heating system instructions - ibidi · 2019-03-14 · 4.operate the heating system in a temperature...

Version 25.04.2017

Heating System Instructions

Instructions

2 Heating System Version (26th April 2017)

Instructions

Contents

Safety Considerations 4

1 Principle – Physiological Conditions in Live Cell Imaging 15

2 Equipment 16

2.1 Components of the Heating System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16

2.2 Combination Options . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17

2.3 Temperature Controller . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18

2.4 Heated Lid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20

2.5 Heated Plate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20

2.6 Heating Inserts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21

2.6.1 Heating Insert for µ–Slides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21

2.6.2 Heating Insert for µ–Dishes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22

3 Operating the Heating System 23

3.1 Installation and Connection of the Parts . . . . . . . . . . . . . . . . . . . . . . . . . . . 23

3.2 Sample Preparation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23

3.3 Start the Heating System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24

3.4 Setting Temperatures in the Control Panel . . . . . . . . . . . . . . . . . . . . . . . . . . 24

3.5 Temperature Adjustment in the Sample . . . . . . . . . . . . . . . . . . . . . . . . . . . 25

4 Maintenance 27

4.1 Sterilization and Cleaning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 27

4.2 Influence of Surrounding Temperature and Airflow . . . . . . . . . . . . . . . . . . . . 27

5 Troubleshooting 28

5.1 Focus not Stable . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28

5.2 Evaporation is Too High . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28

5.2.1 Silicone Oil Procedure (Anti-Evaporation Oil) . . . . . . . . . . . . . . . . . . . 28

Version (26th April 2017) Heating System 3

Instructions

Safety Considerations

To ensure operation safety, the Heating System must be operated correctly and maintained accord-ing to a regular schedule. Carefully read all safety precautions in this manual before operating theinstrument. Note that the signal words WARNING, CAUTION and NOTE have specific meaningsin this manual.

Safety Symbols

Do not proceed beyond a WARNING or CAUTION notice until you have performed the indicatedactions.

WARNING! A potentially hazardous situation which, if not avoided, could result inserious injury or even death.

CAUTION A potentially hazardous situation which, if not avoided, could resultin minor or moderate injury. It is also used to alert against damagingthe equipment or the instrument.

NOTE Additional information to help achieve optimal instrument and assayperformance.

This label is positioned on the back of the device and prompts you toread the manual before using the device.

Warning messageWarning messages in the text are displayed in a gray shaded box.


Instructions

Nomenclature

Temperature Controller

Heated LidHeated Plate

WARNING!

• Do only operate the Heating Devices with the supplied Temperature Controller.

• Do only use the delivered cables and plugs. If not doing so you risk electric shock and fire.

• Do not operate the Heating System with substances or under conditions, that can cause a riskof explosion, implosion or the release of gases.

CAUTION

• Ensure that the external power supply is well accessible. The Heating System must be installedin such manner that none of its components hinders access to the external power supply.

• The weight of the Temperature Controller is approx. 3 kg. Moving the Temperature Controllerduring operation will pose a risk of personal injury or damage to the instrument.

• Some accessible parts of the Heated Plate and Heated Lid can reach temperatures up to 55°C.Avoid touching the temperature-controlled parts of the system when you have set the Temper-ature Controller to high temperatures.

• The glass plate of the Heated Lid can break due to a mechanical impact. If so, the shards canlead to injuries if handled.

• Be aware that when the system is switched on, 10 V AC voltage is applied to the underside ofthe glass on the Heated Lid and bottom of the Heated Plate. Do not touch the underside or putit in contact with anything conductive. This could cause a short circuit that may destroy theTemperature Controller and/or the Heating Devices.

• Only use the Heating System in dry rooms.

• Do not use the Heating System in cold rooms.

• Use the Heating System only for cell experiments with aqueous solutions.

• Do not use the Heating System with hazardous substances or substances/materials that pose arisk of infections.

• Do not use extension cords. Immediately replace damaged cords, plugs, or cables to avoid arisk of personal injury or damage to the instrument.

• Only ibidi technical staff is permitted to open and service the Heating Devices and/or Temper-ature Controller.

• Unplug the external power supply when the Heating System is not in use.


