Fluidigm CyTOF XT User Manual
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Fluidigm CyTOF XT User Manual

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FLDM-00254 Rev 03

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  • Page 1 FLDM-00254 Rev 03...
  • Page 2 © 2021 Fluidigm Corporation. All rights reserved. 07/2021 For technical support visit techsupport.fluidigm.com. North America +1 650 266 6100 | Toll-free (US/CAN): 866 358 4354 | support.northamerica@fluidigm.com Latin America +1 650 266 6100 | techsupportlatam@fluidigm.com Europe/Middle East/Africa/Russia +33 1 60 92 42 40 | eu.support@fluidigm.com Japan +81 3 3662 2150 | techsupportjapan@fluidigm.com...

  • Page 3: Table Of Contents

    Contents About This User Guide Chapter 3: Annotated Figures of CyTOF XT Components Safety Alert Conventions Main Components of the CyTOF XT Instrument Safety Alerts for Chemicals CyTOF XT Status Panel Safety Alerts for Instruments Autosampler Module Safety Data Sheets...
  • Page 4 CyTOF XT Shutdown Procedure CyTOF XT Startup Procedure Appendix C: Preventive Maintenance, Decontamination, and Disposal Preventive Maintenance Recommendations Clean and Decontaminate the CyTOF XT System 139 Clean the System Dispose of Products Appendix D: Related Documents Appendix E: Revision History...

  • Page 5: About This User Guide

    CyTOF Software v8.0 Help for CyTOF XT (FLDM-00045). The software Help also includes an end-to-end daily workflow. For complete instrument specifications and pre-unpacking/installation instructions, see the CyTOF XT System Site Requirements Guide (FLDM-00360).

  • Page 6: Safety Alerts For Instruments

    Safety Data Sheets Read and understand the SDSs before handling chemicals. To obtain SDSs for chemicals ordered from Fluidigm, either alone or as part of this system, go to fluidigm.com/sds search for the SDS using either the product name or the part number.

  • Page 7: Chapter 1: Product Overview

    The Autosampler and CyTOF XT software increase productivity with built-in clog detection, automated declogging, and shutdown. Together, the full range of new features on CyTOF XT maximizes both the breadth and depth of information acquired per sample and accelerates high-parameter studies with enhanced instrument productivity.

  • Page 8: Cytof Xt Site Requirement Specifications

    3.6/12276 * One dedicated electrical branch circuit (single-phase, 30 A single-phase 200–240 V AC, 50/60 Hz) is used to provide power to the CyTOF XT instrument. † For optimal performance the temperature should be 20 ±2 °C. CyTOF XT Specification...

  • Page 9: Workstation Specifications

    ( in) LED Keyboard/mouse Wired Data File Size Specifications File Type Size (up to) .fcs 4 bytes/channel/event .lmd Compressed format. 5–20 GB/hr (typical) _SPR.fcs 3.82 MB/hr _Beads.fcs 0.91 MB/hr (typical) _BG.fcs 0.48 MB/hr _Processed.fcs 4 bytes/channel/event CyTOF XT User Guide...

  • Page 10: Instrument And Computer Connections

    Chapter 1: Product Overview Instrument and Computer Connections Instrument and Computer Connections Figure 1. Computer rear view CyTOF XT User Guide...

  • Page 11: Cytof Xt Equipment And Consumables

    Part Number Standard Equipment* 117002 CyTOF XT™ instrument 108490 Monitor, standard 34-inch LG 118783 CyTOF XT computer, including keyboard and mouse 130069 CyTOF Software v8.0 * Standard equipment is subject to change at sole discretion of Fluidigm. CyTOF XT User Guide...

  • Page 12: Standard Consumables

    Autosampler Probe Line CyTOF XT 118763 Autosampler Sample Loop CyTOF XT 117784 Autosampler Nebulizer Line CyTOF XT 101815 Reducer O-Ring 2-113 CyTOF XT – pack of 5 1 pack 130396 Sampler O-Ring 2-033 CyTOF XT – pack of 10 1 pack 117174...

  • Page 13: Cytof Xt Special Tools And Kits

    Autosampler Sample Loop Agitator CyTOF XT 118342 Autosampler Waste Bottle Cap CyTOF XT 132224 Autosampler Waste Bottle Cap O-Ring Black Size 2-224 CyTOF XT – pack of 5 132161 Autosampler PCB Air Filter—Open Cell, 1/4" Thick GRAY CyTOF XT 118744...

  • Page 14: Required Consumables From Other Suppliers

    Consumables Ordering North America Customers in the US and Canada who have a Fluidigm account are already registered for online ordering. Go to fluidigm.com/catalog. New customers can set up an online account to place orders, view past order history, and see current order confirmations.

  • Page 15: Regulatory Compliance

    Indicates conformity with safety requirements for Canada and the United States. Indicates conformity with European Union requirements for safety and electromagnetic compatibility Refer to the CyTOF XT Site Requirements Guide (FLDM-00360) for more detailed information on the recommended environmental conditions. CyTOF XT User Guide...

  • Page 16: Chapter 2: Mass Cytometry And Cytof Xt

    Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT Instrument Overview Figure 3. CyTOF XT front view CyTOF XT User Guide...

