For Research Use Only. Not for use in diagnostic procedures.
Countess
™
II automated cell counter
USER GUIDE
Catalog Number AMQAX1000
Publication Number MAN0014293
Revision C.0
Life Technologies Corporation | 22025 20th Ave SE Ste. 100 | Bothell, WA 98021
For descriptions of symbols on product labels or product documents, go to thermofisher.com/symbols-definition.
The information in this guide is subject to change without notice.
DISCLAIMER: TO THE EXTENT ALLOWED BY LAW, THERMO FISHER SCIENTIFIC INC. AND/OR ITS AFFILIATE(S) WILL NOT BE LIABLE FOR SPECIAL,
INCIDENTAL, INDIRECT, PUNITIVE, MULTIPLE, OR CONSEQUENTIAL DAMAGES IN CONNECTION WITH OR ARISING FROM THIS DOCUMENT,
INCLUDING YOUR USE OF IT.
Revision history: Pub. No. MAN0014293
Revision Date Description
C.0 07 June 2019 Updated trypan blue handling instructions. Document converted to XML.
B.0 30 June 2017 Add info about edited profile indicator, save profile from results screen,
dilution calculator, and reports.
A.0 01 September 2015 New user guide
Important Licensing Information: These products may be covered by one or more Limited Use Label Licenses. By use of these products, you accept
the terms and conditions of all applicable Limited Use Label Licenses.
TRADEMARKS: All trademarks are the property of Thermo Fisher Scientific and its subsidiaries unless otherwise specified.
©2019 Thermo Fisher Scientific Inc. All rights reserved.
Contents
■
CHAPTER 1 About this guide ............................................ 7
Audience ....................................................................... 7
User documentation ............................................................. 7
Text and keyboard conventions .................................................... 7
User attention words ............................................................ 8
Safety alert words ............................................................... 8
■
CHAPTER 2 Product information ....................................... 9
Product description ............................................................. 9
Product contents ........................................................... 9
Countess
™
II automated cell counter .......................................... 9
Upon receiving the instrument ................................................ 9
Register your instrument ................................................... 10
Instrument exterior components ................................................. 10
■
CHAPTER 3 Getting started ............................................ 11
Installation .................................................................... 11
Operating environment ..................................................... 11
Install the instrument ...................................................... 11
Turn ON the instrument .................................................... 11
Load profile ................................................................... 12
Profiles screen ............................................................ 12
Edit profile screen ......................................................... 13
Load a profile ............................................................. 14
Add/edit a profile .......................................................... 14
■
CHAPTER 4 Cell count and cell viability assays ...................... 16
Prepare sample ................................................................ 16
Recommendations ......................................................... 16
Load Countess
™
chamber slide .............................................. 16
Count cells .................................................................... 17
Count procedure ........................................................... 17
Next steps ................................................................ 18
Countess
™
II automated cell counter User Guide
3
■
CHAPTER 5 Results .................................................... 19
View results ................................................................... 19
Results screen ............................................................ 19
Identify the cells counted ....................................................... 19
Advanced screen .......................................................... 19
Identify live and dead cells .................................................. 20
Graph count results ............................................................ 21
View graph ................................................................ 21
Gate count results .............................................................. 21
Adjust screen ............................................................. 21
Gate count results ......................................................... 21
Save as new protocol ........................................................... 23
Edit and save profile as new protocol ......................................... 23
■
CHAPTER 6 Dilution calculator ........................................ 25
Dilution calculator ............................................................. 25
Calculate dilution .............................................................. 25
■
CHAPTER 7 Save results ............................................... 28
Save count results ............................................................. 28
Save screen ............................................................... 28
Save procedure ............................................................ 28
Report ........................................................................ 31
Report file ................................................................ 31
■
CHAPTER 8 Instrument settings ...................................... 32
Overview ...................................................................... 32
Instrument settings screen ................................................. 32
Software update ............................................................... 32
Guidelines for software update .............................................. 32
Update the Countess
™
II/II FL software ....................................... 33
Date/Time ..................................................................... 34
Set the date and time ....................................................... 34
Contents
4
Countess
™
II automated cell counter User Guide
■
CHAPTER 9 Maintenance .............................................. 36
Instrument care ............................................................... 36
General guidelines for care ................................................. 36
Power supply .............................................................. 36
Clean the Countess
™
II automated cell counter .................................... 37
Introduction ............................................................... 37
Clean the touch-screen ..................................................... 37
Clean the instrument case .................................................. 37
Decontaminate the instrument .............................................. 37
Set nominal focus .............................................................. 38
Overview ................................................................. 38
Set nominal focus .......................................................... 38
■
APPENDIX A Troubleshooting ......................................... 40
■
APPENDIX B Product specifications .................................. 43
Technical specifications ......................................................... 43
Physical characteristics .................................................... 43
Technical specifications .................................................... 43
Optics .................................................................... 43
Analysis slide ............................................................. 43
■
APPENDIX C Ordering information .................................... 44
Countess
™
II automated cell counter ............................................. 44
Accessory products ............................................................ 44
■
APPENDIX D CSV file format ........................................... 45
CSV file format, explained ....................................................... 45
Overview ................................................................. 45
■
APPENDIX E Safety ..................................................... 47
Safety alert words .............................................................. 47
Electrical symbols ............................................................. 48
Safety symbols ................................................................ 48
Environmental symbols ......................................................... 49
Safety labels on instruments .................................................... 50
General instrument safety ...................................................... 50
Operating the instrument ................................................... 50
Safety precautions ......................................................... 51
Cleaning or decontaminating the instrument .................................. 51
Removing covers or parts of the instrument ................................... 51
Contents
Countess
™
II automated cell counter User Guide
5
Chemical safety ................................................................ 52
Chemical waste safety .......................................................... 52
Chemical waste hazard ..................................................... 52
Chemical waste safety guidelines ............................................ 53
Waste disposal ............................................................ 53
Electrical safety ................................................................ 53
Biological hazard safety ......................................................... 54
Safety and electromagnetic compatibility (EMC) standards .......................... 55
■
Documentation and support ............................................. 56
Customer and technical support ................................................. 56
Limited product warranty ....................................................... 56
Contents
6
Countess
™
II automated cell counter User Guide
About this guide
Audience
This user guide is for laboratory sta operating, maintaining, and analyzing data
using the Countess
™
II Automated Cell Counter.
User documentation
The guides listed below are available for the Countess
™
II Automated Cell Counter.
Guide
Pub. no.
Countess
™
II Automated Cell Counters User Guide
MAN0014293
Countess
™
II and Countess
™
II FL Automated Cell Counters Quick
Reference Card (QRC)
MAN0010826
Additional resources are available on the Countess
™
Technical Resources page. Go to
www.thermosher.com/countess to access protocols, application notes, and tutorials.
Text and keyboard conventions
Text and keyboard conventions Text and keyboard conventions used in this user
guide are listed below. For safety alert words and symbols used in this document, see
“Safety alert words“ on page 8.
Convention
Use
Bold Bold text indicates user action. For example: Press More.
► Right arrow symbol (►) indicates a menu choice, and separates
successive commands you execute or select from a drop-down or
shortcut menu. For example: Select More4Adjust.
1
Countess
™
II automated cell counter User Guide
7
User attention words
Two user aention words appear this document. Each word implies a particular level
of observation or action as described below.
Note: Provides information that may be of interest or help but is not critical to the use
of the product.
IMPORTANT! Provides information that is necessary for proper instrument
operation, accurate installation, or safe use of a chemical.
Safety alert words
Four safety alert words appear in this document at points where you need to be aware
of relevant hazards. Each alert word—IMPORTANT, CAUTION, WARNING,
DANGER—implies a particular level of observation or action, as dened below:
IMPORTANT! – Provides information that is necessary for proper instrument
operation, accurate installation, or safe use of a chemical.
CAUTION! – Indicates a potentially hazardous situation that, if not avoided,
may result in minor or moderate injury. It may also be used to alert against
unsafe practices.
WARNING! – Indicates a potentially hazardous situation that, if not avoided,
could result in death or serious injury.
DANGER! – Indicates an imminently hazardous situation that, if not avoided,
will result in death or serious injury. This signal word is to be limited to the
most extreme situations.
Except for IMPORTANT! safety alerts, each safety alert word in this document
appears with an open triangle gure that contains a hazard symbol. These hazard
symbols are identical to the hazard symbols that are axed to the instruments (see
Safety symbols in Appendix E).