Instructions

Regulatory Statement

The following safety and electromagnetic standards were con-sidered:

• IEC 61010-1:2010 + Cor. :2011 Safety requirements forelectrical equipment for measurement, control and labo-ratory use - Part 1: General requirements

• IEC 61326-1:2006 EMC, Electrical equipment for measure-ment, control and laboratory use. EMC requirements.General requirements

• IEC/CISPR 11:2009, modified + A1:2010, Industrial, sci-entific and medical equipment - Radio-frequency distur-bance characteristics - Limits and methods of measure-ment


Instructions

Specifications

External Power Supply:

Heated Lid: AC 10 V, 50-60 Hz

Heated Plate: AC 24 V, 50-60 Hz

Temperature Controller: AC 230 V/50 Hz or 115 V/60 Hz

Operating Conditions

Environmental operating temperature: 18-30°C (indoor use only)

Storage temperature: -5-50°C, humidity up to 80% RH

Humidity: 80% RH up to 30°C

Operating altitude: max. 2000 m (atmospheric pressure 800-1060 hPa)

Dimensions of Heated Plate with Lid

Width: 125.5 mm

Depth: 85.5 mm

Height: 25.5 mm

Weight: 210 g

Temperature Control Range

Heated Lid (Channel 1): Ambient temperature (min. 18°C) to +46°C

Heated Plate (Channel 2): Ambient temperature (min. 18°C) to +45°C

Recommended Temperatures before Adjustment

(see Section3.5)

Heated Lid (Channel 1): 42°C

Heated Plate (Channel 2): 37°C


Instructions

Compatibility

The parts of the Heating System must only be used according to the instructions. The compatibilityof all parts is shown in detail in section 2.2.

Notices

1. ibidi shall not be held liable, either directly or indirectly, for any consequential damage incurredas a result of product use.

2. Prohibitions on the use of ibidi software

• Copying software for other than backup purposes

• Transfering or licensing of the right to use software to a third party

• Disclosure of confidential information regarding software

• Modification of software

• Use of software on multiple workstations, network terminals, or by other methods

3. The contents of this manual are subject to change without notice for product improvement.

4. This manual is considered complete and accurate at publication.

5. This manual does not guarantee the validity of any patent rights or other rights.

6. If an ibidi software program has failed causing an error or improper operation, this may becaused by a conflict from another program operating on the notebook (PC). In this case, takecorrective action by uninstalling the conflicting product(s).

7. ibidi is a registered trademark of ibidi GmbH in Germany and other countries.


Instructions

Limited Warranty

Products sold by ibidi, unless otherwise specified, are warranted for a period of one year from thedate of shipment to be free of defects in materials and workmanship. If any defects in the product arefound during this warranty period, ibidi will repair or replace the defective part(s) or product free ofcharge.

This warranty does not apply to defects resulting from the following:

1. Improper or inadequate installation.

2. Improper or inadequate operation, maintenance, adjustment or calibration.

3. Unauthorized modification or misuse.

4. Use of unauthorized tubing or fluidic connectors.

5. Use of consumables, disposables and parts not supplied by an authorized ibidi distribu-tor.

6. Corrosion due to the use of improper solvents, samples, or due to surrounding gases.

7. Accidents beyond ibidi’s control, including natural disasters.

This warranty does not cover consumables, such as cell culture chambers and dishes, tubes, fluidicconnectors, reagents etc.

The warranty for all parts supplied and repairs provided under this warranty expires on the warrantyexpiration date of the original product. For inquiries concerning repair service, contact ibidi andprovide the model name and serial number of your ibidi System.


Instructions

Installation Requirements

To ensure operation safety and optimal performance, meet the following conditions:

1. Only operate the ibidi Heating Devices with an ibidi Temperature Controller and with the sup-plied power cable.

2. Ensure that the power plug of the power supply is well accessible. The Heating Devices mustbe installed in a manner that they do not hinder the access to the external power supply and itspower plug.

3. Only operate the Heated Plate with original ibidi inserts.

4. Operate the Heating System in a temperature range of 18-30°C.

5. Operate the Heating System in a humidity range of up to 80% RH.

6. Operate the Heating System in an atmospheric pressure range of 800-1060 hPa.

7. Do not operate the Heating System under conditions that pose a risk of explosion, implosion,or the release of gases.

8. Avoid strong magnetic fields and sources of high frequency. The Heating System might notfunction properly when located near a strong magnetic field or high frequency source.

