iota Sciences isoHub User manual

2.1.0-3
isoHub user manual

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In August of 2005, the European Union (EU) implemented the EU WEEE Directive
2002/96/EC and later the WEEE Recast Directive 2012/19/EU requiring Producers of
electronic and electrical equipment (EEE) to manage and finance the collection,
reuse, recycling and to appropriately treat WEEE that the Producer places on the EU
market after August 13, 2005. The goal of this directive is to minimize the volume of
electrical and electronic waste disposal and to encourage re-use and recycling at
the end of life.
Products bearing the symbol of a strikethrough refuse bin must not be disposed of
along with unsorted refuse.
To dispose of your isoHub at the end of its useful life, please contact iotaSciences
for the correct procedure.
NOTE: This equipment has been tested and found to comply with the limits for a Class B digital
device, pursuant to part 15 of the FCC Rules. These limits are designed to provide reasonable
protection against harmful interference in a residential installation. This equipment generates,
uses and can radiate radio frequency energy and, if not installed and used in accordance with
the instructions, may cause harmful interference to radio communications. However, there is no
guarantee that interference will not occur in a particular installation. If this equipment does
cause harmful interference to radio or television reception, which can be determined by turning
the equipment off and on, the user is encouraged to try to correct the interference by one or
more of the following measures:
—Reorient or relocate the receiving antenna.
—Increase the separation between the equipment and receiver.
—Connect the equipment into an outlet on a circuit different from that to which the
receiver is connected.
—Consult iotaSciences for further help.
FCC CAUTION: Any changes or modifications to the isoCell other than those shown in the
manual, demonstrated during training, or directed by iotaSciences authorised personnel may
void compliance and the user’s authority to operate the isoCell.

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Contents
1 Introduction.........................................................................................................................................................................................................................................5
2 Safety instructions.......................................................................................................................................................................................................................5
3 Getting started.................................................................................................................................................................................................................................6
3.1 Familiarisation......................................................................................................................................................................................................................6
3.1.1 Accessories...............................................................................................................................................................................................................8
3.2 Installation................................................................................................................................................................................................................................8
3.2.1 Remove isoHub from packaging and place on a flat stable surface ................................................8
3.2.2 Attach the condenser portion..........................................................................................................................................................9
3.2.3 Insert phase contrast slider..................................................................................................................................................................9
3.2.4 Attach objectives............................................................................................................................................................................................10
3.2.5 Attach controller..............................................................................................................................................................................................10
3.2.6 Insert power cord............................................................................................................................................................................................ 11
3.2.7 Attach 60mm dish holder or well plate holder .......................................................................................................... 11
4 Using the isoHub........................................................................................................................................................................................................................... 11
4.1 Powering-on.........................................................................................................................................................................................................................11
4.2 Moving the stage..........................................................................................................................................................................................................12
4.3 Running programs........................................................................................................................................................................................................12
4.4 Chamber states................................................................................................................................................................................................................13
4.5 Looking around a chamber...............................................................................................................................................................................14
4.6 Moving to the next chamber.........................................................................................................................................................................15
4.7 Moving to the previous chamber.............................................................................................................................................................15
4.8 Changing chamber state......................................................................................................................................................................................15
4.9 Using the touchscreen to move between chambers and change states..............................................16
4.10 Program settings............................................................................................................................................................................................................16
4.10.1 Invert...............................................................................................................................................................................................................................16
4.10.2 Speed.............................................................................................................................................................................................................................16
4.10.3 Centre............................................................................................................................................................................................................................16
4.11 Sleep mode...........................................................................................................................................................................................................................16