  • Page 17: Cytof Xt System Components Overview

    Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT System Components Overview CyTOF XT System Components Overview Autosampler Module The Autosampler Module of the CyTOF® XT system consists of 4 major components: the carousel, the probe line, the pump and valve unit, and the bottle station. Together, these...
  • Page 18 Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT System Components Overview Carousel Figure 5. The Autosampler carousel The carousel housing contains a removable carousel, which can hold 16 tubes. Locations 1–12 are for polypropylene sample tubes (5 mL). •...
  • Page 19 Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT System Components Overview The corner lid is fixed and is removed only when performing maintenance inside the carousel. Figure 6. Autosampler carousel housing Probe Line The probe line draws samples and beads from the carousel into the Autosampler fluidics lines.
  • Page 20 Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT System Components Overview The probe line has 3 positions: Raised and vertical: This is the default rest position for the probe line. • Figure 8. The Autosampler probe line: raised and vertical Raised and horizontal: When the probe line is raised, you can move it to a horizontal •...
  • Page 21 Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT System Components Overview • Lowered and vertical: The probe line is in this position when drawing sample into the Autosampler fluidics lines. Figure 10. The Autosampler probe line: lowered and vertical...
  • Page 22 Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT System Components Overview Bottle Station The bottle station contains 4 bottles. Left to right, they are waste (1 L), Maxpar® Cell Acquisition Solution Plus (Maxpar CAS Plus) (1 L), Maxpar Washing Solution (Maxpar WASH) (500 mL), and Maxpar Water (500 mL).
  • Page 23 Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT System Components Overview Pump and Valve Unit The pump and valve unit contains the Autosampler fluidics lines, where samples loaded into the carousel are automatically introduced into the system, allowing unattended instrument operation and sample acquisition.
  • Page 24 Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT System Components Overview Syringe Pumps The 2 syringes on the pump and valve unit alternate between wash and sample introduction functions. The right fill-pump syringe contains a 3-port valve that connects the syringe to the waste bottle and the switching valve.

  • Page 25: Autosampler Module Functions

    250 µL has been injected. When less than 250 µL of sample is being loaded, the rest of the sample loop remains filled with Maxpar CAS Plus. This function is also used to inject EQ6 beads during tuning. Figure 15. CyTOF XT™ flow diagram of the sample loading process CyTOF XT User Guide...
  • Page 26 Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT System Components Overview Figure 16. CyTOF XT flow diagram of the sample injection process Acquisition of Pelleted Samples The Autosampler can resuspend pelleted samples with Maxpar CAS Plus and add EQ beads prior to acquisition.
  • Page 27 For more recommendations to reduce clogs and manual intervention techniques during acquisition or after acquisition stops, click Help to see the integrated help guide, also available as the CyTOF Software v8.0 Help for CyTOF XT (FLDM-00045). CyTOF XT User Guide...

  • Page 28: Nebulizer Assembly

    The CyTOF XT instrument employs a concentric glass nebulizer consisting of an inner capillary that carries the liquid sample and an outer chamber that carries argon gas (called nebulizer gas).
  • Page 29 Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT System Components Overview Figure 18. The nebulizer assembly The nebulizer holder and the nebulizer mount complete the gas and liquid connections to the nebulizer when it is fully assembled. When not in use, the nebulizer can be placed in the nebulizer rest located above the pump and valve unit on the instrument.

  • Page 30: Heater Assembly

    Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT System Components Overview Heater Assembly Overview The heater assembly is comprised of a heater unit, the spray chamber, and the injector holder, secured by the heater assembly cap. The injector is inserted into the injector holder, which is threaded onto the heater assembly cap.

  • Page 31: Ionization

    Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT System Components Overview Spray Chamber The spray chamber is housed within the heater assembly. Aerosolized sample exits the nebulizer directly into the spray chamber. Makeup gas (argon gas) is pumped into the spray chamber, which heats the sample to 200 ˚C as it passes through the spray chamber.
  • Page 32 Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT System Components Overview Plasma Torch The plasma torch consists of 3 concentric chambers: 2 concentric quartz tubes that are combined in the demountable torch assembly, and a quartz sample injector tube that is inserted inside the torch assembly.
  • Page 33 Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT System Components Overview The torch assembly is mounted inside an induction load coil supplied with current generated by radiofrequency (RF) power. This current generates the electromagnetic field that sustains the plasma.

  • Page 34: Vacuum Interface

    –3 Torr. CyTOF XT uses a 3-cone interface to transport the ion beam into a low-pressure vacuum: sampler, skimmer, and reducer. The 3 cones are made of nickel and the sampler and skimmer cones have a platinum-coated tip. The plasma containing the cell-induced ion...
  • Page 35 Chapter 2: Mass Cytometry and CyTOF XT CyTOF XT System Components Overview High-Pass Ion Optics The ion cloud leaving the vacuum interface is dominated by low mass argon (Ar) ions from the plasma. These ions are not of analytical interest and are of such high abundance that they may quickly damage the detector.

  • Page 36: Event Detection And Acquisition Data File Format

    (the lowest mass registered) to be recorded per push, while the segment length dictates the mass range to be recorded per push. Instruments are set to collect data from up to 135 mass channels (each corresponding to 1 amu), starting at mass 75 for CyTOF XT. CyTOF XT User Guide...