9. Avoid vibrations from vacuum pumps, centrifuges, electric motors, processing equipment, andmachine tools.

10. Avoid dust and corrosive gas. Do not install the Heating System where it could be exposed tohigh levels of dust or to outside air or ventilation outlets.

11. Do not install the Heating System in a location where it could be exposed to direct sunlight.

12. Install the Heating System in a horizontal and stable position, which includes a table, bench ordesk upon which the instrument is installed.

13. Install the Heating System in a location that enables easy access for maintenance.

14. Do not place heavy objects on the instrument.


Instructions

Installing and Connecting Cables

Confirm that the power switch of the Temperature Controller is off (power switch is at the right, rearside of the Temperature Controller).

Connect the Heating Devices to the connectors at the rear of the Temperature Controller. Connecteach device to the connector labeled with the corresponding device name.

WARNING! To avoid risk electric shock and fire, operate the Heating System with the suppliedexternal power supply and the supplied cables and plugs.

CAUTION Ensure that the power supply is easily accessible. The Heating System must be installedin a manner that does not hinder access to the power supply.

Connect the Temperature Controller to a Notebook by using the supplied network cable (only neces-sary when using a PC to control the Heating System).

Switch on the Temperature Controller and the Notebook.


Instructions

Maintenance and Operation

Keep the operating environment according to the installation requirements to maintain the declaredspecifications of the Heating System.

Operating the Heating System

Use the Heating System only as described in this manual.

CAUTION Unintended use which is not in compliance with this manual, can lead to a failure of thedevices’ protective systems. This failure may cause personal injury or damage.

Cleaning the Heating System

Switch off the Temperature Controller and disconnect the Heating Devices from the controller. Toclean the Heating Devices and/or the Temperature Controller, use only dry cloth or a cloth dampenedwith water (ultrapure) or common lab disinfection solutions that are based on quaternary ammoniumcompounds (e.g., Barrycidal 36 or Pharmacidal).

CAUTION The use of ethanol or other types of organic solvents may remove the instrument’s paint.

Transporting the Heating System

First, switch off the Temperature Controller and then disconnect the Heating Devices from the con-troller. Carry the devices carefully and avoid mechanical shocks.

Repairing the Heating System

Do not try to repair the Stage Top Incubator or the Temperature Controller by yourself. Contact theibidi service to repair any damaged components.

CAUTION The manual disassembling of the Heating System is not allowed. Manual disassemblingposes a risk of personal injury or damage to the devices. Contact ibidi service personnel if there isneed to disassemble the devices.


Instructions

Waste Treatment – WEEE/RoHS Compliance Statement

Waste treatment lies in the responsibility of the user. Contact your institution’s waste managementdepartment or a company specializing in waste recovery. Do not dispose the Heating System in atrash bin or at a public waste disposable site. For detailed information please contact the ibidi servicedepartment.

EU Directives WEEE and RoHS

To Our Valued Customers: ibidi is committed to being a good corporate citizen. As part of thatcommitment, we strive to maintain an environmentally conscious manufacturing operation. TheEuropean Union (EU) has enacted two directives, the first on product recycling (Waste Electrical andElectronic Equipment, WEEE) and the second on limiting the use of certain substances (Restrictionon the use of Hazardous Substances, RoHS).

Two Categories of products covered by the WEEE Directive are currently exempt from the RoHSDirective – Category 8, medical devices (with the exception of implanted or infected products) andCategory 9, monitoring and control instruments.

All of our products fall into either Category 8 or 9, and are currently exempt from the RoHS Direc-tive. We will, however, continue to monitor the application of the RoHS Directive in relation to ourproducts and will comply with any changes, as they apply.

Recycling is offered for our products that fall within the scope of the WEEE Directive. These specificproducts, available for sale after August 13, 2005, will be identified with a ”wheelie bin” symbol.

When you see this symbol, do not dispose of this product with Municipal Waste. Special Collec-tion/Disposal is required. Please contact us for feedback or further information.

ibidi GmbHAm Klopferspitz 1982152 [email protected]: +49 89 520 4617 0Fax: +49 89 520 4617 59


Preface

This manual is your guide to using the Heating System for cell culture experiments on an opticalmicroscope. It instructs first-time users how to use the instrument, and serves as a reference forexperienced users.