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4.12 Powering-off....................................................................................................................................................................................................................... 17
5 Workflow...............................................................................................................................................................................................................................................17
5.1 Identify....................................................................................................................................................................................................................................... 17
5.2 Deactivate.............................................................................................................................................................................................................................. 17
5.3 Prep Detach..........................................................................................................................................................................................................................17
5.4 Prep Extract..........................................................................................................................................................................................................................18
5.4.1 Adhesion cells .....................................................................................................................................................................................................18
5.4.2 Suspension cells................................................................................................................................................................................................18
5.5 Prep Redetach...................................................................................................................................................................................................................18
6 Other programs.............................................................................................................................................................................................................................18
6.1 Data................................................................................................................................................................................................................................................18
6.2 Options.......................................................................................................................................................................................................................................19
6.3 Set time.....................................................................................................................................................................................................................................19
6.4 Data sync.................................................................................................................................................................................................................................19
6.4.1 Setting up data syncing........................................................................................................................................................................ 20
6.5 About ...........................................................................................................................................................................................................................................21
7 Bright field/phase contrast observation........................................................................................................................................................21
7.1 Changing between bright field and phase contrast observation....................................................................21
7.2 Adjusting the aperture iris diaphragm................................................................................................................................................21
7.3 Selecting the objective.........................................................................................................................................................................................22
7.4 Changing between visual light path and camera light path..................................................................................22
7.5 Focusing on the specimen................................................................................................................................................................................22
7.6 Adjusting the brightness.....................................................................................................................................................................................23
8 Advanced observation........................................................................................................................................................................................................23
8.1 Adjusting binocular position..........................................................................................................................................................................23
8.1.1 Adjusting the interpupillary distance....................................................................................................................................23
8.1.2 Adjusting the diopter............................................................................................................................................................................... 24
8.1.3 Using the eye shades............................................................................................................................................................................... 24
8.2 Adjusting tension of coarse focusing knob.................................................................................................................................25

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9 Recording with the camera...........................................................................................................................................................................................25
9.1 Changing between the visual light path and the camera light path..........................................................25
9.2 Focusing on the specimen................................................................................................................................................................................25
9.3 Acquiring the image.................................................................................................................................................................................................25
10 Maintenance .............................................................................................................................................................................................................................26
10.1 Cleaning ...................................................................................................................................................................................................................................26
11 Troubleshooting....................................................................................................................................................................................................................26
12 Disposal and recycling ..................................................................................................................................................................................................29
12.1 Disposal of isoHub......................................................................................................................................................................................................29
12.1.1 EU Waste Electrical and Electronic Equipment (WEEE) Directive.....................................................29
12.1.2 Users outside the European Union ..........................................................................................................................................30
13 Technical support................................................................................................................................................................................................................30
14 Additional information.................................................................................................................................................................................................30
14.1 Acknowledgements..................................................................................................................................................................................................30
14.2 Limited warranty............................................................................................................................................................................................................ 31
14.3 Limited liability................................................................................................................................................................................................................. 31
14.4 Medical use restrictions.......................................................................................................................................................................................32
14.5 Registered office...........................................................................................................................................................................................................32
14.6 Legal information.........................................................................................................................................................................................................32

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1Introduction
The isoHub works with the isoCell as part of the iotaSciences’ Cloning Platform to track colonies in
a GRID through the single-cell cloning workflow. The controller and automated stage make it easy
to review chambers in a GRID and tag chambers to include in the next step of the cloning workflow,
from identifying chambers containing single-cells (monoclonal chambers), to selecting colonies
ready to extract from the GRID.
The isoHub manages all of the user dish data in the system and automatically syncs the data
wirelessly with linked isoCells when needed to run programs.
2Safety instructions
Use the isoHub with care, and take the following precautions:
•Only use the isoHub once you have read and understood this manual. Ensure this manual
is accessible for all users, and that each potential user reads and understands it.
•If the isoHub is used in a manner not specified by the manufacturer, then the protection
provided by the isoHub may be impaired.
•Powering the equipment:
oUse only with the supplied XP-POWER VEC50US24 (24VDC, 50W) power supply
and power cord.
oConnect power supply to a suitable AC mains supply with voltage in range 90 –
260V; 50 –60Hz; minimum current 1A at 115V, 0.7A at 230V.
oThe AC mains socket shall act as the disconnect device and be readily accessible.
•Avoid subjecting the isoHub to impact and vibration.
•Keep the original packaging the isoHub came in, and use this when transporting the isoHub.
•Do not connect cables in ways other than those mentioned in this manual. Different
connections may cause damage and fire.
•Do not alter the isoHub in any manner, other than those shown in the manual,
demonstrated during training, or directed by iotaSciences.
•Only use the isoHub when it is set on top of a safe, flat and stable horizontal surface.
•If water or other liquid has been spilled near the isoHub, safety may be compromised.
Ensure that no spilled liquid has been in contact with or entered the isoHub.