  • Page 37: Data Acquisition And Analysis

    The range of data that CyTOF XT collects requires collection of dual data, which means that pulse count and intensity values are collected for every channel. CyTOF Software plots the entire data range on a single dual signal scale, the units of which are actual counts of particles that hit the detector.

  • Page 38: Chapter 3: Annotated Figures Of Cytof Xt Components

    Chapter 3: Annotated Figures of CyTOF XT Components Main Components of the CyTOF XT Instrument Figure 29. Main components of the CyTOF® XT instrument CyTOF XT User Guide...

  • Page 39: Cytof Xt Status Panel

    Chapter 3: Annotated Figures of CyTOF XT Components Main Components of the CyTOF XT Instrument CyTOF XT Status Panel The color of the rectangle LED light indicates the status of the instrument. NOTE The front lid of the instrument must be closed to see the status panel.

  • Page 40: Autosampler Module

    Chapter 3: Annotated Figures of CyTOF XT Components Main Components of the CyTOF XT Instrument Autosampler Module Figure 32. Main components of the Autosampler Module Pump and Valve Unit Figure 33. Autosampler Module pump and valve unit CyTOF XT User Guide...
  • Page 41 Chapter 3: Annotated Figures of CyTOF XT Components Main Components of the CyTOF XT Instrument Probe Line Figure 34. Autosampler Module probe line CyTOF XT User Guide...
  • Page 42 Chapter 3: Annotated Figures of CyTOF XT Components Main Components of the CyTOF XT Instrument Carousel Housing Figure 35. Autosampler Module carousel housing Figure 36. Autosampler Module carousel CyTOF XT User Guide...
  • Page 43 Chapter 3: Annotated Figures of CyTOF XT Components Main Components of the CyTOF XT Instrument Nebulizer Rest Figure 37. Autosampler Module nebulizer rest Bottle Station Figure 38. Autosampler Module bottle station CyTOF XT User Guide...

  • Page 44: Nebulizer Assembly

    Chapter 3: Annotated Figures of CyTOF XT Components Main Components of the CyTOF XT Instrument Nebulizer Assembly Figure 39. The nebulizer assembly Figure 40. The nebulizer assembly, expanded view CyTOF XT User Guide...

  • Page 45: Heater Assembly

    Chapter 3: Annotated Figures of CyTOF XT Components Main Components of the CyTOF XT Instrument Heater Assembly Figure 41. The heater assembly Figure 42. The heater assembly, expanded view CyTOF XT User Guide...

  • Page 46: Torch Assembly

    Chapter 3: Annotated Figures of CyTOF XT Components Main Components of the CyTOF XT Instrument Torch Assembly Figure 43. Torch assembly Figure 44. Torch assembly, expanded view CyTOF XT User Guide...

  • Page 47: Interior View Of Cytof Xt

    Chapter 3: Annotated Figures of CyTOF XT Components Main Components of the CyTOF XT Instrument Interior View of CyTOF XT Interface Assembly Figure 45. Interface assembly Torch Box Figure 46. Torch box CyTOF XT User Guide...

  • Page 48: Other Components Of Cytof Xt

    Chapter 3: Annotated Figures of CyTOF XT Components Main Components of the CyTOF XT Instrument Other Components of CyTOF XT Part Image Location AC distribution box Right side of CyTOF XT™ system Sampler cone Inside the vacuum interface Skimmer-reducer Inside the vacuum interface...
  • Page 49 Chapter 3: Annotated Figures of CyTOF XT Components Main Components of the CyTOF XT Instrument Part Image Location Nebulizer Nebulizer assembly Nebulizer holder with Nebulizer assembly nebulizer Outer torch Torch assembly Inner torch Torch assembly CyTOF XT User Guide...
  • Page 50 Chapter 3: Annotated Figures of CyTOF XT Components Main Components of the CyTOF XT Instrument Part Image Location Spray chamber Heater assembly Injector holder Heater assembly Injector Heater assembly CyTOF XT User Guide...
  • Page 51 Chapter 3: Annotated Figures of CyTOF XT Components Main Components of the CyTOF XT Instrument Part Image Location Injector cap Heater assembly CyTOF XT User Guide...

  • Page 52: Chapter 4: Maintenance

    Always wear non-latex powder-free gloves when working with the instrument and handling its parts. IMPORTANT For rinsing after cleaning the CyTOF XT™ system and its parts, we recommend using only Type 1 ultrapure (18.2 MΩ) water (DIW). Table 1. CyTOF XT components requiring weekly cleaning by operator...

  • Page 53: Clean The Nebulizer

    Chapter 4: Maintenance Clean the Nebulizer Table 2. CyTOF XT tools provided by Fluidigm for use during maintenance procedures Equipment List Source Part Number Torch removal tool Fluidigm 117634 Cone removal tool Fluidigm 105624 Torch verification tool Fluidigm 118383 No. 2 Phillips screwdriver* Various —...

