Cell Dynamics W8 User Manual
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Cell Dynamics W8 User Manual

Physical cytometer
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ORIGINAL INSTRUCTIONS
ENGLISH / AMERICAN

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  • Page 1 ORIGINAL INSTRUCTIONS ENGLISH / AMERICAN...
  • Page 2 CellDynamics – W8 Physical Cytometer User Manual...

  • Page 3: Table Of Contents

    The W8 Physical Cytometer ......................21 3.2. Working principle ........................22 3.3. Applications ..........................22 Schematic overview of the W8 Physical Cytometer ................23 4.1. Component list ..........................27 Installation of the W8 Physical Cytometer ..................28 5.1. Setting Up the W8 Physical Cytometer ..................28 5.2.
  • Page 4 Incorrect tubes positioning ......................52 8.5. Incorrect naming of the LIBRA folder ..................52 8.6. Air Bubbles in the W8 Chip channel .................... 52 8.7. Multiple samples in the W8 Chip channel ................... 53 8.8. Impurities in the W8 Chip channel ....................53 8.9.
  • Page 5 CellDynamics – W8 Physical Cytometer User Manual...
  • Page 6 CellDynamics. However, notwithstanding the foregoing, the owners of the W8 Instrument may make copies solely for purposes of training personnel in the use of the unit within their business or organization.
  • Page 7 ● The appropriate use of the device is specified in the W8 Physical Cytometer User Manual. ● If damage is due to the misuse of the W8 Physical Cytometer, all warranties shall be void. This includes damage to the device itself as well as any consequential damages.
  • Page 8 CellDynamics – W8 Physical Cytometer User Manual EC DECLARATION OF CONFORMITY Referring to the Directive 2006/42/CE II.A on machinery, and amending Directive ECLARATION IN THE ORIGINAL LANGUAGE CellDynamics i.s.r.l. Via Villa Clelia 62/d 40026 IMOLA (BO) - ITALIA as manufacturer of the product, declare that the machine: Physical cytometer for the analysis of mass density, size, shape of samples comprised between 50 and 500 μm:...
  • Page 9 CellDynamics – W8 Physical Cytometer User Manual...

  • Page 10: Warnings And Precautions

    QUICK START GUIDE document. 1.1. Intended use of the instrument The W8 Physical Cytometer is conceived and tested for measuring the mass density, size and weight of sphere-like samples (i.e. spheroids, organoids, organic or hybrid microparticles, etc.). Moreover, the instrument is able to discriminate such samples based on the selected mass density or diameter range by the operator.

  • Page 11: Required Expertise Of The Operator

    18 and 30 °C. DO NOT incline the centrifuge tube, or attempt the shaking of the solution by hand. The W8 Caps are not sealed, and this would lead to a spillage of the solution containing the samples. Moreover, to avoid possible contaminations, never allow the solution to be in contact with the W8 Caps.

  • Page 12: Safety Labels, Hazards And Indicators

    CellDynamics – W8 Physical Cytometer User Manual 2. Safety labels, Hazards and Indicators The following signals are present throughout this User Manual and are associated with information related to specific risks or hazards as reported below: Indicates a potentially hazardous situation, which, if not avoided, could result in serious injuries or death.

  • Page 13: Delivery And Transportation

    The packaging is provided by CellDynamics and it must be used uniquely. Inspection of the delivery. Carefully inspect the entire packaging upon receipt of the W8 Physical Cytometer. In case of signs of mishandling or damage, file a claim with the carrier immediately. If the shipment is separately insured, file a claim with the insurer.

  • Page 14: Area Of Use And Positioning

    2.3. Area of Use and Positioning The W8 Physical Cytometer is designed to be positioned onto a fixed laboratory benchtop. Its highest performance can be reached when placed onto an anti-vibration support (i.e. Atomic Force Microscopy benchtop).