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•If water or other liquid enters the isoHub, stop using it immediately and unplug the power
supply. Continued use may cause a fire. Contact iotaSciences for actions or precautions.
•If any foreign object enters the isoHub, safety may be compromised. Contact iotaSciences
for actions or precautions.
•Unplug the isoHub immediately if it smokes, or produces a strange odour or unusual noise.
Continued use may cause a fire. Contact iotaSciences for actions or precautions.
•Do not autoclave the isoHub.
•The isoHub is for indoor use only.
•The isoHub is designed to work in the following operating environment (operating outside
these limits may degrade performance and damage the isoHub):
oAmbient temperature: 16 –35°C (60 –95°F)
oHumidity: 40 –60%
oAltitude: less than 2000m
oPollution degree 2
•All services shall be performed by iotaSciences authorised personnel only.
•Do not look directly into the LED light source and be careful when looking down the
eyepiece that the LED isn't set too bright.
•When attaching a camera to the isoHub, ensure the isoHub remains stable.
•Use of the isoHub with other separately available items may cause abnormal operation and
may also damage the product. Contact iotaSciences for more information.
•External influences such as electrostatic discharge may cause this equipment to
malfunction. In this case, please disconnect the power and reconnect.
3Getting started
3.1 Familiarisation
Users should familiarise themselves with the key components of the isoHub prior to use:

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Figure 1: Front view showing key components of the isoHub.
Figure 2: Back view showing key components of the isoHub.

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3.1.1 Accessories
The isoHub is supplied with the following accessories:
Documents (also available to download from the iotaSciences customer portal):
•Quickstart user guide
•Training guide
Mains power cable, with appropriate mains plug for country of use
Power supply (24V XP-POWER VEC50US24 )
Condenser portion
Phase contrast slider
Controller
60mm dish holder
Well plate holder
Dust cover
Heated centre plate (optional)
3.2 Installation
3.2.1 Remove isoHub from packaging and place on a flat stable surface
When carrying the isoHub to a different
location, remove all cables and units in
advance.
Hold the lower area [a] of the binocular portion
above the touchscreen, and the lower part of
the column [b] or the hand grip [c] at the
bottom of the column, and install the isoHub at
the predefined location. Alternatively, slide the
isoHub off the table and lift from under the base of the isoHub.

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Do not manoeuvre by holding the eyepieces, light source, stage or the focusing knobs,
as this may cause damage to the isoHub.
Do not carry the isoHub with a specimen on the stage.
3.2.2 Attach the condenser portion
If using the isoHub with the fluorescence option, then attach the umbra shield to the
condenser portion before attaching the condenser portion to the isoHub –see the
isoHub fluorescence user manual available from the iotaSciences customer portal for
more details.
Loosen the condenser fixing knob [a] to the
extent that it does not fall off.
Hold the condenser diagonally, contact the
attaching portion on the top surface [b] of the
condenser to the round dovetail portion on
the bottom of the column and fit it.
Rotate the condenser and set the condenser
fixing knob [a] at the right side position
approximately. Then, tighten the condenser fixing knob [a] to secure it.
3.2.3 Insert phase contrast slider
Set the phase contrast slider [b] with the
surface where the texts are described facing
up, and insert it in the phase contrast slider
hole [a] of the condenser portion.
Align the empty hole in the light path for
bright field observation, or the pre-centred
phase contrast ring for phase contrast
observation.

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3.2.4 Attach objectives
Remove the stage centre plate or the heated
centre plate - if using the heated centre plate,
then first detach the cable from the back of the
isoHub (see Figure 2).
Attach the objectives to the revolving
nosepiece. It is recommended to insert the
objectives into the revolving nosepiece so that
the magnification increases clockwise when
viewing the revolving nosepiece from above.
Attach caps to screw holes [a] of the revolving nosepiece where no objectives are attached to
avoid dust or dirt getting on the lenses.
After all objectives and caps are attached, fit the stage centre plate or the heated centre plate into
the stage.
If using the heated centre plate, then reconnect the heater cable at the back of the isoHub.
When removing the objectives, follow the procedures opposite to those of attaching.
3.2.5 Attach controller
Insert controller cable into the controller USB
port [a] at the back of the isoHub.
Insert the controller into the cradle [b] to the
right of the touch screen.