  • Page 54: Remove The Nebulizer

    Clean the Nebulizer Remove the Nebulizer Disconnect the nebulizer gas line by pulling the quick-connect coupling. Disconnect the nebulizer line by unscrewing the fitting from the nebulizer assembly. Loosen the 2 wing nuts by turning them counterclockwise. CyTOF XT User Guide...
  • Page 55 Ensure that if the plasma is on, the nebulizer port safety shutter completely covers the nebulizer port. Pull the nebulizer release collar away from the nebulizer to remove the nebulizer holder from the nebulizer mount. CyTOF XT User Guide...

  • Page 56: Materials Required For Cleaning

    Label a 5 mL round-bottom tube and fill it with approximately 3 mL of 10% Contrad/Decon. Lubricate one end of Tubing 1 (larger diameter tubing) from the Nebulizer Cleaning Kit with 10% Contrad/Decon. Connect the side arm of the nebulizer to the syringe with Tubing 1. CyTOF XT User Guide...
  • Page 57 DIW before use. The nebulizer is ready for immediate use. If the nebulizer is not to be used immediately, submerge it in DIW until the next use. CyTOF XT User Guide...

  • Page 58: Cleaning Method For Manual Clog Removal: Hot Diw

    (Optional) Visually inspect the nebulizer using a light microscope to verify that the clog has been removed. Check the nebulizer spray. Reassemble the nebulizer: Connect the nebulizer gas line and the nebulizer line to the nebulizer assembly. CyTOF XT User Guide...
  • Page 59 Observe the spray from the nebulizer using a flashlight. It should appear as a fine aerosol that leaves the nebulizer in an even, symmetrical V-pattern. Keep the reassembled nebulizer connected and stored in the nebulizer rest for immediate use or store the nebulizer submerged in DIW until the next use. CyTOF XT User Guide...

  • Page 60: Clean The Nebulizer Rest

    CAUTION Hot surface hazard. The torch components, the vacuum interface, and the heater assembly components remain hot for some time after the plasma has been shut off. Allow sufficient time, at least 20 min, for these items to cool to room temperature before you handle them. CyTOF XT User Guide...

  • Page 61: Remove The Heater Assembly

    Gently slide the heater assembly away from the instrument. CAUTION Finger cut hazard. Broken glass may cause injury or cutting of fingers or hands. To avoid breakage, keep the heater assembly straight while removing and replacing. CyTOF XT User Guide...

  • Page 62: Remove The Injector Cap And Injector

    Chapter 4: Maintenance Clean the Injector and Spray Chamber Place the heater assembly into its holder on the left side of the instrument. Remove the Injector Cap and Injector Unscrew and remove the injector cap. CyTOF XT User Guide...

  • Page 63: Clean The Injector

    Rinse thoroughly with DIW. Spray isopropanol inside the injector to hasten drying. Allow parts to dry thoroughly at room temperature. Inspect the O-ring on the injector cap for damage or discoloration. Replace the O-ring if necessary. CyTOF XT User Guide...

  • Page 64: Remove The Spray Chamber And Injector Holder

    A plastic pipette tip can be used to help remove and replace the injector cap O-ring. Remove the Spray Chamber and Injector Holder With the heater assembly in its holder, gently push down and then turn the heater assembly cap to the unlock position. CyTOF XT User Guide...
  • Page 65 Remove the spray chamber and injector holder from the heater assembly cap. IMPORTANT Carefully remove the spray chamber since the injector holder may loosen from the heater cap and fall out. CyTOF XT User Guide...

  • Page 66: Clean The Spray Chamber And Injector Holder

    CAUTION Hot surface hazard. The torch components, the vacuum interface, and the heater assembly components remain hot for some time after the plasma has been shut off. Allow sufficient time, at least 20 min, for these items to cool to room temperature before you handle them. CyTOF XT User Guide...

  • Page 67: Remove Cones

    Pull the door open fully to access the cones. NOTE Remove the sampler cones using the cone removal tool, which has 2 ends, one to remove the sampler cone and the other to remove the skimmer-reducer assembly. CyTOF XT User Guide...
  • Page 68 Use O-ring removal tool to remove the damaged sampler cone O-ring and replace (105704). Remove the sampler cone from the cone removal tool and place it on a clean surface with the tip facing up. NOTE Place the tip up to avoid damage to the cone orifice. CyTOF XT User Guide...
  • Page 69 Remove the skimmer-reducer assembly using the cone removal tool. Insert the other end of the cone removal tool (with 2 magnets) into the 2 holes on the skimmer-reducer assembly. Turn the cone removal tool counterclockwise to unscrew the skimmer-reducer assembly from the vacuum interface. CyTOF XT User Guide...

  • Page 70: Clean Cones

    Add 10% Citranox as each part is placed in the container to ensure that all parts contact the cleaning solution. Stack the bottom adapter first. Place the sampler cone tip face down on the bottom adapter. CyTOF XT User Guide...
  • Page 71 Add DIW as each part is placed in the container to ensure that all parts contact the DIW. Stack the bottom adapter first. Place the sampler cone face down on the bottom adapter. Add the top adapter. CyTOF XT User Guide...