Chapter 1 About this guide
User attention words
1
8
Countess
™
II automated cell counter User Guide
Product information
Product description
The Countess
™
II Automated Cell Counter is shipped with the components listed
below.
Component
Quantity
Countess
™
II Automated Cell Counter (Cat. no. AMQAX1000) 1 each
Power Cord with 4 adaptor cords (for U.S./Canada/Taiwan/Japan,
Europe, or UK)
1 each
Countess
™
Cell Counting Chamber Slides (50 slides/box) 1 box
Countess
™
II Disposable Slide Holder 1 each
Countess
™
II USB drive 1 each
Countess
™
II Automated Cell Counter Quick Reference Card 1 each
The Countess
™
II Automated Cell Counter is a fully automated cell counter and assay
platform that uses state-of-the-art optics and image analysis algorithms to analyze
trypan blue-stained cells in suspension.
• The Countess
™
II Automated Cell Counter oers an intuitive user interface, and
provides the option to save data and generate a report, which can then be
transferred to a PC using the USB drive supplied with the instrument or available
separately.
• The cells to be counted are loaded into the instrument in disposable Countess
™
Cell Counting Chamber Slides (“Prepare sample“ on page 16). Each chamber
slide contains two enclosed chambers to hold the sample to allow you to measure
two dierent samples or perform replicates of the same sample.
• The Countess
™
II Automated Cell Counter takes 10 seconds per sample for a
typical cell count and is compatible with a wide variety of eukaryotic cells. In
addition to cell count and viability, the instrument also provides information on
cell size.
Examine the instrument carefully for damage incurred during transit. Ensure that all
parts of the instrument, including accessories listed above, are included with the
product. Damage claims must be led with the carrier; the warranty does not cover
in-transit damage.
See “Installation“ on page 11 for instructions on installing the instrument.
2
Product contents
Countess
™
II
automated cell
counter
Upon receiving the
instrument
Countess
™
II automated cell counter User Guide
9
Visit www.thermosher.com/registercountess to register your instrument. You will
be asked to supply the serial number, your name, and your contact details.
Registering your instrument ensures that you will receive notications of software
upgrades and information on new assays for use with the Countess
™
II Automated
Cell Counter.
Instrument exterior components
①Touch-screen display: The 7-inch capacitive touch-screen display is the main user
interface of the Countess
™
II Automated Cell Counter. It contains the buons for all
instrument functions and displays data from the cell count.
②⑤ USB ports: The USB ports allow you to transfer and save the cell count data and
image to an external computer for record keeping and printing purposes. You can use
the USB drive supplied with the instrument or any other standard, FAT32-formaed
USB drive for data transfer. If desired, you can plug in a USB mouse into the rear USB
port for instrument control.
Note: The USB ports located in the front and the back of the instrument function the
same. However, the rst USB drive connected will be the preferred saving location
and both USB drives cannot be accessed at the same time.
③Slide port: The slide port is used to insert the Countess
™
Cell Counting Chamber
Slide containing the sample into the instrument.
④Power switch: The ON/OFF rocker switch is the main power switch. It is not
necessary to use the power switch for day-to-day operation of the instrument.
⑥Power input jack: The power input jack connects the instrument to an electrical
outlet through the supplied power cord and the appropriate plug, based on the
electrical outlet conguration in your country.
Note: The Countess
™
II Automated Cell Counter (Cat. no. AMQAX1000) uses
brighteld illumination only and does not support the EVOS
™
LED light cubes.
Register your
instrument
Chapter 2 Product information
Instrument exterior components
2
10
Countess
™
II automated cell counter User Guide
Getting started
Installation
• Place the instrument on a level surface away from vibrations emanating from
other pieces of equipment.
• Allow at least 5 cm (2 in) free space at the back of the instrument to allow for
proper ventilation and prevent overheating of electronic components.
• Set up the instrument away from direct light sources, such as windows. Ambient
room lighting can enter the imaging path and aect the image quality.
• Operating temperature range: 4°–32℃ (40°–90°F).
• Relative humidity range: 30–90%.
IMPORTANT! Do not position the instrument so that it is dicult to turn o the main
power switch located on the back of the instrument (see “Instrument exterior
components“ on page 10). In case of an instrument malfunction, turn the main power
switch to the OFF position and disconnect the instrument from the wall outlet.
1.
Unpack the instrument and place the instrument on a at, level, dry surface.
2.
Remove the thin plastic protector lm from the touch-screen display.
3.
Plug one end of the power cord appropriate for your region into the instrument.
4.
Plug the power cord into the electrical outlet. Be sure to use only the power cord
supplied with your instrument. Powering the instrument with an unapproved
power cord may damage the instrument.
1.
Turn on the instrument by ipping the power switch on the back of the
instrument (“Instrument exterior components“ on page 10) to the ON position.
The instrument initializes and displays the Home screen.
3
Operating
environment
Install the
instrument
Turn ON the
instrument
Countess
™
II automated cell counter User Guide
11
2.
From the Home screen, you can proceed immediately to the assays by inserting a
slide (Chapter 4, “Cell count and cell viability assays“).
Alternatively, you can change or add a prole (Step 3 on page 12) or change
instrument seings (Step 4 on page 12).
3.
To change the current prole or to add a new prole to the instrument, press
the Proles buon in the upper left corner.
Proles allow you to create customized count preferences (i.e., gate counts based
on cell size, brightness, and/or circularity) (“Load prole“ on page 12).
4.
To change instruments seings, press the Instrument Seings buon in
the
upper right corner.
Instrument seings allow you to update the Countess
™
II software and change
the date and time (Chapter 8, “Instrument seings“).
Load profile
Proles screen allows you to create and save up to 9 customized proles. Each custom
prole denes the count parameters (size, brightness, and circularity) and the Auto
Lighting function for a consistent and streamlined workow.
• You can access the Proles screen from the Home, Capture, Results, Advanced, or
Adjust screens.
• The current prole is displayed on the upper left corner of the Home, Capture,
Results, Advanced, or Adjust screens .
• The Auto Lighting function and count parameters are set in the “Edit prole
screen“ on page 13.
• The Default prole contains default count seings and cannot be edited.
• The count parameters specied in the selected prole are applied to all new cell
counts.
• If you have already performed a count, loading a new prole from the Results
screen applies the count preferences to the current counts results (total cells,
viability etc.) and to all new counts.
• If you change any seing that is saved as part of the protocol (size, brightness, or
circularity) on the Results screen, the prole name is appended with the (*)
symbol.
Profiles
screen
Chapter 3 Getting started
Load profile
3
12
Countess
™
II automated cell counter User Guide
The Auto Lighting function and count parameters are set in the Edit prole screen.
Auto lighting
Auto Lighting function automatically illuminates the sample in the brighteld
channel for increased sample-to-sample consistency and decreased user-to-user
variability. By default, the Auto Lighting function is turned on; to toggle it on and o,
use the Auto Lighting checkbox (see “Add/edit a prole“ on page 14).
Count parameters
Count parameters are set using the parameter sliders, which correspond to a single
channel that is selected via the channel selection radio buons.
• Size: As you move the slider up, the algorithm includes larger objects in the
count. As you move the slider down, only the smaller objects are counted.
= larger objects
= smaller objects
• Brightness: As you move the slider up, the algorithm includes the brighter
objects in the count. As you move the slider down, only the dimmest of objects
are counted.
= brighter objects = dimmer objects
• Circularity: As you move the slider up, the algorithm includes more objects with
shapes other than circular in the count. As you move the slider down, only the
objects that are perfect circles are counted.
= less circular = more circular
• Size and brightness sliders are range sliders.
To adjust the upper and lower boundaries without changing the data range, drag
the slider by its middle section (i.e., the slider bar).
To adjust only the upper or the lower boundary, move the upper or the lower
handle in the desired direction. This will also change the range of values within
which the cells are counted.
• The circularity slider only sets a single threshold value; cells that fall below the
set value are counted, and cells that are beyond this range are excluded.
To adjust the threshold for circularity, drag the slider in the desired direction.
Edit
profile screen
Chapter 3 Getting started
Load profile
3
Countess
™
II automated cell counter User Guide
13
1.
Press the Proles buon located on the upper left corner of the screen to open
the Proles screen.
2.
Press the desired prole to select, and then press Load.
The instrument will load the count parameters specied in the selected prole
and return to the previous screen.
3.
