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Formulatrix NT8 User Manual

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NT8
User's Guide
Original Instructions
Applies to LCP & Plate Copy Options
Third Edition, Revised June 2018
NTG-V263R115
© 2018 Formulatrix. All Rights Reserved.

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Summary of Contents for Formulatrix NT8

  • Page 1 User’s Guide Original Instructions Applies to LCP & Plate Copy Options Third Edition, Revised June 2018 NTG-V263R115 © 2018 Formulatrix. All Rights Reserved.
  • Page 2 Support, Europe Phone: +31 652 451106 flip@formulatrix.com Acknowledgements The NT8 was developed in collaboration with Vadim Cherezov of The Scripps Research Institute (TSRI) and the JCIMPT Center led by Ray Stevens and supported by the NIH Common Fund in Structural Biology.

  • Page 3: Table Of Contents

    Introduction..................................3 Equipment Safety Guidelines............................4 Mechanical Hazards................................4 Electrical Hazards................................4 Specifications.............................5 Dimensions..................................5 Weight....................................5 Computer Requirements...............................5 Electrical Requirements..............................6 Overview..............................7 Key Features..................................7 Options....................................8 Hardware Overview...........................9 The NT8 Deck..................................9 The NT8 Head.................................11 Initial Setup..............................13 Setting Up the NT8...............................13 Preparing the NT8 for a Dispense ..........................15...
  • Page 4 Sitting Drop Experiments........................42 Hanging Drop Experiments........................45 Creating Daughter Plates (Plate Copying)...................50 Mixing LCP Drops...........................53 LCP Experiments.............................56 Running Rock Maker Experiments with NT8..................60 System Maintenance..........................62 Sterilizing the Liquid Lines............................62 Lubricating the Plate Copy Plungers........................64 Lubricating the O-rings..............................67 Enabling the Programmatic Tab..........................68 Troubleshooting and FAQs........................70...
  • Page 5 Thank you for purchasing the NT8, an automated nanoliter volume liquid handler with drop dispensing capabilities. Your NT8 must be used in the manner described in this user’s guide. Any other use may result in damage to your NT8 or personal injury. Formulatrix, Inc. is not responsible in whole or in part for any damages caused by: improper use, unauthorized alterations, adjustments or modifications, failure to comply with the procedures outlined in the NT8 User’s Guide, or use of...

  • Page 6: Introduction

    NT8 User's Guide | Chapter 1: Introduction Symbols and Conventions This icon denotes a caution and advises you of precautions to take. This icon denotes a prohibited action. This icon denotes a compulsory action and advises you of actions you must take.

  • Page 7: Regulatory Compliance

    Introduction The safety recommendations in this guide are basic guidelines. If the lab where the NT8 is to be kept has additional safety guidelines they should be followed as well, along with all applicable national and international safety codes. Regulatory Compliance Formulatrix products are CE (Conformité...

  • Page 8: Equipment Safety Guidelines

    ● Turn OFF power before inserting or removing power cables. Mechanical Hazards The NT8 is a complex electromechanical device. Only persons with the proper training should service or operate the product. All facilities to the product must be disconnected before servicing, or injury may result from the automatic operation of the equipment.

  • Page 9: Dimensions

    The NT8 should be placed on a sturdy table that has dimensions of at least 120 cm (47.2”) wide by 80 cm (32.5”) long. The water carboy should be placed to either the left or right of the NT8. The waste carboy should be placed under the table.

  • Page 10: Electrical Requirements

    NT8 User's Guide | Chapter 3: Specifications ● At least 1 GB RAM ● 1 GB Hard drive space ● FTDI 2.08.24 Electrical Requirements ● 110 V - 240 V, 50 Hz - 60 Hz, 130 W typical, 200 W max ●...

  • Page 11: Key Features

    At the core of the NT8 is an eight-channel nanopipettor that reliably dispenses drops from 50 nL to 1.8 µL with CVs averaging 5%. The NT8’s pipette tips feature a very small geometry and a hydrophobic coating, ensuring precise dispensing across the full range of buffers and reagents encountered in protein crystallization.

  • Page 12: Options

    NT8 User's Guide | Chapter 4: Overview LCP Dispensing The NT8’s LCP option can accurately and quickly dispense drops as low as 30 nL. Its fully- automated calibration system ensures that the syringe tip is centered to within 50 µm of the target drop location, and it can dispense LCP and well solution drops to a 96-drop LCP plate in about 7 minutes.