Before using the Heating System, please read this instruction manual carefully, and make sure thatthe contents are fully understood. This manual should be easily accessible to the operator at all timesduring instrument operation. When not using the Heating System, keep this manual in a safe place.If this manual gets lost, order a replacement from the company website.

Instructions

1 Principle – Physiological Conditions in Live Cell Imaging

The Heated Lid and Heated Plate are designed to keep cells on–stage at 37°C in microscopy slidesand dishes (Figure 1). The Heated Lid prevents condensation effects inside the entire incubationsystem (Figure 2).

The Heated Plate provides outer dimensions of 127.5 mm × 85.5 mm which fits perfectly into anuniversal mounting frame K. The glass top of the Heated Lid and the Heated Plate are actively heat–controlled by the Temperature Controller.

The modular concept of the Heating Inserts enables the use of different consumables ranging from60 mm and 35 mm dishes to regular slides.

Combined with a Gas Mixer. humidity, CO2–enriched, and/or O2–reduced air can be flushed in,which makes the incubation chamber a fully controlled incubator – right on the microscope stage.

Figure 1: Schematic view of the Heating System.

Figure 2: The Heated Lid prevents condensation effects on the lid of the cell culture vessel.


Instructions

2 Equipment

The Heating System is a stage top incubator to keep samples on the microscope stage at the righttemperature. It consists of the Temperature Controller, Heated Lid, Heated Plate, and Inserts to holdthe samples.

The Heated Plate fits into a universal mounting frame for multiwell plates (outer dimensions 127.5 mm× 85.5 mm). The complete height of the assembled Heated Lid and Heated Plate is 25.5 mm.

This section provides a brief overview including the nomenclature for later descriptions in this doc-ument.

2.1 Components of the Heating System

The components of the Heating System are listed with a description and the short name that is usedin this instruction manual.

Description Short name Drawing

Temperature Controller TemperatureController

Heated Lid with electrical cableand D-sub connector to connect tothe Temperature Controller

Heated Lid

Heated Plate with electrical cableand D-sub connector to connect tothe Temperature Controller

Heated Plate

Heating Insert for µ–Dish 35mm, high


Instructions

Description Short name Drawing

Heating Insert for µ–Slides.

1 temperature sensor1 perforated ibidi µ–Slide 8 Well1 perforated ibidi µ–Dish 35mm, high

1 perforated ibidi µ–Dish 35mm, low

Temperature Ad-justment Set

Power cable for the TemperatureController

Power cable

USB cable to connect the Tempera-ture Controller with a computer USB cable

2.2 Combination Options

The parts of the Heating System are combined as shown in Figure 3. The Heated Lid and HeatedPlate fit on a microscope stage equipped with a universal mounting frame for multiwell plates. Theyare connected to the Temperature Controller.

The Heating System can be combined with the Gas Mixer unit, that provides CO2 and O2 (optional)control and a defined humidified atmosphere. Detailed information on the Gas mixer unit is givenin the Gas Mixer instructions.

To hold several geometries of Slides and Dishes, the Heated Plate can be equipped with differentInserts (Section 2.6): Insert for µ–Dish 35mm, high, Insert for µ–Dish 35mm, low and Insert for µ–Slides.


Instructions

Figure 3: Overview of the parts of the Heating System with options to combine

2.3 Temperature Controller

The Temperature Controller is designed to precisely operate the different heating devices. To achievethis, the controller has four control channels that control the Heated Lid, Heated Stage (Insert), andtwo optional channels. In addition, this controller can be run either as a stand–alone system orPC–controlled. The display shows the measured (’I’ = is) and set (’S’ = set) temperatures of all thechannels. You can navigate through the menu by using the control buttons on the right. The LEDson the front indicate the status and feedback of the channels, connection to the power supply, andUSB connection (Figure 4).

Control LEDs

LED off Channel inactiveLED on/blinking slowly Channel activeLED blinking fast Channel error


Instructions

Figure 4: Front view of the Temperature Controller.

All plugs for the electrical connections are integrated into the rear of the Temperature Controller(Figure 5). The setup of the connections is shown in Section 11.

The available connections are:

• Power supply

• USB to connect to the computer

• 4 control channels

• External thermosensor (Type KTY-81)

• On/Off switch

Figure 5: Rear view of the Temperature Controller.