To return to the previous screen without loading the new prole, press
the
previous buon.
The instrument will keep the saved prole, but return to the previous screen
without loading it.
1.
Press the Proles buon located on the upper left corner of the screen to
navigate to the Proles screen.
2.
To add or edit a new prole, select an empty or an existing prole, and then
press Edit. The Edit screen for the selected prole opens.
Note: The Default prole contains default count seings and cannot be edited.
3.
Select or deselect the Auto Lighting checkbox to turn the Auto Lighting function
ON or OFF.
4.
To dene the new count parameters, select Live or Dead from the Trypan Blue
radio buons.
5.
Adjust the size, brightness, and circularity thresholds using the appropriate
parameter slider (“Count parameters“ on page 13).
Note: Although the Auto FL Threshold checkbox and the FL channel radio
buons are visible on the Edit prole screen, they are not functional on
Countess
™
II Automated Cell Counters, which only feature a single brighteld
channel.
Load a
profile
Add/edit a profile
Chapter 3 Getting started
Load profile
3
14
Countess
™
II automated cell counter User Guide
6.
To assign a name to the new prole or to change the name of the existing prole,
press the Prole name text box. The alpha-numeric keypad opens.
7.
Type in the desired prole name using the alpha-numeric keypad.
To enter symbols, press the symbol (@%&) key. To return to the alpha-numeric
keypad, press ABC.
8.
Press Enter to save the name and return to the Edit prole screen.
To return to the Edit prole screen without saving the name, press the close
buon.
9.
Press Save to save the new prole, and then press Close in the conrmation
screen to return to the Proles screen.
To return to the Proles screen without saving, press Cancel.
10.
On the Proles screen, press Load. The instrument will load the count
parameters specied in the selected prole and return to the previous screen.
11.
To return to the previous screen without loading the new prole, press the
previous
buon. The instrument will keep the saved prole, but return to the
previous screen without loading it.
Chapter 3 Getting started
Load profile
3
Countess
™
II automated cell counter User Guide
15
Cell count and cell viability assays
Prepare sample
To obtain the best results, follow these recommendations:
• Ensure that the cell sample is homogeneously mixed.
• The measurement range extends from 1 × 10
4
–1 × 10
7
cells/mL, but the optimal
range is 1 × 10
5
–4 × 10
6
cells/mL.
• For accurate results in cell viability assays, ensure that the counting area is
covered with the cell suspension and count the cells immediately after staining
per the assay protocol.
• Do not press the optical surfaces of the chamber slides. Hold the slides by the
edges.
• Take care to avoid forming bubbles in the sample.
• Sterile ltering and centrifugation can be used to remove precipitates common
within trypan blue solutions. Alternatively, avoid mixing and vortexing trypan
blue stock solutions to allow precipitates to remain at the boom of tube, thereby
promoting more accurate cell counts. Also, precipitates can be reduced by gentle
heating at 37°C for 10 minutes.
1.
Prepare the sample by adding 10 µL of your cell suspension to 10 µL of 0.4%
trypan blue stain. Mix the sample mixture well by pipeing up and down a few
times.
2.
Gently pipet 10 µL of the trypan blue-stained sample into the half moon-shaped
sample loading area. The sample is loaded into the chamber through capillary
action.
Note: Each chamber in the Countess
™
Cell Counting Chamber Slide has a 10-µL
sample capacity. Do not overll the slide chambers.
3.
Let the sample mixture sele in the chamber for 30 seconds, and then insert the
slide into the slide port (see “Instrument exterior components“ on page 10). You
will hear a soft click, if the slide is pushed in correctly.
4
Recommendations
Load Countess
™
chamber slide
16
Countess
™
II automated cell counter User Guide
4.
To remove the slide, push the slide gently into the instrument until it “clicks”
and a spring pushes the slide out. Grasp the slide and pull it out the rest of the
way.
Note: After using the Countess
™
Cell Counting Chamber Slides, appropriately
dispose of them as biohazardous waste. Do not reuse the disposable chamber
slides.
Count cells
1.
Load the Countess
™
Cell Counting Chamber Slide with your trypan blue-stained
sample and insert it into the slide port of the instrument as described in “Prepare
sample“ on page 16. You will hear a soft click, if the slide is pushed in correctly.
2.
When the slide is inserted, the instrument automatically illuminates the sample,
sets the intensity of brighteld illumination, and auto focuses on the cells.
Note: To turn o the Auto Lighting function, see “Auto lighting“ on page 13.
3.
Optional: Press the Proles buon and load the desired prole as
described in “Load a prole“ on page 14.
4.
Optional: If desired, press the Focus buon, and then use the Focus slider to
manually bring your sample into focus (see Note on page 17).
If needed, Zoom in on the image while adjusting the focus.
Note: The auto focus algorithm is designed to highlight the dierences between
live and dead cells. Therefore, the optimal focus level is where the “live” cells
have a light colored center and the “dead” cells are dark throughout. To enable
optimal auto focus functionality, you may need to initially rene the focus by
adjusting it manually and then seing it as the nominal focus. For more
information, see “Set nominal focus“ on page 38.
5.
Optional: Manually adjust exposure using the light source slider.
The light source slider controls the LED intensity, camera gain, and exposure
time and it is used for adjusting the image brightness.
Count procedure
Chapter 4 Cell count and cell viability assays
Count cells
4
Countess
™
II automated cell counter User Guide
17
6.
Press Capture. The instrument temporarily captures the image and displays the
results (total concentration, percentage and concentration of live and dead cells).
For more information, see “Chapter 5, “Results“”.
• To identify the objects (i.e., cells) counted as live or dead, press More to navigate
to the Advanced
™
screen (“Identify the cells counted“ on page 19).
• To see the distribution of live and dead cells in a graphical format, press the
Graph buon (see “Graph count results“ on page 21).
• To gate the results by object size, brightness, or circularity, rst press More to
open the Advanced
™
screen, and then press Adjust to navigate to the Adjust
screen (“Gate count results“ on page 21).
• To permanently save the results, press Save (see Chapter 7, “Save results“).
• To perform a new count, remove the slide and reinsert it into the instrument to
count the sample in the second chamber.
Next steps
Chapter 4 Cell count and cell viability assays
Count cells
4
18
Countess
™
II automated cell counter User Guide
Results
View results
The Results screen for cell count and cell viability assays displays a composite image
of the objects counted and the results of the cell count and cell viability calculations
(total concentration, percentage and concentration of live and dead cells).
Note: When performing brighteld (BF) cell counts and cell viability assays, the
counting algorithm assumes that you have diluted your cells 1:1 in trypan blue and
takes this dilution into account when calculating the total cell concentration. The cell
concentration displayed in the Results screen is the original cell concentration before
dilution into trypan blue.
Identify the cells counted
The Advanced screen allows you to identify the objects (i.e., cells) counted in each
channel and included in the count results for further review. After reviewing the
marked objects, you can adjust the threshold for size, brightness, and/or circularity as
desired for your application.
5
Results screen
Advanced screen
Countess
™
II automated cell counter User Guide
19
1.
On the Results screen, click More. The Advanced screen opens.
2.
To identify the cells that are included in the count as “"live"”, press the Live
buon. "“Live"” cells will be circled in green on the screen.
To identify the cells that are included in the count as "“dead"”, press the Dead
buon. “"Dead"” cells will be circled in red on the screen.
Note: You may select either or both options. In the example below, both Live
and Dead buons are pressed and "“live"” and “"dead"” cells are marked with
green and red circles around them, respectively.
3.
To unmark the cells identied as "“live"” (green) or "“dead"” (red) on the screen,
press the Live or the Dead buon again, respectively.
Identify live and
dead cells
Chapter 5 Results
Identify the cells counted
5
20
Countess
™
II automated cell counter User Guide
Graph count results
For cell count and cell viability assays, you can view the distribution of cells (live
and/or dead) based on size in a graphical format.
Note: You can view the Graph on Results, Advanced
™
, and Adjust screens.
1.
To view the graph showing the distribution live and/or dead cells based on cell
size, press the Graph buon.
2.
To view the distribution of only the live or dead cells, check the corresponding
Live or Dead check box on the graph.
The graph will automatically update and display the distribution of cells based
on size only in the selected population.
3.
Optional: Using the size, brightness, and circularity sliders, adjust the count
parameters. As you adjust the count parameters, the count results and the graph
will be automatically updated.
4.
To close the graph, press the Graph buon again.