  • Page 13: The Nt8 Deck

    To begin, the two major components of the NT8 are the head and deck. Both are located at the front of the NT8. The head primarily contains tools to aspirate and dispense liquids, and the deck contains plates, tip caddies, the protein source, and the wash and waste stations.
  • Page 14 384 replacement LV tips. 5, 6, 7 – Microplate Stations. On the deck of the NT8 there are designated areas for three SBS footprint microplates. Three microplates can be placed on the deck to transfer reagents from one plate to another.

  • Page 15: The Nt8 Head

    8 is the one closest to the front. ● Optional: High Volume (HV) Tip Attachments HV tips are located at the bottom-right of the NT8's head. Each HV tip attachment is equipped with a sensor that detects any existing liquid in a deep well plate. The HV tip attachment uses 200 µL pipette tips (eight at a time) to aspirate liquid from a...
  • Page 16 Note: You can adjust the lever speed and height to fit the LCP syringe by clicking the toolbar and moving the lever button on the NT8's head up and down as necessary. ● Optional: Lipidic Cubic Phase (LCP) Syringe Mount...

  • Page 17: Setting Up The Nt8

    Setting Up the NT8 Setting up the NT8 for initial use means connecting all of the cables, setting up the water and waste carboys and installing the .NET framework as well as the FTDI drivers to your computer. A diagram of the setup is shown below for reference.
  • Page 18 Repeat steps a-g for USB Serial Converter B. Your NT8 is now ready for use. If you want to, you can find the NT8’s icon in the NT8’s files and make a shortcut for your desktop by right-clicking the icon, selecting Send to and...

  • Page 19: Preparing The Nt8 For A Dispense

    Hardware Overview. This topic will explain how to station every possible component of the NT8. You may not have to set up each component for a dispense, so make sure to read the quick start guide before prepping your hardware to avoid unnecessary hardware setup.
  • Page 20 Installing the HV Tip Caddy 1. Click the button to move the NT8's head to the left and stage back. This will allow you to slide the HV tip caddy into place. 2. Slide the tip caddy into position.

  • Page 21: Adding Plate Types

    NT8 User's Guide | Chapter 6: Initial Setup Adding Plate Types When you add a plate to the Plate Type Library, you will need to do three things: 1. Create a plate definition. 2. Define well properties. 3. Define drop properties.

  • Page 22: Adding Commands To The Task List

    ● Adding the LV or HV Dispose Tip Command Prerequisites ● You are familiar with the NT8 interface (see Software Overview on page 28). Adding the HV or LV Pick Tip Command 1. Drag and drop the HV Tip Caddy or LV Tip Caddy from the Plate Type Library into the HV tip caddy station or LV tip caddy station.
  • Page 23 NT8 User's Guide | Chapter 6: Initial Setup Adding the Wash Tip Command 1. Double-click the Wash Station. 2. Double-click the Wash Tip command on the Task List and type a new value into the corresponding field, or click the up or down arrows to increase or decrease the wash time.
  • Page 24 NT8 User's Guide | Chapter 6: Initial Setup 5. Double-click the Protein Station. 6. Select Aspirate next to Action. 7. Select the location of your protein sample. This should correlate to the number button next to the Column field where you've put your protein.
  • Page 25 NT8 User's Guide | Chapter 6: Initial Setup 4. Select Aspirate next to Action. 5. Select the columns you wish to dispense to. To select multiple columns, hold the SHIFT key and click the desired columns. In the example above we selected all well columns, 1-12.

  • Page 26: Liquid Class Guidelines

    Liquid Class Guidelines Selecting the correct liquid class for your protein and well solutions results in more precise dispenses, as the NT8 has been programmed to handle liquids of different viscosities according to their specific properties. Use the information below to help you select the correct liquid class.

  • Page 27: Attaching Lcp Attachments

    NT8 User's Guide | Chapter 6: Initial Setup Attaching LCP Attachments An LCP dispensing attachment is available for the NT8 that can dispense LCP drops of 50 - 100 nL. The dispenser uses an 8 mm barrel syringe. A sensor ensures the syringe is properly mounted before the NT8 begins dispensing drops.
  • Page 28 Occasionally you might want to empty and clean the system liquid and humidifier chamber that serve your NT8. To do so, you'll need to turn the pump control settings off while you're cleaning the chambers and then turn them back on.
  • Page 29 NT8 User's Guide | Chapter 7: Configuring Humidifiers To empty the system liquid and humidifier chambers, take the following steps: 4. Click on the toolbar to start draining the chambers. 5. Clean the chambers. To fill the system liquid and humidifier chambers, take the following steps: 1.