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3.2.6 Insert power cord
Insert power cable into the power supply.
Insert the cable from the power supply into the
power port [a] at the rear of the isoHub.
Connect the power supply's plug to the power
outlet.
3.2.7 Attach 60mm dish holder or well plate holder
Press points [a] and [b] together.
Slide the 60mm dish holder [c] or the well plate
holder into position and release points [a] and
[b].
Ensure the 60mm dish holder or well plate
holder are flat on the stage.
4Using the isoHub
4.1 Powering-on
To power-on the isoHub, ensure that it is
connected to a suitable mains power socket by
the supplied mains power cable and 24V
power supply (see Section 3.2.6). Push the
power switch to the on position (located at the
back of the isoHub on the left-hand side).
After a few seconds, the isoHub touchscreen
will display the home screen (see Figure 3).

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Figure 3: isoHub home screen
4.2 Moving the stage
While the isoHub is showing the home screen (Figure 3) or a sub-menu screen, the controller can
be used to move the 60mm dish holder or well-plate holder around the stage in order to view
different parts of the dish or well plate.
The buttons on the controller can be used to change the speed that the stage will move around
at (the upper button will increase the speed while the lower button decreases the speed).
4.3 Running programs
The programs on the isoHub can be used to scan through the chambers in a GRID made with an
isoCell, and change the state of individual chambers. For example, the Identify program can be run
to review the chambers in a recently plated GRID, and mark chambers containing single cells
(monoclonal chambers).
Ensure the 60mm dish holder is attached to the isoHub before starting a program.
Running a program works as follows:
1. Select user . The user should select their own username. If the user is using the isoHub for
the first time, they will have to enter a new username at this point (only available in the
identify program).
2. Select colour . The user needs to select the colour of the locating ring attached to their
dish. Each colour is assigned a number of dots, which corresponds to the number of
notches on the locating ring on the dish.
3. Place dish in 60mm dish holder . Ensure that the triangular protrusion of the locating ring
fits into the triangular notch of the 60mm dish holder. Press the button to continue.

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4. Select chambers . A screen similar to the Figure 4 will appear on the touch screen, and the
dish holder will move so that the first available chamber of the GRID is in view.
In each program, chambers that are available to be viewed and edited will depend on the program
being run. See the section 0for more details on each program.
Figure 4: Identify screen showing a blank dish (user IOTA’s PINK dish) before identifying monoclonal chambers.
The chamber that is currently being viewed is indicated on the touchscreen by a blue circle - in
the above example, the isoHub is viewing chamber A01. A larger icon to the left of the GRID
shows the state of the chamber that is currently being viewed.
All programs on the isoHub require the user to select a username and dish colour. The
isoHub supports up to thirty-six users. Under each username the isoHub can
simultaneously track up to nine individual dishes via the dishes’ coloured locating rings.
Stored user data can be managed via the Data program.
4.4 Chamber states
As a dish progresses through the workflow, the Cloning Platform tracks the state of the chambers.
The current chamber states for a dish can be viewed at any time via the Data program.
The chamber states are:

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Non-clonal –Chamber contains either no cells or multiple cells. This is the default
state –a chamber is assumed to be non-clonal until it is marked as clonal unfilled
via the Identify program.
Clonal unfilled –Chamber contains a single cell (or its derivative monoclonal
colony). Chamber volume is 200nl, indicating that the Fill program on the isoCell
has not yet been run on the dish.
Clonal filled –Chamber contains a single cell (or its derivative monoclonal colony).
Chamber volume is 600nl, indicating that the Fill program on the isoCell has been
run on the dish.
Detached –Dissociation reagent has been administered to the chamber using the
Detach program on the isoCell.
Harvested –The colony has been harvested from this chamber using the Extract
program on the isoCell.
(Most recently) Harvested –The harvested chambers produced by the most recent
run of the Extract program on the isoCell are distinguished by a small circle in their
corner. These chambers are the only ones that can be selected in the Prep
Redetach program.
Inactive –The cell or colony in this chamber is no longer active in the single-cell
cloning workflow. Chambers can be marked as Inactive in the Deactivate program.
Unknown –The chamber is in an unknown state. This can happen if, for example,
a program is cancelled while operating on the chamber. The Recover program on
the isoCell can be used to attempt to return these chambers to a clonal filled state.
4.5 Looking around a chamber
Whilst in a program, the joystick on the controller can be used to move to defined points in the
chamber to allow viewing the extremities of the chamber. When the joystick is released, the dish
holder will move back so that the chamber is centred in the field of view again.
The chamber should be centred in the field of view. If this is not the case, then you will
need to run through centre calibration (see section 4.10.3).

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Alternatively, the controller can be used to move freely around the chamber by pressing the
button in the top right corner of the program screen. When in this mode, the joystick will move
the dish holder around so that different areas of the chamber can be viewed, and the buttons will
change the speed. Releasing the joystick will not cause the dish holder to move back to the centre
of the dish. To exit this mode, press anywhere on the touch screen.
The direction the specimen moves in response to input from the joystick can be
changed by going to the program settings and changing the invert options (see section
4.10.1).
4.6 Moving to the next chamber
To move to the next available chamber without changing the state of that chamber, press the
lower button on the controller. The chambers that are available to be viewed depends on the
program that is being run on the isoHub –see the individual sections on each program in section
0for more details.
For continuous scanning of chambers, press and hold the lower button on the controller.
How long the isoHub will remain on each chamber when continuously scanning can be
changed by going to the program options, and adjusting the speed slider (see section
4.10.2).
4.7 Moving to the previous chamber
To move to the previous chamber, move the joystick over to the left and press the lower button
on the controller simultaneously.
To move back more than one chamber, press and hold the lower button on the controller whilst
keeping the joystick over to the left.
4.8 Changing chamber state
The upper button on the controller can be used to change the state of the chamber that is currently
being viewed.
The state that the chamber will be changed to depends on the program being run - see the
individual sections on each program in section 0for more details.
When changing the state of a chamber, the LED illumination will change colour and the dish holder
will then move so that the next available chamber is visible.

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4.9 Using the touchscreen to move between chambers and change states
Instead of using the controller to scan through chambers and change the state of chambers, the
touchscreen can be used to jump directly to a chamber and to change the state of chambers.
On the screen in Figure 4, click on the quadrant containing the chamber to move to. This will display
a zoomed in view of that quadrant. Then click on the target chamber, and the dish holder will move
so that chamber is currently being viewed. Clicking on the same chamber again will change the
state of the chamber (the state change depends on the program being run).
4.10 Program settings
Some settings are available during a program, and are available by clicking the button in the
top right corner of the program screen (Figure 4).
4.10.1 Invert
This option will change the direction that the dish moves in response to input from the joystick.
4.10.2 Speed
This option will set how fast the isoHub scans through chambers when holding down the button
on the joystick to jump between chambers.
4.10.3 Centre
If the chamber isn’t centred in the field of view when looking through the eyepieces or via a
camera, then the isoHub needs to be calibrated.
To do this, go to the program settings and follow the on-screen instructions.
4.11 Sleep mode
The isoHub will go to sleep after a certain amount of inactivity while on a menu screen. When this
happens, the touch screen will dim and the LED illumination will turn off. Whilst in sleep mode, the
isoHub will still respond to requests to sync data with any isoCells that have been setup to do this
with the isoHub.
The isoHub can also be put to sleep by pressing the button in the bottom-left corner of a menu
screen.
To wake the isoHub up, touch anywhere on the screen or press any button on the controller.
The amount of inactivity before the isoHub goes to sleep can be changed by going to More →
Settings → Options (see section 6.2). Setting the time to 0 means the isoHub won’t go to sleep
after a period of inactivity.