Gate count results
The Adjust screen contains the controls for gating count results based on size,
brightness, and circularity. You can adjust the count parameters before or after
performing a count.
1.
On the Results screen, press More to open the Advanced screen.
2.
Optional: Press the Live and/or the Dead buon to identify the cells in the
selected population (see “Identify the cells counted“ on page 19).
View graph
Adjust screen
Gate count results
Chapter 5 Results
Graph count results
5
Countess
™
II automated cell counter User Guide
21
3.
On the Advanced screen, press Adjust to open the Adjust screen.
4.
Optional: Press the Graph buon to view the distribution of cells (live and/or
dead) based on size as you gate the count results (see “Graph count results“ on
page 21).
5.
Select the channel (Live or Dead) you wish to gate.
6.
Using the size, brightness, and circularity sliders, adjust the count parameters.
Note: For a description of the count parameters and count parameter controls
(i.e., parameter sliders), see “Count parameters“ on page 13.
7.
When nished, press Done to save the changes to count parameters and return
to the Advanced screen.
Press Cancel to return to the Results screen without saving the changes.
8.
On the Advanced screen, press Count to recalculate your results with the new
count parameters.
9.
To save the changes to size, brightness, or circularity parameters to the current
prole or to create a prole with the new count parameters, see “Save as new
protocol“ on page 23.
10.
To permanently save your results to an external computer using a USB ash
drive, see Chapter 7, “Save results“.
Chapter 5 Results
Gate count results
5
22
Countess
™
II automated cell counter User Guide
Save as new protocol
1.
If you have made any changes to the count parameters before or after performing
a count, the displayed prole name is appended with the (*) symbol and the
Advanced results screen displays the New protocol buon, which allows you to
save the changes to the current prole or as a separate protocol.
2.
To save the changes to the count parameters to the current prole or to create a
new prole with the edited parameters, press the New protocol buon. The
Select prole to edit screen opens.
Note: By default, the current prole buon is selected on the Select prole
screen. If you are using the Default prole for the count, no prole buon is
selected on this screen, because the Default prole cannot be edited.
3.
Select the prole you wish to edit, then press Import seings.
Note: You can select only a previously saved or an empty prole. The Default
prole cannot be edited.
Edit and save
profile as new
protocol
Chapter 5 Results
Save as new protocol
5
Countess
™
II automated cell counter User Guide
23
4.
The Edit prole screen opens and displays the edited count parameters from the
Adjust screen (21).
Note: If you have selected a prole that had been previously saved, the name of
that prole populates the Prole name text box by default. Otherwise, the
textbox remains empty.
5.
To change the name of the selected prole, press the Prole name text box and
enter the desired name using the alpha-numeric keypad.
6.
(Optional): If desired, make additional changes to the prole and the count
parameters as described in “Add/edit a prole“ on page 14.
7.
Click Save to save the new prole seings and return to the Results page for the
last count. The prole name will be displayed without the “(*)” symbols.
Click Cancel to return to the Results page for the last count without saving the
changes to the prole. The prole name will be displayed with the (*) symbol,
indicating that the count parameters for the selected prole had been altered, but
not yet saved.
Chapter 5 Results
Save as new protocol
5
24
Countess
™
II automated cell counter User Guide
Dilution calculator
Dilution calculator
Dilution calculator function allows you to calculate the volume of cell sample and
buer needed to reach a desired concentration using the count results.
Calculate dilution
1.
On the Results screen, press Dilution Calculator to open the Dilution calculator
screen.
2.
Press the cell type buon located to the right of the current concentration, then
select the count result you wish to use for the dilution calculation from the
dropdown. The current concentration changes to reect the results for the cell
type selected.
Available Cell type options are Live cells or Total cells. By default, Live cells is
selected.
6
Countess
™
II automated cell counter User Guide
25
3.
Enter the desired cell concentration (“What is your desired cell concentration?”):
a.
Press the value text box, then enter the value using the number pad.
You can enter one digit to the left of the decimal separator (integer part) and
one to the right (fractional part). If you do not enter the fractional part, the
software enters a 0 by default.
Press Enter or touch anywhere outside the number pad to close it.
b.
Press the exponent buon, then select the exponent value for the desired
concentration.
By default, the exponent is (n–1), where n = the exponent value of the
current count. The maximum selectable exponent is same exponent value as
the current count.
Pressing an exponent buon selects that exponent and closes the window.
Pressing anywhere outside the window keeps the previously selected
exponent and closes the window.
4.
Press the total volume text box (“How many mL do you need?”), then enter the
total volume of the sample you wish to have at the new concentration using the
number pad.
By default, the total volume box is blank. The maximum volume you can enter is
999.9 mL and you can use only a single decimal.
Press Enter or touch anywhere outside the number pad to close the number pad.
Chapter 6 Dilution calculator
Calculate dilution
6
26
Countess
™
II automated cell counter User Guide
5.
When you have made valid entries for cell type, desired cell concentration, and
total volume, and closed the last popup window, the boom line of the dilution
calculator displays the volumes of cell solution and buer needed.
If you enter a combination of values that is not valid (e.g., desired concentration
is greater than the current concentration), the results line remains on (or return
to) the blank state, and screen displays a warning message.
If you make any changes to any of the input areas above, the results are
recalculated automatically upon closing the popup window.
6.
Press Done or the Back buon to return to the main Results screen.
Chapter 6 Dilution calculator
Calculate dilution
6
Countess
™
II automated cell counter User Guide
27
Save results
Save count results
The Countess
™
II FL Automated Cell Counter allows you to save your data and
images using a USB ash drive. To save your experiment, you can choose from the
following options, in any combination:
• Result: Saves the Results screen as it is displayed on the instrument, with or
without the Graph, in the selected image format (JPEG, BMP, PNG, or TIFF).
• Images: Saves only the raw captured image in the selected image format (JPEG,
BMP, PNG, or TIFF).
• Data: Saves the data from the experiment as a CSV le (comma separated values).
The CSV format allows for processing or re-displaying results with any third
party software or spreadsheet program. For more information on the CSV le
format, see “Appendix D, “CSV le format“”.
• Report: Saves a printer-friendly report of the results, graph(s), and image in the
selected format (PDF, PNG, or JPEG). For more information, see “Report“ on
page 31.
Note: If you wish to save your results with the Graph showing the distribution of
cells based on cell size or uorescence intensity, make sure that the desired graph is
displayed on the Results screen.
1.
To save your data, insert the Countess
™
II USB drive (or equivalent) into an
available USB port on the instrument (see “Instrument exterior components“ on
page 10).
Note: The USB ports located in the front and the back of the instrument function
the same. However, the rst USB drive connected will be the preferred saving
location.
2.
On the Results screen, press Save to navigate to the Save screen.
7
Save screen
Save procedure
28
Countess
™
II automated cell counter User Guide
3.
To assign a name to your experiment, press the File name text eld. The alpha-
numeric keypad opens.
4.
Enter the le name using the alpha-numeric keypad.
To enter symbols, press the symbol (@%&) key. To return to the alpha-numeric
keypad, press ABC.
5.
Press Enter to save the name and return to the Save screen.
To return to the Save screen without saving the name, press the close buon.
6.
Select the desired mode(s) to save your experiment (Result, Images, Data,
Report). You can select an individual mode (e.g., Result only) or any combination
of modes (e.g., Result, Images, Data, and / or Report).
In the example below, Data and Report are selected.
7.
By default, Result and Images are saved as JPEG les, and Report is saved as
PDF.
To choose a dierent le format, press the le type buon. The Choose le type
screen opens.
Note: Data can only be saved as a CSV le.
Chapter 7 Save results
Save count results
7
Countess
™
II automated cell counter User Guide
29
8.
Press to select the desired le type. Available options are JPEG, BMP, PNG, and
TIFF.
After you make your selection, the instrument returns to the Save screen.
To return to the Save screen without changing the le format, press the close
buon.
9.
Press Save to save your experiment in the selected mode(s) in the USB drive.
10.
Press Close and then transfer the USB drive to the desired location.
Chapter 7 Save results
Save count results
7
30
Countess
™
II automated cell counter User Guide
Report
The Report function allows you to save a printer-friendly report of the results, graphs,
and images in the selected format (PDF, PNG, or JPEG).
You can create reports using the Report dropdown as described in “Save count
results“ on page 28.