  • Page 30: The Experiment Designer Tab

    The Experiment Designer Tab The Experiment Designer is open by default when you first launch the NT8 software. The experiment designer enables you to interact with the NT8 platform and set up sitting drop, hanging drop, LCP, seeding, serial dilution, and/or plate copy experiments.
  • Page 31 NT8 User's Guide | Chapter 8: Software Overview 2 – The Task List Panel The Task List panel is divided into two tabs: the Task List and Task List Properties. The Task List The Task List tab provides the Repeat and Repeat Gradient buttons and displays detailed task list(s) or sequences that you have added.
  • Page 32 You can search for a plate type by typing the plate name in the Available Plate Types box. 5 – Status Bar The Status Bar is located at the bottom of the NT8 window. It keeps you apprised of important operating parameters.
  • Page 33 For multiple selection, hold the SHIFT key and click the desired well column number. If a column is selected, it appears in dark blue. The NT8 will perform the action on the selected number. Liquid Class Provides four liquid class options: Normal (N), Medium (M), High (H), and Very High (V).
  • Page 34 Exits the Task Details dialog. TIP CADDY STATIONS In order for the NT8 to pick up tips from the tip caddy on the machine, you must configure the tip caddies in the software. Setting Up the Low Volume Tip Caddy 1.
  • Page 35 NT8 User's Guide | Chapter 8: Software Overview 3. Click Set Full if you want to set the entire pipette tip caddy. For individual settings, click the desired pipette tip (represented by a circle inside the box). 4. Click OK.
  • Page 36 Protein Volume. If you are using a HV protein block, select Set High Volume Protein Tray. The Protein Volume Setup dialog appears 2. Leave the box next to Auto checked. If it is unchecked, the NT8 will not automatically use the reagent liquid class as defined in the Task Details dialog box.
  • Page 37 THE WASTE STATION If you want to tell the NT8 to dispose tips, then you need to interact with the Waste Station on the interface. Prerequisites ● You have already added the Pick Tip command to the task list.
  • Page 38 (indicated by the red button on the lower right of the software screen). At this point, the NT8 will stop moving, and you cannot operate it until the hood is closed. If the door is closed, the button you will see is this:...
  • Page 39 If the liquid detection sensors are not active, you must manually specify the solution levels on the well columns before running the plate copy experiment. This step is required for the NT8 to smoothly detect and aspirate the liquid from the deep-well plate.
  • Page 40 1. Make sure you have added the Pick Tip command for the HV tips to the Task List. 2. Optional: Click the Repeat button on the Task List to tell the NT8 that you want to repeat this task for all columns.

  • Page 41: The Rock Maker Integration Tab

    NT8 User's Guide | Chapter 8: Software Overview ● Volume: Set the liquid volume to be aspirated or dispensed by typing a new value into the corresponding field, or clicking the up and down-arrows to increase and decrease the volume.
  • Page 42 NT8 User's Guide | Chapter 8: Software Overview 2 – Experiment Properties Panel Shows the detailed experiment properties as defined in Rock Maker. There are three different tabs in experiment properties: Instruction tab, Canvas tab, and Experiment Info tab. The experiment properties display after the experiment's barcode is validated.
  • Page 43 NT8 User's Guide | Chapter 8: Software Overview Canvas tab has two different sections: experiment panel and plate view. ● Experiment Panel Item Configurable? Description Experiment ● Stock Name: The stock name. ● Type: The experiment layer type as defined in Rock Maker.
  • Page 44 Change Tips √ This is where you instruct the NT8 when to wash the LV tips. Options include: ● After Each Plate: Select this option enables the NT8 to wash the LV tips after dispensing to one plate is complete.
  • Page 45 NT8 User's Guide | Chapter 8: Software Overview Item Configurable? Description √ Should Dispense This property is only available for system with Screen Plate Copy feature. You can select whether you want to dispense screen by using the Plate Copy or not.
  • Page 46 ● Run the task list you created Set Up the NT8 Hardware 1. Place the tip caddy and the MRC 2 Well Crystallization plate on the NT8. Each well should contain 50 µL of solution. 2. Place the protein tray on the protein block, and put the protein tray clamp on top of the protein tray to hold it in place.