  • Page 72: Reinstall Cones

    Place the other side of the cone removal tool (with 4 magnets) in the non-threaded holes of the sampler cone. Seat the sampler cone flush with the interface. Turn the sampler cone clockwise while applying gentle forward pressure. CyTOF XT User Guide...
  • Page 73 While pressing on the sampler cone with 2 fingers, detach the cone removal tool. Press the edges of the sampler cone to make sure it is firmly seated taking care not to touch the orifice. CyTOF XT User Guide...

  • Page 74: Reassemble The Injector And Spray Chamber

    Close the instrument front door. Reassemble the Injector and Spray Chamber Ensure that the O-ring is seated correctly in the spray chamber. NOTE If this O-ring is damaged or lost, contact Fluidigm Technical Support to order a new spray chamber. CyTOF XT User Guide...
  • Page 75 Place the spray chamber (narrow end with O-ring first) in the heater assembly. Press down gently on the spray chamber. Place the injector holder in the heater assembly cap. Ensure that THIS SIDE OUT can be seen on the injector holder. CyTOF XT User Guide...
  • Page 76 Make sure the injector is completely dry. With the injector cap off, insert the injector into the injector holder until it reaches a hard stop. IMPORTANT If the injector tip is chipped or broken, do not use. Replace the injector. CyTOF XT User Guide...
  • Page 77 Hold and turn the heater assembly so that the injector is facing the torch assembly. Align the heater assembly on the 2 guide pins and push gently to insert the injector completely into the torch assembly. CyTOF XT User Guide...
  • Page 78 Chapter 4: Maintenance Reassemble the Injector and Spray Chamber Firmly push the 2 wing nuts while turning clockwise to lock the heater assembly in place. CyTOF XT User Guide...

  • Page 79: Replace The Nebulizer Port O-Ring

    Replace the Nebulizer Port O-Ring Using the torch removal tool provided, insert the 4 pins of the torch removal tool into the holes on the nebulizer retainer for O-ring on the base of the heater assembly. CyTOF XT User Guide...
  • Page 80 Remove the damaged O-ring using a 100 µL plastic pipette tip and replace it with a new nebulizer port O-ring. Use the torch removal tool once again to secure the retainer onto the base of the heater assembly by turning clockwise. CyTOF XT User Guide...

  • Page 81: Check The Nebulizer Holder O-Ring

    Check the Nebulizer Holder O-Ring Pull on the nebulizer release collar to remove the nebulizer holder from the nebulizer mount. Firmly hold the nebulizer holder with one hand. With the other hand gently pull the nebulizer off the nebulizer holder. CyTOF XT User Guide...
  • Page 82 Unscrew the O-ring retainer to remove it from the nebulizer shell. Inspect and replace the O-ring if damaged. If damaged remove the O-ring with a 100 µL pipette tip. Reconnect the O-ring retainer to the nebulizer shell. CyTOF XT User Guide...

  • Page 83: Reassemble The Nebulizer Assembly

    Carefully tighten the wing nuts to secure the nebulizer assembly to the heater assembly. When tightening the upper wing nut, ensure that the nebulizer port safety shutter does not interfere. CyTOF XT User Guide...
  • Page 84 Ensure that the nebulizer gas line is disconnected from the nebulizer whenever the nebulizer is stored in the nebulizer rest to prevent DIW from backing up and entering the gas port. Store the nebulizer soaking in DIW when not in use. CyTOF XT User Guide...

  • Page 85: Clean The Autosampler Fluidic Lines And Nebulizer With 10% Bleach

    Using 2 sample tubes, load at least 1 or 2 mL of 10% bleach solution in Location 1 and DIW in Location 2 of the carousel. Change the autosampler settings. In CyTOF Software, on Instrument Configuration, click Autosampler. CyTOF XT User Guide...
  • Page 86 Under Sample Settings, set Sample Location to 1. Click OK to close the Flow Rate notification. Click to save the changes. To load the sample loop with bleach solution, under Actions > Workflows > Sample Prepare, click Load. CyTOF XT User Guide...
  • Page 87 Under Ion Optics > Settings > return Nebulizer Gas Flow to 0.15, press Set. NOTE It is necessary to tune the instrument before proceeding. Replace the bleach waste tube with a new nebulizer rest tube filled with DIW. CyTOF XT User Guide...

  • Page 88: Cleaning Summary

    10% bleach solution empty tube labeled Bleach Waste. (diluted to 10% v/v in DIW) 3 Run 10% bleach solution through the Autosampler fluidic lines and nebulizer. 4 Run DIW through the Autosampler fluidic lines and nebulizer. CyTOF XT User Guide...

  • Page 89: Maintenance Checklist

    The injector must be in the lowest position to avoid damage during operation. Cones ☐ Cones are clean and properly installed. The skimmer/reducer assembly is installed fully (hand tight). • The sampler cone is flush with the interface. • CyTOF XT User Guide...

  • Page 90: Periodic Maintenance

    Nebulizer holder O-ring Every 3 months Nebulizer mount Every 3 months Autosampler air filter Every 3 months Table 7. Tools required for periodic maintenance and supplied by Fluidigm Tools Provided by Fluidigm Adjustable wrench No. 2 Phillips screwdriver CyTOF XT User Guide...

  • Page 91: Autosampler Module Maintenance

    Connect the new sample loop to the LOOP positions on the switching valve of the pump and valve unit, taking care not to leave an air gap between the end of the line and the valve port. CyTOF XT User Guide...