  • Page 15: Electricity And Fire Hazards

    Do not use the instrument in a wet location or environment. Particularly, do not place the instrument on areas with high humidity. Maintain at least 15 cm of distance between the W8 Physical Cytometer and any wall, object or PC. This distance is also needed for proper ventilation as discussed in Section 2.8.
  • Page 16 Do not use extension cables or power strips. Do not overload the electrical outlet. The main power supply source must be visible and accessible at any time to be rapidly unplugged in case of danger. A computer, or laptop, is the only peripheral device that can be connected to the W8 Physical Cytometer.

  • Page 17: Electromagnetic Compatibility (Emc)

    Particular attention should be focused on the correct position of the small o-rings inside the Pin at the top of the W8 Caps (Section 6.4). Take special care while handling liquids.

  • Page 18: Mechanical Hazards

    Do not disconnect or adjust in any manner the fluidic network, nor evade safety measures, during operation. Always stop a procedure (Pause button), before handling hardware accessories such as centrifuge tubes, W8 analysis solution or waste, except when it is part of the procedure indicated by the software.

  • Page 19: Service

    CellDynamics – W8 Physical Cytometer User Manual Even when not highlighted on the SDS of the substances used, wear protective gloves and clothing, as well as safety glasses to prevent contact of the substance with skin and eyes. This is highly recommended for state-of-the-art research when no side effects are known.

  • Page 20: Disposal

    CellDynamics – W8 Physical Cytometer User Manual Upgrades Only use options and upgrades recommended by CellDynamics or delegate representatives. These can be related to both the instrument and the software. Information can be gathered on the website www.celldynamics.it, or contacting the CellDynamics Tech Team at support@celldynamics.it.

  • Page 21: Introduction And Applications

    The W8 Physical Cytometer introduced on the market as a validated analytic device. In fact, the validation of the method was successfully developed by analyzing several dimensions of commercial polystyrene micro- beads, and comparing the declared mass density values with the obtained values.

  • Page 22: Working Principle

    3.3. Applications The W8 Physical Cytometer is designed for measuring mass density, size and weight out of 3D cell models like small cell clusters, cancer spheroids or organoids. The resulting physical characterization constitutes a...

  • Page 23: Schematic Overview Of The W8 Physical Cytometer

    The front panel includes the following areas or components:  The W8 Chip Compartment, designed to host the W8 Chip. Graphical detail on the W8 Chip orientation can be seen in Figure 4.3. The insertion procedure, as well as information on the preservation of the W8 Chip, are described in Section 6.6.
  • Page 24 CellDynamics – W8 Physical Cytometer User Manual 4.2 A ’ IGURE NGLED GRAPHICAL REPRESENTATION AND INDICATION OF THE INSTRUMENT S AREAS 4.3 G W8 C IGURE RAPHICAL REPRESENTATION OF THE ORIENTATION OF THE HIP FOR THE INSERTION INTO THE DEDICATED OMPARTMENT OF THE INSTRUMENT The front view of the instrument in Figure 4.4 highlights the order, name and volumes of the centrifuge...
  • Page 25 Starting from the left, there are three 15 mL centrifuge tubes (Vessel A, Vessel B and the Sample Holder), and a 50 mL one, which is mainly used for the W8 Analysis Solution. However, based on the procedure, the 50 mL tube it is intended to also contain the Cleaning Solution or the Milli-Q water.
  • Page 26 CellDynamics – W8 Physical Cytometer User Manual 4.5 G IGURE RAPHICAL REPRESENTATION OF THE REAR PANEL OF THE INSTRUMENT AND MAGNIFICATION OF THE POWER AND SWITCH COMPARTMENT ADHESIVE PLATE IN BLACK IS POSITIONED ON THE BOTTOM RIGHT AREA The CE adhesive plate, indicates the model, the serial number, the year of construction and the weight of the instrument, as well as the ratings information including power (VAC), frequency (Hz) and Ampere (A) (Figure 4.6).