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4.12 Powering-off
To turn the isoHub off, push the power switch to the off position (located at the back of the isoHub
on the left-hand side).
5Workflow
5.1 Identify
Once cells have been plated into chambers on the isoCell, the chambers that contain single cells
(monoclonal chambers) need to be identified.
This is done on the isoHub in the Identify program. The isoHub will move to non-clonal chambers,
and users can mark chambers containing single cells as unfilled clonal chambers.
To distinguish between chambers that are non-clonal or unfilled clonal, the LED illumination will
be white when viewing non-clonal chambers and green when viewing unfilled clonal chambers.
5.2 Deactivate
During the Culture stage of the workflow (after clonal chambers have been filled on the isoCell),
the isoHub can be used to view all clonal filled chambers by running the Deactivate program. Any
chambers that are no longer of interest can be marked as inactive, and will no longer be included
in the Cloning Platform workflow.
The isoHub will only go to filled clonal chambers to allow users to mark them as inactive, and
chambers that have previously been marked as inactive so users can reactivate them and put them
back into a filled clonal state as necessary.
To distinguish between chambers that are filled clonal or inactive, the LED illumination will be
white when viewing filled clonal chambers and blue when viewing inactive chambers.
5.3 Prep Detach
The Prep Detach program ( Prep Harvest → Prep Detach ) is used to select colonies in filled clonal
chambers that are ready to have dissociation reagent applied to them during the Detach program
on the isoCell.
The LED illumination will be white when reviewing the chambers and turn green for chambers that
have been selected.
The Prep Detach program should not be run on a dish containing suspension cells –this
would result in the cells being removed during the PBS washes when running the
Detach program on the isoCell.

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5.4 Prep Extract
The Prep Extract program ( Prep Harvest → Prep Extract ) is used to select chambers to be removed
from the dish during the Extract program on the isoCell.
It can be run following a run of the Detach program on the isoCell for adherent cells, or alone for
suspension cells.
5.4.1 Adhesion cells
If the dish contains chambers that have had dissociation reagent added to them, then the isoHub
will move to all detached chambers during the Prep Extract program. Detached chambers will have
been pre-selected to be extracted, but the user unselect chambers with colonies that aren’t ready
to be extracted. These colonies will then need to be reselected when they are ready to extract.
The LED illumination will be white when viewing chambers that have been marked as being ready
for extraction, and blue for chambers that have been marked as not being ready for extraction.
5.4.2 Suspension cells
If the dish doesn't contain any chambers that have had dissociation reagent added to them, then
the isoHub will move to all filled clonal chambers during the Prep Extract program, and users can
mark which colonies should be extracted.
The LED illumination will be white when viewing chambers not marked as being ready for
extraction, and green for chambers that have been marked as being ready for extraction.
5.5 Prep Redetach
The Prep Redetach ( Prep Harvest → Prep Redetach ) program is used in the case where the Extract
program on the isoCell has failed to remove one or more colonies from the dish.
The user must select harvested chambers for re-detaching - the user can only select from harvested
chambers produced by the most recent run of the Extract program.
The LED illumination will be white when viewing chambers not marked for re-detaching, and green
for chambers that have been marked for re-detaching.
6Other programs
6.1 Data
The data program ( more → data ) enables the user to:
•Add or remove usernames.
•Erase individual dishes under a chosen username.

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•View current chamber states (section 4.4) of dishes under a chosen username.
Removing a username will erase all stored dish data for that username.
6.2 Options
The options program ( more → settings → options ) allows the user to:
•Adjust display screen brightness.
•Adjust volume.
•Invert the direction the dish/well plate holder moves in when using the controller.
•Turn bed heater on or off.
•Set the time before the isoHub goes to sleep (see section 4.11).
6.3 Set time
The set time program ( more → settings → set time ) is used to update the date and time on the
isoHub.
6.4 Data sync
The data sync program ( more → settings → data sync ) is for viewing the isoCells that the isoHub
has been set up to sync data with as part of the iotaSciences’ Cloning Platform. It also allows
disabling data syncing with individual isoCells.
Figure 5: Data sync screen when data syncing with an isoCell is setup.