  • Page 47: Sitting Drop Experiments

    Option 2: Manually Set Up the NT8 Software 1. Mimic the NT8’s physical setup with the software: ● Select each microplate you stationed on the NT8 from the Plate Type Library, and drag and drop it into the appropriate location in the software.
  • Page 48 ● If you are done adding commands for that plate, click Add. If you will dispense to drop locations on this plate, click Add More. 7. Instruct the NT8 to dispense the protein and well solution mixture to the drop location on the microplate.
  • Page 49 ● Transferring the hanging drops to the experiment plate. When you set up a hanging drop experiment with the NT8, you must be careful to set up the dispense in the opposite direction by clicking the dispense direction arrow. This is because you will flip the plate when you put the plate seal on the MRC 2 Well Crystallization Plate.

  • Page 50: Hanging Drop Experiments

    You will pipette your protein into the protein block after you set up the software. Option 1: Set Up the NT8 Software Using the Hanging Drop Templates A hanging drop experiment template is preinstalled with your software. You may use it if you plan to combine 100 nL protein with 100 nL well solution for a 200 nL dispense.
  • Page 51 ● Next to Z-Offset, select the height you want the pipette tips to stop at before aspirating from the well. ● Click Add. 7. Instruct the NT8 to dispense the solution to the UVP Hanging Drop Plate Seal: ● Double-click the UVP Hanging Drop Plate Seal image to open the Task Details dialog box.
  • Page 52 ● Next to Volume, enter the desired dispense volume (generally the combined volume of the well solution and the protein volume). ● Optional: Next to Mixing, select the number of mixing cycles you want the NT8 to perform before dispensing drops.
  • Page 53 NT8 User's Guide | Chapter 10: Hanging Drop Experiments 3. Carefully slide the reference plate to the right. The adhesive surface on the UVP Seal should keep it in place while you remove the reference plate. 4. Use a microplate seal roller to secure the UVP seal to the microplate surface to assure...
  • Page 54 Option 2. Set Up the NT8 Hardware ● Place the HV tip caddy and deep well block on the NT8. Place the MRC 2 Well Crystallization plate on microplate station 2. Option 1: Set Up the NT8 Software Using the Plate Copy Template A plate copy template is preinstalled with your software.

  • Page 55: Creating Daughter Plates (Plate Copying)

    Option 2: Manually Set Up the NT8 Software 1. Mimic the NT8’s physical setup with the software: ● Select each microplate you stationed on the NT8 from the Plate Type Library, and drag it to the appropriate location in the software.
  • Page 56 HV Tip Caddy, picks up a column of tips, moves to the deep well and aspirates solution, then moves to the daughter plate and dispenses solution. Finally, the NT8 head moves back to the HV Tip Caddy and disposes the used tips in the empty column, then picks up new tips from the next column and repeats the process until your daughter place is created.
  • Page 57 ● Number of mixing cycles: Sets number of mixing cycles to be performed by the NT8. ● Mixing speed: Sets the mixing speed to be performed by the NT8. 2. Specify the settings on the LCP Mixing Wizard as needed.
  • Page 58 NT8 User's Guide | Chapter 12: Mixing LCP Drops 3. Click OK to confirm. A notification appears and asks you to connect the mixer plunger to the stage. Set up the NT8 hardware: 1. Connect the mixer plunger to the stage.
  • Page 59 Mix the LCP drops After you are done setting up the NT8 software and hardware, you are now ready to mix the LCP drops. To mix the LCP drops, follow these steps: 1. In the software, click OK to confirm that you have connected the syringe to the mixer station.

  • Page 60: Mixing Lcp Drops

    ● Using your left hand, align the syringe with the syringe bracket and hold it in place. ● Using your right hand, move the joystick to align the plunger lock on the NT8 with the top of the plunger. ● Using your left hand, place the top of the plunger into the plunger lock, and gently press the barrel of the syringe into the syringe bracket until it snaps into place.