Instructions

Supply Voltage Adjustments

The temperature controller must be adjusted to the corresponding voltage (230 V or 115 V). Theactual voltage setting may be read and adjusted at the integrated fuse box of the power cableconnector, located on the rear panel. In order to change these settings, it is necessary to pull outthe fuse holder from the casing. Replace the fuses to match the input voltage (1 A (T) for 230V /50Hz or 2 A (T) for 115 V / 60 Hz). Rotate the grey fuse holder, and then reinsert it so that thecorrect voltage is shown in the window. Snap the fuse box back into the housing. The ”On/Off”switch is at the rear side of the controller. After selecting ”On”, the controller will automaticallystart a self-test, and then the measured values will be shown.

2.4 Heated Lid

The Heated Lid (127.5 mm × 85.5 mm) provides excellent optical quality, and also allows for the useof all standard microscopy techniques, including differential interference contrast (DIC). Due to theheight of the lid, we recommend using condensers with a working distance of ≥ 26 mm.

The upper glass part of the Heated Lid is electrically heat controlled. The electrical cable (1.5 m) isconnected to the Temperature Controller. The Heated Lid fits exactly and securely onto the HeatedPlate.

For gas incubation, the Heated Lid is equipped with inlets for the gas flow and the humidity sensor.Detailed information on the Gas mixer unit is given in the Gas Mixer instructions.

Figure 6: Heated Lid

2.5 Heated Plate

The Heated Plate provides the base for the Heating System. The lower part of the Heated Plate fitsinto any microscope stage with a universal mounting frame for multiwell plates. The electrical cable(1.5 m) is connected to the Temperature Controller.


Instructions

Figure 7: Heated Plate, top view

2.6 Heating Inserts

The Heating Insert holds the sample and fits into the Heated Plate. The Insert is held in place bystrong neodymium magnets. Use the handles to install/remove the insert to/from the Plate of theHeating System.

The Heating Inserts have a lower part (Insert) that fit into the Heated Plate, and an upper part (holder)to press the sample down. The two–part inserts have two functions that both use the force of theintegrated magnets. The first function is to hold the Slides or Dishes firmly into position, so as toavoid displacement during microscope stage movements. The second is to create a tight contact ofthe Slides or Dishes to the metallic insert, which then maximizes the heat transfer and ensures thestable heat control of the sample. The Heated Plate passively heats the inserts.

2.6.1 Heating Insert for µ–Slides

The Heating Insert for µ–Slides fits 75 mm × 25 mm µ–Slides (e.g. all ibidi µ–Slides).

Figure 8: Heating Insert for µ–Slides


Instructions

2.6.2 Heating Insert for µ–Dishes

The Heating Insert for µ–Dishes fits dishes with a diameter of 35 mm (e.g. ibidi µ–Dish 35mm, high).Center the dish in the insert, before you put the holder on.

Figure 9: Heating Insert for µ–Dish 35mm, high

To make sure that µ–Dishes are properly placed, the holder of the insert must apply a diagonal forcein order to snap the dishes into a fixed position. This will leave an inclined gap between the holderand the insert, as shown in Figure 10, but this is intended to keep the dishes in place.

Figure 10: Schematic Drawing of µ–Dish 35mm, high in the Heating Insert. To ensure a tight fit, the lidof the Insert is tilting when closed.

Warning!

Heated Plate, Heated Lid, and Heating Inserts are positioned by strong Neodym Magnets!Please contact us for a non-magnetic system, in the case that permanent magnetic fields willdisturb your experiment!


Instructions

3 Operating the Heating System

Before starting an experiment, check that the Heating System fits on your microscope stage, and thatyour cell culture vessels are compatible to the Heating Insert. Connect all parts (Section 3.1) andperform a temperature adjustment as explained in Section 3.5.

3.1 Installation and Connection of the Parts

The components of the Heating System are connected as shown in Figure 11.

Figure 11: Installation of the components of the Heating System.

To set up the Heating System follow these steps:

1. Place the Temperature Controller next to the microscope and connect the power supply.

2. Insert the Heated Plate into the opening of the microscope stage.

3. Insert a Slide or Dish into the Heating Insert and put the assembly in the opening of the HeatedPlate

4. Place the Heated Lid onto the Heated Plate.

5. Connect the electrical cables of Heated Lid (Channel 1) and Heated Plate (Channel 2) to theplugs on the back of the Temperature Controller.