  • Page 92: Replace The Probe Line

    Replace the Probe Line Turn the probe line counterclockwise to loosen the fitting and then remove the probe line in the PROBE position on the selector valve of the pump and valve unit. CyTOF XT User Guide...

  • Page 93: Recalibrate The Probe Line

    Recalibrate the Probe Line Place an empty tube in the carousel. Note the location number. In the CyTOF Software, on the Instrument Control tab, click Autosampler camera. The tube below the probe line in the carousel is now visible. CyTOF XT User Guide...
  • Page 94 Carousel to display the Carousel tab. Under Carousel Tray in Sample Location, enter the location number of the empty tube. Click Probe Calibration Starting Position. The probe line lowers into the empty tube until it reaches the calibration start position. CyTOF XT User Guide...
  • Page 95 Watch the Autosampler Camera to see the probe line enter the tube. An example of the probe line in a 5 mL and a 15 mL sample tube is shown below. Enter 2000 in the Steps field. CyTOF XT User Guide...
  • Page 96 Click Save New Lower Position after the correct probe line position is achieved. NOTE Use the Autosampler camera to verify the correct location of the probe line. The images below are as examples only. CyTOF XT User Guide...

  • Page 97: Replace A Bottle Line

    Feed the bottle line through the tube guides and then through the guide hole on the Autosampler top plate. Connect the new bottle line to the bottle cap. Repeat Steps 1–8 for additional bottle lines one at a time if necessary. CyTOF XT User Guide...

  • Page 98: Clean The Carousel

    Clean the Carousel NOTE Clean the carousel monthly or immediately after spillage. Tilt the probe line stir assembly from its vertical position over the carousel to its horizontal position. Lift to remove the entire carousel housing. CyTOF XT User Guide...
  • Page 99 Remove the carousel from the base by holding it at the top and pulling straight up away from the carousel tray. Wipe the carousel and the carousel tray with 10% bleach solution, followed by a thorough wipe-down with DIW. CyTOF XT User Guide...
  • Page 100 Reinstall the carousel by lining up the pin on the carousel base with the notch inside the carousel. Replace the carousel housing over the carousel. Tilt the probe line stir assembly back to its vertical position over the carousel assembly. CyTOF XT User Guide...

  • Page 101: Clean The Bottle Tray

    Remove the bottle tray and rinse with tap water. Dry with a Kimwipe. Reinstall the bottle tray. Reattach the caps and reinstall bottles. NOTE Wipe the tubing from each cap with a separate Kimwipe moistened with DIW before putting it back into the bottle. CyTOF XT User Guide...

  • Page 102: Replace Syringes

    Chapter 4: Maintenance Autosampler Module Maintenance Replace Syringes IMPORTANT If a syringe leak is observed, contact Fluidigm technical support before replacement. Replace the syringes only after leaking is observed and acquisition has been stopped. NOTE Before inserting the replacement plunger into the syringe barrel, condition the plunger tip.
  • Page 103 Using a No. 2 Phillips screwdriver, remove the screw from the right side of the syringe guard on the pump and valve unit. This allows the syringe guard to be displaced from both notches located on the left. CyTOF XT User Guide...
  • Page 104 Chapter 4: Maintenance Autosampler Module Maintenance Fold the autosampler fluidic lines forward while connected to the syringe guard to gain access to the syringe pumps. Rotate the syringe from the luer end to detach from the pump. CyTOF XT User Guide...
  • Page 105 Chapter 4: Maintenance Autosampler Module Maintenance Depress the syringe halfway towards the mount. Be mindful as some liquid may eject. While holding the syringe, unscrew the thumbscrew under the mount from the plunger and remove the leaking syringe. CyTOF XT User Guide...
  • Page 106 Insert the luer end of the new syringe into the valve and rotate it until finger tight. Pull the plunger down until it reaches the syringe drive stem. Position the plunger so that the plunger button sets into the channel of the drive stem. CyTOF XT User Guide...
  • Page 107 Autosampler Module Maintenance Hold the plunger button in the channel with both thumbs and tighten the thumbscrew into the plunger using both index fingers. Initialize the fill-pump and push-pump. Under Actions > Syringe Pumps, click Initialize. CyTOF XT User Guide...

  • Page 108: Nebulizer Assembly Maintenance

    Restart the Maxpar CAS Plus flow. No leaks should be observed. Continue with acquisition. Nebulizer Assembly Maintenance Replace the Nebulizer Line IMPORTANT Turn off Maxpar CAS Plus flow before removing the nebulizer line or the nebulizer assembly. CyTOF XT User Guide...
  • Page 109 Chapter 4: Maintenance Nebulizer Assembly Maintenance In CyTOF Software, click Instrument Configuration. Click Autosampler to open the Autosampler tab. In the Autosampler tab under the Actions > Continuous CAS Flow to Nebulizer, click Off. CyTOF XT User Guide...
  • Page 110 Turn clockwise to install the new nebulizer line to the NEB position on the pump and valve unit. Take care not to leave an air gap between the end of the line and valve port. CyTOF XT User Guide...