  • Page 61: Lcp Experiments

    Wipe the tip of the LCP syringe with a chem wipe. 6. The sensors on the NT8 will detect the presence of the LCP syringe. A dialog box will open asking if you want to calibrate the syringe. Click Yes.
  • Page 62 ● Next to Volume, enter the volume of solution you want to dispense. This should match the volume you entered in step 7. ● Next to Mixing, enter the number of times you want the NT8 to mix the LCP sample and well solution before dispensing.
  • Page 63 LCP sample across a column and then dispenses a column of well solution to the LCP sample drops. This process repeats for each column you specified for the LCP plate. After the NT8 has completed the tasks for each column, it disposes the tips in the waste station.
  • Page 64 You can create experiments in Rock Maker, and then dispense them with the NT8 using Rock Maker Integration feature. Prerequisites ● You have created an experiment in Rock Maker. If you don't know how, please refer to the "Experiment Design" section of the Rock Maker help file.

  • Page 65: Running Rock Maker Experiments With Nt8

    NT8 User's Guide | Chapter 14: Running Rock Maker Experiments with NT8 4. Tick all the checkboxes to confirm that you have completed all the required instructions as shown in the picture below. 5. A notification message will appear. Click Yes to continue, then follow the on-screen...

  • Page 66: Sterilizing The Liquid Lines

    Sterilizing the Liquid Lines The NT8’s liquid lines should be sterilized every three months to discourage the growth of mold, fungus, and other undesirable elements that can thrive in an aqueous environment. Before you begin, please make sure that the NT8 is powered off.
  • Page 67 ● Click the Run Task List button to run the task list you created in step 5. ● When the NT8 finishes the task list, wait for 5 seconds to let the pump rest in order to prevent overheating. ● Repeat this process two additional times for a total of three wash cycles.

  • Page 68: Lubricating The Plate Copy Plungers

    ● Click the Run Task List button to run the task list. ● When the NT8 finishes running the task list, wait for 5 seconds to let the pump rest in order to prevent overheating. ● Repeat this process two additional times for a total of three rinse cycles.
  • Page 69 3. Using your hands, gently move the head to the center of the NT8. 4. Select an appropriately sized hex key from the hex key set, and remove the four screws that attach the dark gray cover to the NT8’s head. There are two screws on the right and two on the left.
  • Page 70 HV tip caddy. Reinstalling the NT8 Head's Cover 1. Exit the software, power-off the NT8, and unplug it. 2. Manually move the head to the center of the NT8.

  • Page 71: Lubricating The O-Rings

    4. Using the hex key, screw the four screws back in to the side of the NT8 (two on the left, two on the right).

  • Page 72: Enabling The Programmatic Tab

    2. Using the paintbrush, lightly but evenly apply grease to the front and back of each of the eight O-rings on the NT8’s plate copy head. 3. When you are finished, power-on the NT8 and continue with “Running a Plate Copy Test".
  • Page 73 NT8 User's Guide | Chapter 15: System Maintenance 4. Locate the following string: and edit it so that it looks like this (without the highlight): <add key="UserInterface.ProgrammaticTab.Visible" value="true" /> 5. Save the changes. 6. Restart the software. You should now see the Programmatic tab, as shown below.

  • Page 74: Aspiration Or Dispense Problems

    Aspiration or Dispense Problems If the NT8 is not aspirating or dispensing as expected, it could be that your pinch valve is stuck. The pinch valve is not accessible by users, but you can "unstick" it by running an extended wash cycle.

  • Page 75: Nt8 Doesn't Recognize The Device

    3 columns total, for a total of 72 µL). With NT8 2.4, we introduced a HV protein block that can hold up to 13 µL per well (8 wells per column, 3 columns total, for a total of 312 µL). We also introduced a specialized protein block that fits into Microplate Station 3, which holds the equivalent of 4 HV protein blocks, or 1248 µL.
  • Page 76 The reference plate is a clear plate that has crosshairs located at each corner. The Reference Plate crosshairs are targets for you to use if you need to tune drop position. The NT8’s software then determines the remaining drop positions from these four points The HD seal jig transfer is used to attach the HD seal to a microplate.
  • Page 77 Power Cable NT8. USB Cable The USB cable connects the NT8 to a PC. The water carboy is the water reservoir for the NT8. It is used by the humidifiers and the Water Carboy wash station Waste Carboy The waste carboy is used for liquid waste.
  • Page 78 Moves the NT8 head to HV tip caddy so that you can interact with the protein block or other components on the left side of the stage. Moves the stage back into the NT8's body so that you can slide an HV tip caddy into the HV tip caddy station.

  • Page 79: Toolbar Buttons

    Note: This button displays only if you have the LCP attachment. Displays the settings and calibration operations. Note: Calibration operations should only be performed by Formulatrix support or advanced users. Displays options to open the data directory, open the config directory, or open the logs directory.

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