  • Page 27: Component List

    Power supplier  One USB 3.1 SuperSpeed cable  W8 Caps (two standard for Vessel A and Vessel B, one for the Sample Holder and one for the 50 mL centrifuge tube)  Waste bottle (including lid and connection tube) ...

  • Page 28: Installation Of The W8 Physical Cytometer

     Finally, connect the power supply plug to the main power source. 5.2. PC requirements To operate, the W8 Physical Cytometer is coupled with the LIBRA software. Here the PC requirements needed:  Processor i7-9700 Octa Core 3 GHz or equivalent, 16 GB RAM, SSD 256, USB 3.1 SS.

  • Page 29: Operative Protocols

    6.1. Starting Up the W8 Physical Cytometer To start the W8 Physical Cytometer simply verify that all the cables are correctly plugged (power source and PC connection), switch the instrument on from the switch button on the rear panel, then turn the PC on.

  • Page 30: Waste Bottle Positioning And Connection

    ASTE BOTTLE POSITION 6.4. W8 caps positioning The W8 Caps allow to cover the centrifuge tubes and connect them to the fluidic network through the metal inlet channel. Do not try to separate the W8 cap from its attached metal channel.

  • Page 31: Reservoirs: Preparation And Connection

    6.3 W8 C IGURE AP POSITIONING SEQUENCE NOTE: Each W8 Cap has a small o-ring inserted into its protrusion at the top. Always verify the correct position of the o-ring before connecting the centrifuge tube. 6.5. Reservoirs: preparation and connection...
  • Page 32 To insert the 50 mL tube proceed as follow:  Fill a new 50 mL centrifuge tube with the W8 Analysis solution. Add a minimum of 20 mL to analyze up to 10 samples, or a minimum of 24 mL to run more than 10 samples.
  • Page 33 Do not use samples outside of the 50-500 m range in diameter. NOTE: The W8 Cap for the 15 mL Sample Holder tube has a small rotating part on the top called Pin. To insert the 15 mL Sample Holder tube proceed as follow: ...

  • Page 34: W8 Chip Information And Insertion Procedure

    Slide the W8 Chip into the W8 compartment for the insertion.  Make sure that the W8 chip is vertical and it has reached the end of the path by pushing with a finger.  Close the vertical door making sure that it is properly hooked.

  • Page 35: Cleaning Solution Preparation

    HIP INSERTION PROCEDURE Remove the W8 Chip To remove the W8 Chip the procedure requires to simply unlock and open the vertical door, then pull the W8 Chip out. Ensure to keep the vertical door closed and locked (Figure 6.8).
  • Page 36 CellDynamics – W8 Physical Cytometer User Manual NOTE: The Cleaning Solution has to be stored in a tightly closed bottle, in a cool, dry, well-ventilated area away from incompatible substances. NOTE: If stored for more than one week, prepare a new Cleaning Solution.

  • Page 37: Libra: User Manual

    CellDynamics – W8 Physical Cytometer User Manual 7. LIBRA: User Manual The LIBRA software was conceived to convey the simplest and most user-friendly working experience for the operator. This makes, both the manual procedures and measurements, easy-to-perform by all users, regardless their level of experience.
  • Page 38 CellDynamics – W8 Physical Cytometer User Manual At this stage, the operator is asked to type the experiment name in the Name box. To maintain a proper order of the experiments, it is suggested to type a name that includes the following information: year;...
  • Page 39 CellDynamics – W8 Physical Cytometer User Manual 7.4). “Pictures'' contains the sample images for each measure repetition, acquired with the High-Quality camera. “Speed” contains the terminal velocity data of that specific sample for each repetition. For the “Spheroids” example, when three samples are selected with the Number of Samples slider, the data of the “Sample0”...

  • Page 40: Sorting Options

    The Size Range information is used by the algorithm to lead the sample to the most suitable measuring area of the analysis channel inside the W8 Chip. Moreover, it allows the operator to perform an initial rough selection.