  • Page 111: Replace The Nebulizer Gas Line

    Pull to disconnect the quick connect nebulizer gas line from the nebulizer assembly. Remove the nebulizer gas from the CyTOF XT instrument using an adjustable wrench. Replace the line with a new nebulizer gas line and tighten it using an adjustable wrench.
  • Page 112 Remove the spring assembly from the nebulizer mount. Clean it by wiping with a Kimwipe moistened with isopropanol to remove any residue. CyTOF XT User Guide...
  • Page 113 The spring block assembly can now be easily removed from inside the nebulizer mount. Using the smaller-diameter end of the torch removal tool, align the 4 pins with the holes in the spring block assembly cap. Turn the cap counterclockwise to unthread it and access the nebulizer seal washer. CyTOF XT User Guide...
  • Page 114 IMPORTANT If the seal washer/threaded cap can be seen after the spring block assembly has been placed in the nebulizer mount, it has been inserted incorrectly. Remove and flip it over, then reinstall. CyTOF XT User Guide...
  • Page 115 Place your thumb on the flat surface of the spring assembly and turning clockwise to lock the pins in place. Refer to Reassemble the Nebulizer Assembly to prepare the nebulizer for immediate use or storage. CyTOF XT User Guide...

  • Page 116: Remove And Clean The Torch Assembly

    Allow sufficient time, at least 20 min, for these items to cool to room temperature before you handle them. Remove and Disassemble Torch Assembly Table 8. Cleaning equipment for torch assembly not provided by Fluidigm. Parts Cleaning Equipment...
  • Page 117 To avoid breakage, keep the torch assembly straight while removing and replacing. Carefully turn the white retaining ring counterclockwise to remove the ring. CAUTION Finger cut hazard. Broken glass may cause injury to fingers or hands. CyTOF XT User Guide...
  • Page 118 The torch removal tool has 2 ends, one to remove the outer torch and the other to remove the inner torch. Line up the 2 pins on the larger-diameter end of the torch removal tool with the 2 notches on the outer torch. CyTOF XT User Guide...
  • Page 119 Pivot the torch removal tool left and right to loosen the outer torch. Once the outer torch is loosened, lift the torch removal tool (which is now attached to the outer torch) from the torch holder. Then, carefully detach the outer torch from the torch removal tool. CyTOF XT User Guide...
  • Page 120 Line up the 4 pins of the smaller-diameter end of the torch removal tool with the 4 notches on the inner torch. Using the torch removal tool, turn counterclockwise to unscrew the inner torch from the torch holder. CyTOF XT User Guide...

  • Page 121: Clean The Inner And Outer Torch

    Spray isopropanol inside both inner and outer torch and allow to air-dry thoroughly. Clean the torch holder by wiping with a Kimwipe moistened with isopropanol to remove any residue. Inspect O-rings for damage or discoloration and replace if necessary. CyTOF XT User Guide...

  • Page 122: Reassemble The Torch Assembly

    Repeat Step 3a to verify that the torch verification tool now spins freely between the inner and outer torch. If the torch verification tool still does not spin freely, remove the inner torch and outer torch tools using the torch removal tool and repeat the reassembly procedure. CyTOF XT User Guide...
  • Page 123 Repeat Step 3a to check the alignment once more using the torch verification tool. Align the pin at the top of the torch assembly with the notch on the torch box. Once the pin is in the torch box, rotate it clockwise to fully seat the torch. CyTOF XT User Guide...

  • Page 124: Fill The Chiller With Coolant

    Before opening the instrument door, close CyTOF Software. If the software is not closed, the alignment may shift, and the system may not reach XY boundaries (120,000 steps). If this occurs, manually rehome the system from Instrument Configuration > Alignment. CyTOF XT User Guide...
  • Page 125 Chapter 4: Maintenance Fill the Chiller with Coolant Loosen the level sensor. Remove the level sensor. CyTOF XT User Guide...
  • Page 126 Chapter 4: Maintenance Fill the Chiller with Coolant Insert tubing into the opening of the level sensor and fill coolant to max line. Reinstall the level sensor. CyTOF XT User Guide...

  • Page 127: Check The Cooling System Air Filter

    LED light appears and the door pops open. IMPORTANT Before opening the instrument door, close CyTOF Software. If the software is not closed, the alignment may shift, and the system may not reach XY CyTOF XT User Guide...
  • Page 128 Pull on the air filter bracket to remove the air filter. If air filter is dirty, contact Fluidigm technical support to order a new one. Return the air filter to its position and tighten the thumb screws if air filter condition is good or while waiting for a replacement filter.

  • Page 129: Check The Autosampler Air Filter

    Carousel tab. Disable the Carousel Chiller. Under Carousel Chiller > Thermal Electrical Cooling, click Disable. Remove the Maxpar Water bottle from the bottle station to gain access to the magnetic autosampler air filter cover. CyTOF XT User Guide...
  • Page 130 Chapter 4: Maintenance Check the Autosampler Air Filter Pull to remove the magnetic cover. Pull the air filter bracket straight out to remove the air filter. CyTOF XT User Guide...
  • Page 131 If the air filter is dirty, replace it with a new one. Otherwise return the air filter to its original position. Place the magnetic Autosampler air filter cover back in its original position. Return the Maxpar Water bottle to the bottle station and Enable the Carousel Chiller once again in the CyTOF Software. CyTOF XT User Guide...