To setup a computer communication to the TempControl Software, the USB cable must be connectedbetween Temperature Controller and computer.

To adjust the sample temperature, use the temperature sensor of the Temperature Adjustment Set andplug it into the correspondent connector on the back of the Temperature Controller. The temperaturesensitive ending is placed inside the sample.

3.2 Sample Preparation

Check that the cell cultureware you intend to use fits the Insert of the Heating System.

Prepare the cells according to your protocol and place the culture vessel in the Heating Insert. TheHeating Insert can be removed from the Heating System even with the holder in place. Then put thewhole assembly in the opening of the Heated Plate and close the Heated Lid.


Instructions

3.3 Start the Heating System

The Temperature Controller is switched on by the dip–switch on the back. The system immediatelystarts heating up the heated components.

Warning!

Be aware that when the system is switched on, 10V AC voltage is applied to the underside ofthe glass plate. Do not touch the underside or contact it with anything conductive. This couldcause a short circuit that may destroy the controller and/or the lid.

If you are not sure which temperature is set (e.g. when operating the system the first time), it isrecommended to disconnect all heated devices (Heated Plate, Heated Lid, and other optional heateddevices) and then switch the system on. By doing this, you ensure that you do not start the heat-ing process with the wrong temperature settings. It is now possible to set the temperatures for theindividual channels (Section 3.4). After this, you can re–connect the heated devices.

For the first operation, before adjusting the temperature in the sample for your specific setup, thefollowing temperature settings are recommended:

Table 4: Recommended temperature settings before optimization

Device Temperature

Heated Lid (channel 1) 42°CHeated Plate (channel 2) 37°C

Note!

Let the temperature of the system equilibrate for a minimum of 30 min before you start yourexperiments.

3.4 Setting Temperatures in the Control Panel

Most control parameters can be manually set on the controller using the buttons and the display onthe front panel.

The functions of the button are:

• up/right

• down/left

• select/confirm


Instructions

The cursor position is indicated with square brackets (”[xxx]”). You can move the cursor using the”Left” and ”Right” buttons. If you want to select a parameter or a function, you need to press the”Select” button. When you select a parameter, the square brackets will change to angle brackets(”<xxx>”). Now you are able to change the value up or down. To confirm the changed value, youmust press the ”Confirm” button once more. When selecting a function, you will see the individualfunction view, which is where you can change the parameters or access further functions.

Figure 12: Temperature Controller with Control Panel.

3.5 Temperature Adjustment in the Sample

The Heating System controls the temperature of the actively heated components. The temperaturein the sample will result from the interaction of all heated parts. Therefore, the temperature at theposition of the cells must be adjusted. An adjustment is recommended before starting the first ex-periment. The temperature adjustment should be done for each chamber type, in order to controltemperatures with an absolute accuracy of less than 1°C.

Note!

The displayed temperatures are valid for the heated components only. The temperature near thecells will be different!

The temperature adjustment must be renewed from time to time (at least once a year), especially ifone of the following conditions has been changed:

• Room temperature

• Air conditioning

• Chamber type or objective lens

• Humidification and gas flow

• Use of an XL-Incubator

• Use of an Objective Heater


Instructions

Note!

Before shipment, all controllers are run through an in–house calibration with the correspond-ing Heated Lid and Heated Plate. Only run the Temperature Controller with the correspondingHeated Lid and Heated Plate to ensure the correct calibration of the sensors.

To measure and calibrate the sample temperature, use the provided Temperature Adjustment Set.The temperature of the heated components has to be adjusted in small steps. Follow this procedureto adjust the sample temperature:

1. Connect the temperature sensor (thermocouple type K) to the plug on the Temperature Con-troller’s back.

2. Fill the provided µ–Dish or µ–Slide (depending on what you want to use for your experiments)with water. Volumes should resemble the amount you need for your experiments.

3. Put the loose end of the thermocouple through the hole in the lid of the µ–Dish/µ–Slide andmake sure it dips into the water.

4. Place the µ–Dish/µ–Slide in the Insert and put the holder on.

5. Place the Insert in the Heating System, close the Heated Lid.

6. Set all parameters to create your experimental conditions, such as room temperature, air con-ditioning, airflow, illumination, microscope settings, etc. For 37°C sample temperature, werecommend to start with set temperatures of 37°C for the Heated Plate and 42°C for the HeatedLid.