1
The top section of the Report contains a table with the results as displayed on the
Countess
™
Results screen, showing the concentration of the sample, and the percentage
and number for the total, Live, and Dead channels.
2
Below the results table, the report contains the “number of cells vs. cell size” graph.
3
Under the graph, the report contains the brightfield count image, with the live and dead
cells identified by the green and red circles, respectively.
4
At the bottom, the report displays the profile information used to gate these images.
Report file
Chapter 7 Save results
Report
7
Countess
™
II automated cell counter User Guide
31
Instrument settings
Overview
To access the Instrument Seings screen, press the Instrument Seings buon on the
Home page (“Turn ON the instrument“ on page 11).
In the Instrument Seings screen, you can:
• perform software update (“Software update“ on page 32)
• set the date and time (“Date/Time“ on page 34)
Note: Although the Change Light Cube buon is visible on the Instrument Seings
screen, this buon is not functional on the Countess
™
II Automated Cell Counter,
which is a brighteld-only instrument and does not support EVOS
™
light cubes.
Software update
• The USB drive used for transferring the software update le must be FAT32
formaed; verify this before proceeding. If necessary, reformat the USB drive to
FAT32 following the recommended procedure for your operating system.
Note: Reformaing the USB drive will result in the loss of all les. Back up the
les in the USB drive prior to reformaing.
• The software update le must be saved on the top level of the USB drive, not
within a folder or a subfolder.
• The software update le must be uncorrupted during transfer. Do not rename,
zip, or compress the software update le.
8
Instrument
settings screen
Guidelines for
software update
32
Countess
™
II automated cell counter User Guide
1.
Go to www.thermosher.com/countessupdate, and download the latest
Countess
™
II/II FL software version to your desktop.
Note: The software update le has a version-specic name followed by the
extension .lft (e.g., Countess
™
_II_v_1_0_202.lft for software version 1.0.202).
2.
Copy the software update le onto the USB drive, making sure that it is saved on
the top level and not hidden within a folder.
3.
Insert the USB drive into one of the USB ports of the instrument (see “Instrument
exterior components“ on page 10).
4.
Press the Instrument Seings buon on the Home page (“Turn ON the
instrument“ on page 11) to open
the Instrument Seings screen
(“Overview“ on page 32).
5.
Select Software Update from the Instrument Seings menu. The instrument
scans the USB drive for the latest software version.
6.
When prompted, select Update Now.
7.
Once the update has completed, restart the instrument.
Update the
Countess
™
II/II FL
software
Chapter 8 Instrument settings
Software update
8
Countess
™
II automated cell counter User Guide
33
Date/Time
1.
Press the Instrument Seings buon on the Home page (“Turn ON the
instrument“ on page 11) to open the Instrument Seings screen.
2.
Press Date/Time on the Instrument Seings menu to open the Date/Time screen.
3.
Select the Date format you wish to use.
4.
Press any Date text box (MM, DD, or YYYY) to open the Edit Date keypad.
5.
Using the keypad, enter the date into Month, Day, and Year text boxes, pressing
Enter after each entry.
6.
After you are nished entering the date, press the close buon to return
to the
Date/Time screen.
Set the date and
time
Chapter 8 Instrument settings
Date/Time
8
34
Countess
™
II automated cell counter User Guide
7.
Select the Time format you wish to use. Available options are 12 Hour and
24 Hour formats.
8.
Press any Time text box (Hours or Minutes) to open the Edit Time keypad.
9.
Using the keypad, enter the time into Hours and Minutes text boxes, pressing
Enter after each entry.
10.
After you are nished entering the time, press the close buon to return to
the Date/Time screen.
11.
If you have selected the 12 Hour format, select AM or PM.
12.
Press Save to set the Time and Date and return to the Instrument Seings screen.
Press Cancel to return to the Instrument Seings screen without saving your
changes.
Chapter 8 Instrument settings
Date/Time
8
Countess
™
II automated cell counter User Guide
35
Maintenance
Instrument care
• Use the appropriate cleaning solutions for each component, as indicated in the
cleaning procedures in “Clean the Countess
™
II automated cell counter“ on
page 37.
• If liquid spills on the instrument, turn o the power immediately and wipe dry.
Always use the correct power supply. The power adaptor specications appear on the
serial number label (boom of the instrument) and in the Technical specications
section of this user guide (“Technical specications“ on page 43). Damage due to an
incompatible power adaptor is not covered by warranty.
CAUTION! Never disassemble or service the instrument yourself. Do not
remove any covers or parts that require the use of a tool to obtain access to
moving parts. Operators must be trained before being allowed to perform the
hazardous operation. Unauthorized repairs may damage the instrument or
alter its functionality, which may void your warranty. Contact your local
distributor to arrange for service.
IMPORTANT! If you have any doubt about the compatibility of decontamination or
cleaning agents with parts of the equipment or with material contained in it, contact
Technical Support () or your local distributor for information.
9
General
guidelines for care
Power supply
36
Countess
™
II automated cell counter User Guide
Clean the Countess
™
II automated cell counter
We recommend cleaning the Countess
™
II Automated Cell Counter periodically to
prevent the buildup of dust and dirt that might reduce its performance and cause
contamination.
CAUTION! To avoid electrical shock, always turn o the Countess
™
II
Automated Cell Counter and unplug the power cord before cleaning or
decontaminating the instrument.
CAUTION! All biological samples and materials that come into contact with
them have the potential to transmit infectious diseases and are considered
biohazardous. Follow all applicable local, state/provincial, and/or national
regulations. Wear appropriate protective eyewear, clothing, and gloves.
IMPORTANT! Using a cleaning or decontaminating method other than that specied
by the manufacturer may result in damage to the instrument.
• Wipe the touch-screen of the Countess
™
II Automated Cell Counter using a soft,
lint-free cloth moistened with an LCD cleaning solution. Do not apply excessive
force during cleaning. Wipe the touch-screen dry immediately after cleaning.
• Ensure that the cleaning solution does not enter the power buon, the power
inlet, the slide port, or the USB ports.
• Never pour or spray any liquids directly on the instrument to avoid electrical
shock when the instrument is plugged in.
• Do not use abrasive cleaning solutions or material to prevent the touch-screen
from geing scratched.
• Wipe the instrument case of the Countess
™
II Automated Cell Counter using a
soft, lint-free cloth moistened with distilled water. Wipe the instrument dry
immediately after cleaning.
• Ensure that water or other cleaning solutions do not enter the power buon, the
power inlet, the slide port, or the USB ports.
• Never pour or spray any liquids directly on the instrument to avoid electrical
shock when the instrument is plugged in.
• Wipe the instrument case of the Countess
™
II Automated Cell Counter using a
soft, lint-free cloth moistened with 70% alcohol. Wipe the instrument dry
immediately after cleaning.
• Avoid using a bleach solution, because it may leave a residue of bleach crystals
on the instrument.
• Ensure that water or other cleaning solutions do not enter the power buon, the
power inlet, the slide port, or the USB ports.
• Never pour or spray any liquids directly on the instrument to avoid electrical
shock when the instrument is plugged in.
Introduction
Clean the touch-
screen
Clean the
instrument case
Decontaminate
the instrument
Chapter 9 Maintenance
Clean the Countess
™
II automated cell counter
9
Countess
™
II automated cell counter User Guide
37
Set nominal focus
Nominal focus is the Z-point (i.e., depth) around which the auto focus function
searches to provide ne focus to the sample.
The auto focus algorithm of the Countess
™
II FL Automated Cell Counter is designed
to highlight the dierences between live and dead cells in the brighteld channel. The
optimal focus level is where the “live” cells have a light colored center and the “dead”
cells are dark throughout (see examples below).
Focus is not optimal Focus is optimal
To enable optimal auto focus functionality, you may need to initially rene the
brighteld focus by adjusting it manually and then seing the nominal focus. This
allows the auto focus function to have a set point from which to focus on the cells in
subsequent samples.
1.
Prepare the sample by adding 10 µL of cell suspension to 10 µL of 0.4% trypan
blue stain. Mix the sample mixture well by pipeing up and down a few times.
2.
Load 10 µL of the sample mixture into the Countess
™
Cell Counting Chamber
Slide (“Load Countess
™
chamber slide“ on page 16). Let the sample mixture
sele for 30 seconds to ensure a uniform focal plane.
3.
Insert the sample slide into the slide port of the instrument, making sure that the
side containing the sample is inserted completely.
4.