  • Page 41: Operative Panel

    The next screen displays the Operative Panel (Figure 7.6), which is divided into three main areas. On the left side the two bright-field views of the W8 Chip channel. The two cameras allow an overview of the entire channel, through the lower resolution optics, LQ camera, and a higher resolution zoomed view of the channel restriction (HQ camera).

  • Page 42: Control Panel: Fill

    Focusing on the Control Panel, the first operation regards the filling of the fluidic network, which includes the centrifuge tubes and the W8 Chip, with the W8 Analysis Solution (Fill). As displayed in Figure 7.7, the Fill procedure needs to be performed before starting a new experiment.
  • Page 43 At this stage, all the fluidic network is filled with the W8 Analysis Solution, and it will be possible to proceed. NOTE: the FILL procedure is not required after the Reset Settings protocol. In this case, the fluidic network is...

  • Page 44: Control Panel: Measure

    LQ and HQ cameras. NOTE: Check the volume of the Sample Holder tube after analyzing 3 samples consecutively without any W8 Analysis Solution refill. Always maintain a volume higher than 3.5 mL, but lower than 8.0 mL, inside the...
  • Page 45 CellDynamics – W8 Physical Cytometer User Manual NOTE: If more than one sample is introduced inside the camera of analysis, the system will automatically separate them to allow a proper measurement. One sample will be analyzed, whereas the other will be transported into the Sample Holder.
  • Page 46 NOTE: For daily and proper use of the instrument it is suggested to change the Filter monthly.  Refill 1mL: This function allows the operator to add 1 mL of W8 Analysis Solution into the Sample Holder. The process lasts 10 seconds. This can be done when the volume inside the Sample Holder decreases (always maintain a volume higher than 3.5 mL).

  • Page 47: Control Panel: Clean

    The Rinse procedure removes any residue or impurity, for example due to the previous sample, by rinsing the entire fluidic network with fresh W8 Analysis Solution. A safety pop-up is displayed after clicking on the Rinse procedure. Once confirmed, the Loading information...
  • Page 48 Replace them with empty ones.  Refill the 50 mL centrifuge tube with 30 mL of W8 Analysis Solution. When these steps are completed Click Confirm to start the RINSE protocol, which has a duration of 10 minutes (wait until the Loading information disappears).
  • Page 49 Insert the two 15 mL centrifuge tubes, empty and clean, related to Vessel A and Vessel B.  Insert the 15 mL Sample Holder centrifuge tube containing the new sample dispersed into fresh W8 Analysis Solution (maximum volume 7 mL).
  • Page 50 Once this final step is completed the LIBRA software switches off automatically. 7.17 F IGURE INAL LEAN TERILIZATION The W8 Physical Cytometer is now clean, empty from liquids and it can be turned off from the main switch button (on the back of the instrument) to be ready for new measurements.

  • Page 51: Troubleshooting: A Short Guideline On Possible Issues

    8.2. Out-of-scale liquids volumes  Fill a new 50 mL centrifuge tube with the W8 Analysis solution. Add a minimum of 20 mL to analyze up to 10 samples, or a minimum of 24 mL to run more than 10 samples.

  • Page 52: Incorrect Tubes Positioning

    CellDynamics – W8 Physical Cytometer User Manual  In order to prolong the life and maintain the best efficiency of the W8 Chip, it is crucial to perform the CLEAN procedure at the end of every experiment.  Make sure that the W8 Chip is positioned vertically and it has reached the end of the insertion path by pushing with a finger.

  • Page 53: Multiple Samples In The W8 Chip Channel

    8.8. Impurities in the W8 Chip channel Cellular debris may be present in the Sample Holder tube and may enter in the W8 Chip channel during the test. Impurities are removed by clicking the Pause button and then selecting the Remove Impurities function.
  • Page 54 CellDynamics – W8 Physical Cytometer User Manual...

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