  • Page 132: Appendix A: Troubleshooting

    Check the argon regulator on the tank. The value should read approximately 80 psi. If a liquid argon tank is being used, ensure that the pressure build valve on the tank is open. CyTOF XT User Guide...
  • Page 133 11–13 L/min Auxiliary gas pressure 0.1–3 psi Plasma gas pressure 0.1–3 psi If the readings are outside of these ranges, contact Fluidigm technical support. Liquid in the spray Remove the nebulizer assembly, remove and disassemble the heater chamber assembly, and check the spray chamber for liquid and wipe with a dry Kimwipe.
  • Page 134 XY Calibration failed Rerun tuning, applying the settings from the most recent passed (no optimum found) tuning. Refer to the CyTOF Software v8.0 Help for CyTOF XT (FLDM-00045) 2 Manually tune XY alignment. Refer to the CyTOF Software v8.0 Help for CyTOF XT (FLDM-00045)
  • Page 135 V-pattern. Manually clean the nebulizer or replace it if necessary. Clean the cones or replace ✓ ✓ them if necessary. One or more of the ✓ ✓ subcalibrations is not being performed. Run full tuning protocol. CyTOF XT User Guide...
  • Page 136 If level of contamination is very high, the sample should not be run to prevent damage to the detector. Cell concentrations Dilute sample to the recommended concentration of no greater than too high 1 x 10 cells/mL. CyTOF XT User Guide...

  • Page 137: Appendix B: Recommendations For Extended Shutdown

    IMPORTANT Leave the argon supply on during shutdown if you plan to restart the CyTOF XT™ instrument immediately. Venting the vacuum chamber with argon results in a more rapid achievement of minimum vacuum levels needed before plasma start can be done.

  • Page 138: Cytof Xt Startup Procedure

    CyTOF XT Startup Procedure If the argon supply was on during the instrument shutdown, proceed to Step 2. Otherwise, turn on argon gas supply 2 hr prior to restarting CyTOF XT. This allows the instrument to achieve the necessary vacuum levels.

  • Page 139: Appendix C: Preventive Maintenance, Decontamination, And Disposal

    For more information, see the BMBL guidelines online at cdc.gov/biosafety/publications/index.htm. Ensure that the CyTOF XT™ system is cleaned and/or decontaminated prior to servicing the equipment, removing it from use, or transporting it for disposal. Refer to the instructions contained in this document and use only those materials specified.

  • Page 140: Dispose Of Products

    Do not dispose of this product in unsorted municipal waste. This equipment may contain hazardous substances that could affect health and the environment. Use appropriate take-back systems when disposing of materials and equipment. Learn more at fluidigm.com/compliance. CyTOF XT User Guide...

  • Page 141: Appendix D: Related Documents

    Appendix D: Related Documents Go to fluidigm.com to download these related documents. Title Document Number CyTOF XT™ System Site Requirements Guide FLDM-00360 CyTOF Software v8.0 Help for CyTOF XT FLDM-00045 CyTOF XT User Guide...

  • Page 142: Appendix E: Revision History

    GRAY, CyTOF® XT 132163 118737 Autosampler Maxpar Water Bottle Cap O-Ring Blue 2-223, CyTOF® XT 132225 118735 Autosampler Maxpar CAS Bottle Cap O-Ring Green 2-224, CyTOF® XT - 5 pack 132162 905544 Nebulizer Holder O-ring 2-105 CyTOF XT User Guide...

  • Page 143: Appendix F: Safety

    Maintain clean work areas. • Wash hands before leaving the lab. • Notify Fluidigm prior to servicing if biohazardous or radioactive materials are being used • on the instrument. Instrument Safety The system should be serviced by authorized personnel only.

  • Page 144: Symbols Used In Documentations And On The Instrument

    The following table describes the hazard symbols that may be used in this document or on labels on the system. Symbol Description Hazard. Consult the user guide for further information. Hot surface hazard. Do not touch; potential for personal injury. Biohazard. CyTOF XT User Guide...

  • Page 145: Electrical Safety

    Fluidigm personnel should install or repair the system. WARNING The system power supplies are capable of generating potentially lethal voltages and currents. Maintenance should be performed only by a Fluidigm field service engineer or by maintenance personnel, employed by the customer, who have been trained by Fluidigm and are appropriately certified.

  • Page 146: Chemical Safety

    Detailed information on exhaust vents is included in the site requirements guide. WARNING Use of the instrument or system without adequate ventilation to outside air may constitute a health hazard. Take extreme care to vent exhaust gases properly. CyTOF XT User Guide...

  • Page 147: Pressurized Gas Safety

    WARNING Inductively coupled plasma-based systems generate high levels of radio frequency (RF) energy within the RF power supply and the torch box. RF energy is potentially hazardous if allowed to escape. Do not bypass or disconnect safety devices and safety interlocks. CyTOF XT User Guide...
  • Page 148 Fluidigm Corporation South San Francisco, CA USA T: 650 266 6000 For technical support visit techsupport.fluidigm.com.

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