When the slide is inserted, the instrument automatically illuminates the sample,
sets the intensity of the brighteld light source, and auto focuses on the cells.
5.
To manually adjust the focus, press the Focus
™
buon.
Overview
Set nominal focus
Chapter 9 Maintenance
Set nominal focus
9
38
Countess
™
II automated cell counter User Guide
6.
Use the Focus
™
slider or the plus and minus buons to rene the brighteld
focus.
Note: If needed, Zoom in on the image to adjust focus or lighting.
7.
After nding the optimal focus, press Set to set the nominal focus.
Once the focus has been set, the Set buon on the focus slider becomes inactive,
conrming that the focus seing has been stored.
Chapter 9 Maintenance
Set nominal focus
9
Countess
™
II automated cell counter User Guide
39
Troubleshooting
Note: The software for the Countess
™
II is updated regularly. If you are having any
issues with your experiments, rst check the website to see if a new software version
is available. You can download the most recent version of the software from
www.thermosher.com/countessupdate. You can also register your Countess
™
II
instrument at www.thermosher.com/registercountess to be informed of any future
software updates.
Problem
Possible solutions
Autofocus does not seem to focus on the cells very well
• Make sure there are no bubbles or debris visible on
the screen that could interfere with the autofocus
and make it more difficult to get the sample in the
correct focal plane.
• Ideally, the live cells should have bright centers
compared to the dead cells, which are dark
throughout (“Count procedure“ on page 17).
• Setting the nominal focus will improve autofocus
consistency with future slides. To set the nominal
focus, see “Set nominal focus“ on page 38.
Some cells appear on the image but are not included in
the count
• Adjust the size, brightness, and circularity gates for
both live and dead cells to include all of the cells in
the count (“Graph count results“ on page 21).
• After including all of the cells in the count, you can
narrow the count criteria, if you wish to exclude cells
of a certain size or certain brightness.
• When the gates are fully maximized, the CSV should
indicate 0–60 for cell size and 0–255 for brightness.
Images are very bright and washed out Enable Auto Lighting from the Profiles menu, or decrease
the bright field light intensity before counting the cells.
Getting incorrect concentration for the Countess
™
test
beads
• The beads can settle quickly in solution, which will
affect the concentration reading.
• Vortex the bead stock on high for a full 30 seconds to
resuspend, and add 10 µL of the bead suspension to
10 µL of trypan blue without delay.
• Pipet the bead and trypan blue mixture up and down
several times to make sure it is well mixed, and
immediately load 10 µL into the slide.
A
40
Countess
™
II automated cell counter User Guide
Problem Possible solutions
Variable counts for the same sample of cells
• If you are pipetting different samples from the same
cell sample, the variability could be due to pipetting
or mixing.
• Use recently calibrated pipettors and make sure that
the cells are well suspended by pipetting up and
down several times before adding trypan blue.
• Pipet the bead and trypan blue mixture up and down
several times to make sure it is well mixed, and load
10 µL into the slide without delay.
Variable counts when performing replicate counts of the
same slide
• If you are counting replicates of the exact same
slide, visually inspect that all cells are counted
correctly in the image.
• There may be a slightly different field of view each
time a slide is inserted. Depending on the
concentration and uniformity of the cells, this will
cause some variability when performing replicate
counts of the same slide, although it should be less
than 10%.
• When counting fewer cells, a small field of view
change for only a small number of cells can have a
larger affect. Count cells at a higher concentration to
reduce variability.
• Make sure that you do not shake or agitate the slide
between counts.
Abnormally high percentage of dead cells or live cells
counted as dead
• Ensure that the cells are focused correctly so that
live cells have bright centers and dead cells are dark
throughout (see “Count procedure“ on page 17). If
the cells are not well focused and look dark on the
screen, the Countess
™
II will count them as dead
cells.
• If cells are well focused, have bright centers, and are
being counted as dead, confirm that they are within
the appropriate cell size range and try adjusting the
settings.
• If cells are exposed to trypan blue for a long period
of time, viability could be affected so slide should be
prepared and counted fresh each time for best
results.
• Gate out the debris using the size, brightness, and
circularity sliders.
USB drive not recognized by the instrument
• The USB drive must be FAT32 formatted to be
recognized by the instrument. If it is not, reformat
the USB drive to FAT32 (“Software update“ on
page 32).
• Try another correctly formatted USB drive.
Appendix A Troubleshooting
Set nominal focus
A
Countess
™
II automated cell counter User Guide
41
Problem Possible solutions
Unable to update the Countess
™
software
• Make sure the USB drive is formatted to FAT32. If it
is not, reformat the USB drive to FAT32 before
transferring the files onto the USB drive for software
update.
• The update file must sit on the top level of the USB
drive, not within a folder or a subfolder.
• File cannot be renamed in any way.
• File cannot be zipped or compressed during
distribution. It must be uncorrupted during transfer
and have a .lft suffix.
• If needed, check that the USB port is functional by
testing a USB mouse.
Appendix A Troubleshooting
Set nominal focus
A
42
Countess
™
II automated cell counter User Guide
Product specifications
Technical specifications
Instrument type
Benchtop cell counter and suspension cell-based assay
platform
Instrument dimensions 9 (W) × 5½ (D) × 9 inches (H)
Weight 8 lbs
Operating power 100–240 VAC, 0.58 A MAX
Frequency 50/60 Hz
Electrical input 12 VDC, 2 A
Installation site Indoor use only, Class A Environments (i.e., non-residential
or light industrial); Pollution degree 2.
Operating temperature 4–32°C (40–90°F)
Operating humidity <80% (non-condensing)
Processing time
~15 seconds
Sample concentration range 1 × 10
4
–1 × 10
7
cells/mL
Particle/cell diameter range 4–60 µm (particles); 7–60 µm (cells)
Required sample volume 10 µL
Firmware Countess
™
Automated Cell Counting Platform Software
USB Drive 4 Gigabyte
Optics
1 channel (brightfield)
Camera 5 Mega pixels, 2.5× Optical Magnification
Material
Poly(methyl methacrylate) (PMMA)
Dimensions 25 mm (W) × 75 mm (D) × 1.7 mm (H)
Chamber volume 10 µL
B
Physical
characteristics
Technical
specifications
Optics
Analysis slide
Countess
™
II automated cell counter User Guide
43
Ordering information
Countess
™
II automated cell counter
The Countess
™
II instrument and instrument accessories are available from Thermo
Fisher Scientic. For more information, visit www.thermosher.com or contact
Technical Support (“Customer and technical support“ on page 56).
Product
Quantity Cat. no.
Countess
™
II Automated Cell Counter 1 each AMQAX1000
Countess
™
II power cord with four adapter cords 1 each AMEP4716
Countess
™
II USB drive 1 each A25751
Countess
™
II Disposable Slide Holder 1 each AMEP4745
Accessory products
The following products can be used with the Countess
™
II Automated Cell Counter
and are available separately from Thermo Fisher Scientic. For more information, visit
www.thermosher.com or contact Technical Support (“Customer and technical
support“ on page 56).
Product
Quantity Cat. no.
Countess
™
Cell Counting Chamber Slides, 50 Slides (100
counts)
1 box
[1]
C10228
Countess
™
Cell Counting Chamber Slides, 500 Slides (1000
Counts)
10 boxes
[1]
C10312
Countess
™
Cell Counting Chamber Slides, 1250 Slides
(2500 Counts)
25 boxes
[1]
C10313
Countess
™
Cell Counting Chamber Slides, 2500 Slides
(5000 Counts)
50 boxes
[1]
C10314
Countess
™
Cell Counting Chamber Slides, 5000 Slides
(10,000 Counts)
100 boxes
[1]
C10315
Countess
™
Test Beads (1 × 10
6
beads/mL) 1 mL C10284
Trypan blue stain (0.4 %) 2 × 1 mL T10282
[1]
Each box of Countess
™
Cell Counting Chamber Slides contains 50 slides and 2 × 1 mL vials of trypan blue
(0.4%), sufficient for 100 counts.
C
44
Countess
™
II automated cell counter User Guide
CSV file format
CSV file format, explained
A comma-separated values (CSV) le stores tabular data (numbers and text) in plain-
text form. Plain text means that the le is a sequence of characters, with no data that
has to be interpreted as binary numbers. A CSV le can be opened with any third
party software or spreadsheet program. The table below describes the categories of
the Countess
™
II data saved as a CSV le and opened with a spreadsheet program.