7. After 30 minutes, check the temperature of the external sensor in the InCuControl software. Ifthe sample temperature is still too low, raise the set values for Heated Plate and Heated Lid for0.5-1°C and wait until the sample temperature is stable again (at least 10-15 min).

8. When the sample temperature has reached the desired value, write down set values for HeatedPlate and Heated Lid. (As well as the Objective Heater – if used)

Note!

The temperature of the Heated Lid must be set to at least some degrees warmer than the tem-perature of the Heated Plate!


Instructions

4 Maintenance

4.1 Sterilization and Cleaning

The Heated Lid, Heated Plate and the Temperature Controller do not have to be sterile, because theincubation chamber has no direct contact with the cells and the cell culture medium. If sterilizationis necessary for some reason, we recommend using isopropanol (70%) or common lab disinfectionsolutions based on quaternary ammonium compounds (e.g. Barrycidal 36 or Pharmacidal).

All parts of the Heating System can be cleaned from the outside. We recommend using ultrapurewater for cleaning. Fingerprints on the Heated Lid can be removed using isopropanol (70%) orlens/eyeglass cleaning wipes.

Note!

When cleaning the Heated Lid, be careful when wiping the inner surface of the lid. The electro-conductive coating may be damaged. Also take care not to damage the glass plate of the HeatedLid.

4.2 Influence of Surrounding Temperature and Airflow

The surrounding, ambient temperature affects the temperature inside the microscopy chambers. De-vices, such as computers and camera controllers, can significantly heat up small rooms. In this case,we recommend equilibrating the room temperature to the typical experimental conditions at least2-3 hours before calibrating.

The airflow can enhance the effect of temperature changes in the vicinity of the incubation chamber.In a case where the airflow (e.g., air conditioning, etc.) cannot be stopped, we recommend protectingthe microscope as much as possible from any strong airflow. Please contact us for support.

The use of an XL–Incubator and/or an objective heater minimizes those effects and helps significantlyto stabilize surrounding conditions.


Instructions

5 Troubleshooting

5.1 Focus not Stable

Focus drift is a disturbing effect, especially during time-lapse experiments. Focus stability is mainlyinfluenced by mechanical changes and temperature variations. Follow these recommendations tokeep your cells in focus:

• Switch on all components (e.g., heating, gas incubation, computer, or other equipment) at least60 minutes before starting the experiment.

• After you put the sample onto the microscope, wait 20 minutes before starting a time-lapseexperiment to achieve temperature and immersion oil equilibration1.

• Keep the room temperature as stable as possible. Air conditioning should either be workingcontinuously or switched off.

• Do not change the temperature during the experiments. Avoid door/window openings, as thiscould rapidly change the temperature.

• Eliminate all sources of mechanical vibrations. Use a damped table for your microscope.

5.2 Evaporation is Too High

Depending on the incubating conditions, small volumes might evaporate quickly, especially duringlong–term experiments. We suggest using silicone oil (e.g. Anti-Evaporation Oil, ibidi, 50051) todecrease evaporation. If you have an actively controlled humidifying device, increase the set valuefor relative humidity.

5.2.1 Silicone Oil Procedure (Anti-Evaporation Oil)

Covering the medium with sterile silicone oil prevents all evaporation effects and is compatible withcell culture. Please do not use mineral oil, as this can be harmful to your cultureware.

Equilibrate oil and medium inside the incubator overnight. This step helps avoid the formation ofair bubbles, and pre-warms the solutions to 37°C. Afterwards, fill your slide with cells and medium.Cover the medium’s surface with an appropriate amount of silicone oil. Don’t drip the oil directlyonto the surface, but let it run down the edges by pressing the pipette tip directly on the upper sideof the reservoir. For example, when using the µ–Slide VI0.4, fill each reservoir with 30 µl cell–freemedium and 30 µl of silicone oil.

1In case the experiment needs to be started immediately, either after placing the sample on the microscope or afterclosing the lid, we recommend controlling the focus for 20 minutes. In the first minutes after starting the experiment,temperature equilibration might influence the focus/z-position of the cells.


heating system instructions - ibidi · 2019-03-14 · 4.operate the heating system in a temperature...

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