Benefits MACS Column Technology Gentle isolation of viable functionally active cells Excellent purity recovery and viability Flexible cell sorting strategies e positive selection depletion
6. e vials 6 Combine all fractions and centrifuge at 350 xg for 10 minutes 7 Discard the supernatant and apply cells to a reseparation as soon as possible Keep cells on ice until the separation 8 Reconnect all tubing at the appropriate positions and reposition up take needle in needle holder 8 Troubleshooting General o What are the aim and the experiment o How often was the experiment performed before Did it work then Results o How many cells were found in the different fractions original positive negative o What was the frequency of target cells found in the different fractions original positive negative o Are flow data available Which gating was performed Where dead cells stained Protocol o Which sample material was used Which species Frozen or fresh sample How was the sample stored prior to the separation procedure o How was the cell suspension prepared Was the single cell suspension checked o Which MACS reagent was used Was it still within the shelf life o Which buffer was used composition o Which fluorochromes were used When was the fluorescent staining performed o How was the magnetic labelling performed time temperature amount of reagent total labelling volume o Which instrument program was used How many cells were loaded onto the column or instrument
7. llow labeling instructions in datasheet Same protocol used for manual and automated MACS cell separation 3 How to work with the Instrument 1 Choose a cell ln autoMACSTM Pro separation approach Separator 5 Program a 2 Prepare cell a separation template Sample up to six samples x 3 optional l manual labeling en o gt of cell samples 6 Run autoMACS Pro Cell Separation 7 Run Sleep or Store program for storage A 3 Choose a Separation Program Select cell populations according Eliminate cell population s Goal to particular cell surface antigen obtain untouched cells Positive selection Depletion Strat Magnetic labeling of target cells Magnetic labeling of cells ates Normal to high Rare cells or other than the target cells frequency cells purity increase Normal to POSSEL POSSELD DEPLETE high antigen Positive Double positive Depletion expression selection selection 0 5 mL Low antigen POSSEL S POSSELD2 DEPLETES expression Sensitive Double positive Sensitive depletion positive selection selection 2 mL POSSELDS DEPLO5 and DEPL025 E ana a Sensitive double Special sensitive depletions automatic Dilution 1 3 prior to uptake positive selection Stage loading up to 7 mL not POSSELWB A_Depl A_Depls Blood marrow Large scale depletion applicable to autolabelling Program 3 Priming Rinsing and Cleaning Priming Program
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