Category
Column Name Description
General A Number Sequential sample run number
B File Name Name of file
C Date & Time Date and time of sample run
D Mode BF-Brightfield
Trypan
Blue/Brightfield
E Total Concentration Concentration of the entire sample
F Total cells counted Total number of cells counted in the sample
G Live concentration Concentration of just the "live" portion of the sample
H Live cells counted Total number of "live" cells counted
I Dead concentration Concentration of just the "dead" portion of the sample
J Dead cells counted Total number of "dead" cells counted
K Viability (%)
Percent viability of the sample based on trypan blue
staining
L Average size (um) Average cell size in microns
General Details X Focus value Focal position number
Y BF Light intensity Brightfield light intensity value from 0–100%
Trypan
Blue/Brightfield
Count Parameters
Z Live Size min Minimum size of "live" cells in microns
AA Live Size max Maximum size of "live" cells in microns
AB Live Brightness min "Live" adjustment slider value for minimum brightness
AC Live Brightness max "Live" adjustment slider value for maximum brightness
AD Live Circularity "Live" adjustment slider value for circularity
AE Dead Size min Minimum size of "dead" cells in microns
D
Overview
Countess
™
II automated cell counter User Guide
45
Category Column Name Description
Trypan
Blue/Brightfield
Count Parameters
AF Dead Size max Maximum size of "dead" cells in microns
AG Dead Bright min "Dead" adjustment slider value for minimum brightness
AH Dead Bright max "Dead" adjustment slider value for maximum brightness
AI Dead Circ "Dead" adjustment slider value for circularity
Appendix D CSV file format
CSV file format, explained
D
46
Countess
™
II automated cell counter User Guide
Safety
WARNING! GENERAL SAFETY. Using this product in a manner not specied
in the user documentation may result in personal injury or damage to the
instrument or device. Ensure that anyone using this product has received
instructions in general safety practices for laboratories and the safety
information provided in this document.
·
Before using an instrument or device, read and understand the safety
information provided in the user documentation provided by the
manufacturer of the instrument or device.
·
Before handling chemicals, read and understand all applicable Safety Data
Sheets (SDSs) and use appropriate personal protective equipment (gloves,
gowns, eye protection, and so on). To obtain SDSs, see the “Documentation
and Support” section in this document.
Safety alert words
Four safety alert words appear in this document at points where you need to be aware
of relevant hazards. Each alert word—IMPORTANT, CAUTION, WARNING,
DANGER—implies a particular level of observation or action, as dened below:
IMPORTANT! – Provides information that is necessary for proper instrument
operation, accurate installation, or safe use of a chemical.
CAUTION! – Indicates a potentially hazardous situation that, if not avoided,
may result in minor or moderate injury. It may also be used to alert against
unsafe practices.
WARNING! – Indicates a potentially hazardous situation that, if not avoided,
could result in death or serious injury.
DANGER! – Indicates an imminently hazardous situation that, if not avoided,
will result in death or serious injury. This signal word is to be limited to the
most extreme situations.
Except for IMPORTANT! safety alerts, each safety alert word in this document
appears with an open triangle gure that contains a hazard symbol. These hazard
symbols are identical to the hazard symbols that are axed to the instruments (see
Safety symbols in Appendix E).
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Electrical symbols
The following table describes the electrical symbols that may be displayed.
Symbol Description
Indicates the On position of the main power switch.
Indicates the Off position of the main power switch.
Indicates a standby switch by which the instrument is switched on to the
Standby condition. Hazardous voltage may be present if this switch is on
standby.
Indicates the On/Off position of a push-push main power switch.
Indicates a terminal that may be connected to the signal ground reference of
another instrument. This is not a protected ground terminal.
Indicates a protective grounding terminal that must be connected to earth
ground before any other electrical connections are made to the instrument.
Indicates a terminal that can receive or supply alternating current or
voltage.
Indicates a terminal that can receive or supply alternating or direct current
or voltage.
Safety symbols
The following table describes the safety symbols that may be displayed. Each symbol
may appear by itself or in combination with text that explains the relevant hazard (see
“Safety labels on instruments”). These safety symbols may also appear next to
DANGERS, WARNINGS, and CAUTIONS that occur in the text of this and other
product-support documents.
Symbol
Description
Caution, risk of danger. Consult the manual for further safety information.
Caution, risk of electrical shock.
Appendix E Safety
Electrical symbols
E
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Symbol Description
Caution, hot surface or other high-temperature hazard.
Caution, laser.
Caution, moving parts.
Caution, potential biohazard.
Caution, ultraviolet light.
Environmental symbols
The following symbol applies to all Thermo Fisher Scientic electrical and electronic
products placed on the European market after August 13, 2005.
Symbol
Description
Do not dispose of this product as unsorted municipal waste. Follow local
municipal waste ordinances for proper disposal provisions to reduce the
environmental impact of waste electrical and electronic equipment (WEEE).
European Union customers:
Call your Customer Service representative for equipment pick-up and
recycling. See www.thermofisher.com for a list of customer service offices in
the European Union.
Appendix E Safety
Environmental symbols
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Safety labels on instruments
The following CAUTION, WARNING, and DANGER statements may be displayed
Thermo Fisher Scientic instruments in combination with the safety symbols
described in the preceding section.
Symbol English Français
CAUTION! Hazardous chemicals. Read the Safety
Data Sheets (SDSs) before handling.
ATTENTION! Produits chimiques dangereux. Lire
les fiches techniques de sûreté de matériels avant
toute manipulation de produits.
CAUTION! Hazardous waste. Refer to SDS(s) and
local regulations for handling and disposal.
ATTENTION! Déchets dangereux. Lire les fiches
techniques de sûreté de matériels et la régulation
locale associées à la manipulation et l’élimination
des déchets.
DANGER! High voltage. DANGER! Haute tension.
WARNING! To reduce the chance of electrical
shock, do not remove covers that require tool
access. No user-serviceable parts are inside. Refer
servicing to Thermo Fisher Scientific qualified
service personnel.
AVERTISSEMENT! Pour éviter les risques
d’électrocution, ne pas retirer les capots dont
l’ouverture nécessite l’utilisation d’outils.
L’instrument ne contient aucune pièce réparable
par l’utilisateur. Toute intervention doit être
effectuée par le personnel de service qualifié
venant de Thermo Fisher Scientific.
DANGER! Class 3B visible and/or invisible laser
radiation present when open. Avoid exposure to
beam.
DANGER! Rayonnement visible ou invisible d’un
faisceau laser de Classe 3B en cas d’ouverture.
Evitez toute exposition au faisceau.
CAUTION! Moving parts. Crush/pinch hazard. ATTENTION! Pièces en mouvement, risque de
pincement et/ou d’écrasement.
General instrument safety
WARNING! PHYSICAL INJURY HAZARD. Use this product only as specied
in this document. Using this instrument in a manner not specied by Thermo
Fisher Scientic may result in personal injury or damage to the instrument.
Ensure that anyone who operates the instrument has:
• Received instructions in both general safety practices for laboratories and specic
safety practices for the instrument.
• Read and understood all applicable Safety Data Sheets (SDSs).
Operating the
instrument
Appendix E Safety
Safety labels on instruments
E
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• Do not install the instrument in heavy humidity, such as a greenhouse or an
incubator, to avoid a danger of electric shock. If water or other material enters the
instrument, the adaptor, or power inlet, disconnect the power cord and contact a
service person.
• Do not press the main plug or power cord with wet hands.
• Always ensure that the power supply input voltage matches the voltage available
in your location.
• Do not install the instrument on a slant or a place prone to vibrations, which
induces the risk of instrument malfunction or damage of the instrument.
• Never insert any objects into the air vents of the instrument as this could result in
electrical shock, personal injury, and equipment damage.
• Plug the power cord rmly into the wall outlet and the instrument.
• To avoid potential shock hazard, make sure that the power cord is properly
grounded.
• Be sure to position the equipment such that it is easy to disconnect the
instrument.
• Turn o the instrument before unplugging the power cord and/or moving the
instrument.
• If the instrument is broken or dropped, disconnect the power cord and contact a
service person. Do not disassemble the instrument.
• Use only authorized accessories (adaptor, power cord, and USB drive).
• If the instrument emits smoke, disconnect the power cord from the wall outlet
and contact a service person.
CAUTION! Using cleaning or decontamination methods other than those
recommended by the manufacturer may compromise the safety or quality of
the instrument.
CAUTION! PHYSICAL INJURY HAZARD The instrument is to be serviced
only by trained personnel or vendor specied in the user guide.
Safety precautions
Cleaning or
decontaminating
the instrument
Removing covers
or parts of the
instrument
Appendix E Safety
General instrument safety
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Chemical safety
WARNING! GENERAL CHEMICAL HANDLING. To minimize hazards,
ensure laboratory personnel read and practice the general safety guidelines for
chemical usage, storage, and waste provided below. Consult the relevant SDS
for specic precautions and instructions:
·
Read and understand the Safety Data Sheets (SDSs) provided by the
chemical manufacturer before you store, handle, or work with any chemicals
or hazardous materials. To obtain SDSs, see the "Documentation and
Support" section in this document.
·
Minimize contact with chemicals. Wear appropriate personal protective
equipment when handling chemicals (for example, safety glasses, gloves, or
protective clothing).
·
Minimize the inhalation of chemicals. Do not leave chemical containers open.
Use only with sucient ventilation (for example, fume hood).
·
Check regularly for chemical leaks or spills. If a leak or spill occurs, follow
the manufacturer cleanup procedures as recommended in the SDS.
·
Handle chemical wastes in a fume hood.
·
Ensure use of primary and secondary waste containers. (A primary waste
container holds the immediate waste. A secondary container contains spills
or leaks from the primary container. Both containers must be compatible
with the waste material and meet federal, state, and local requirements for
container storage.)
·
After emptying a waste container, seal it with the cap provided.
·
Characterize (by analysis if needed) the waste generated by the particular
applications, reagents, and substrates used in your laboratory.
·
Ensure that the waste is stored, transferred, transported, and disposed of
according to all local, state/provincial, and/or national regulations.
·
IMPORTANT! Radioactive or biohazardous materials may require special
handling, and disposal limitations may apply.
WARNING! HAZARDOUS WASTE (from instruments). Waste produced by
the instrument is potentially hazardous. Follow the guidelines noted in the
preceding General Chemical Handling warning.
WARNING! 4L Reagent and Waste Bole Safety. Four-liter reagent and waste
boles can crack and leak. Each 4-liter bole should be secured in a low-density
polyethylene safety container with the cover fastened and the handles locked in
the upright position.
Chemical waste safety
CAUTION!
HAZARDOUS WASTE. Refer to Safety Data Sheets (SDSs) and
local regulations for handling and disposal.
Chemical waste
hazard
Appendix E Safety
Chemical safety
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To minimize the hazards of chemical waste:
• Read and understand the Safety Data Sheets (SDSs) provided by the
manufacturers of the chemicals in the waste container before you store, handle, or
dispose of chemical waste.
• Provide primary and secondary waste containers. (A primary waste container
holds the immediate waste. A secondary container contains spills or leaks from
the primary container. Both containers must be compatible with the waste
material and meet federal, state, and local requirements for container storage.)
• Minimize contact with chemicals. Wear appropriate personal protective
equipment when handling chemicals (for example, safety glasses, gloves, or
protective clothing). For additional safety guidelines, consult the SDS.
• Minimize the inhalation of chemicals. Do not leave chemical containers open. Use
only with adequate ventilation (for example, fume hood). For additional safety
guidelines, consult the SDS.
• Handle chemical wastes in a fume hood.
• After emptying the waste container, seal it with the cap provided.
• Dispose of the contents of the waste tray and waste bole in accordance with
good laboratory practices and local, state/provincial, or national environmental
and health regulations.
If potentially hazardous waste is generated when you operate the instrument, you
must:
• Characterize (by analysis, if necessary) the waste generated by the particular
applications, reagents, and substrates used in your laboratory.
• Ensure the health and safety of all personnel in your laboratory.
• Ensure that the instrument waste is stored, transferred, transported, and disposed
of according to all local, state/provincial, and/or national regulations.
IMPORTANT! Radioactive or biohazardous materials may require special handling,
and disposal limitations may apply.
Electrical safety
Part
Symbol Description
General DANGER! ELECTRICAL SHOCK HAZARD. Severe electrical shock
can result from operating the Countess
™
II Automated Cell
Counter or Countess
™
II FL Automated Cell Counter without its
instrument panels in place. Do not remove instrument panels.
High-voltage contacts are exposed when instrument panels are
removed from the instrument.
Fuses WARNING! FIRE HAZARD. For continued protection against the
risk of fire, replace fuses only with fuses of the type and rating
specified for the instrument.
Chemical waste
safety guidelines
Waste disposal
Appendix E Safety
Electrical safety
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Part
Symbol Description
Power DANGER! ELECTRICAL HAZARD. Grounding circuit continuity is
vital for the safe operation of equipment. Never operate equipment
with the grounding conductor disconnected.
DANGER! ELECTRICAL HAZARD. Use properly configured and
approved line cords for the voltage supply in your facility.
DANGER! ELECTRICAL HAZARD. Plug the system into a properly
grounded receptacle with adequate current capacity.
Overvoltage rating The Countess
™
II Automated Cell Counter and Countess
™
II FL
Automated Cell Counter have an installation (overvoltage)
category of II, and are classified as portable equipment.
Biological hazard safety
WARNING! Biological samples such as tissues, body uids, and blood of
humans and other animals have the potential to transmit infectious diseases.
Follow all applicable local, state/provincial, and/or national regulations. Wear
appropriate protective eyewear, clothing, and gloves. Read and follow the
guidelines in these publications.
ATTENTION! BIOHAZARD. Les échantillons biologiques tels que les tissus, les
uides corporels et le sang des humains et d’autres animaux ont la possibilité de
transmere des maladies infectieuses. Suivre tous les règlements municipaux,
provinciaux/provincial et / ou nationales en vigueur. Porter des lunees de protection
approprié, des vêtements et des gants.
In the U.S.:
• U.S. Department of Health and Human Services, Biosafety in Microbiological and
Biomedical Laboratories (BMBL), 5th Edition, HHS Publication No. (CDC) 21-1112,
Revised December 2009; found at: www.cdc.gov/labs/pdf/CDC-
BiosafetymicrobiologicalBiomedicalLaboratories-2009-P.pdf
• Occupational Safety and Health Standards, Bloodborne Pathogens (29
CFR§1910.1030; www.access.gpo.gov/nara/cfr/waisidx_01/29cfr1910a_01.html)
• Your company’s/institution’s Biosafety Program protocols for working
with/handling potentially infectious materials.
• Additional information about biohazard guidelines is available at: www.cdc.gov
Appendix E Safety
Biological hazard safety
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In the EU:
• Check your local guidelines and legislation on biohazard and biosafety
precaution, and the best practices published in the World Health Organisation
(WHO) Laboratory Biosafety Manual, third edition
www.who.int/csr/resources/publications/biosafety/WHO_CDS_CSR_LYO_2004
_11/en/
Safety and electromagnetic compatibility (EMC) standards
Symbol
Description
U.S. and Canadian safety standards.
The CSA C/US Mark signifies that the product meets applicable U.S. and Canadian standards,
including those from CSA, CSA America, ANSI, ASME, ASSE, ASTM, NSF and UL.
European safety and EMC standards.
The CE Mark symbolizes that the product conforms to all applicable European Community
provisions for which this marking is required. Operation of the instrument is subject to the
conditions described in this manual.
The protection provided by the instrument may be impaired if the instrument is used in a manner
not specified by Thermo Fisher Scientific.
Australian EMC standards
The C-Tick Mark indicates conformity with Australian and New Zealand standards for
electromagnetic compatibility.
Appendix E Safety
Safety and electromagnetic compatibility (EMC) standards
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Documentation and support
Customer and technical support
Visit thermosher.com/support for the latest service and support information.
• Worldwide contact telephone numbers
• Product support information
– Product FAQs
– Software, patches, and updates
– Training for many applications and instruments
• Order and web support
• Product documentation
– User guides, manuals, and protocols
– Certicates of Analysis
– Safety Data Sheets (SDSs; also known as MSDSs)
Note: For SDSs for reagents and chemicals from other manufacturers,
contact the manufacturer.
Limited product warranty
Life Technologies Corporation and/or its aliate(s) warrant their products as set forth
in the Life Technologies' General Terms and Conditions of Sale at
www.thermosher.com/us/en/home/global/terms-and-conditions.html. If you have
any questions, please contact Life Technologies at www.thermosher.com/support.
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