Invitrogen Countless C10227 User manual

User Manual
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Invitrogen Corporation
1600 Faraday Avenue
Carlsbad, CA 92008
T: 1 760 603 7200
F: 1 760 602 6500
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For country-specic contact information visit our web site at www.invitrogen.com
Countess™Automated
Cell Counter
Catalog no. C10227
Rev. Date: 15 September 2009
MP10227

ii

iii
Table of Contents
Product Contents......................................................................................................... iv
Safety Information ........................................................................................................v
Product Specifications ............................................................................................... vii
Description of Countess™Automated Cell Counter ........................................... viii
Introduction.......................................................................................... 1
Product Overview.........................................................................................................1
Methods................................................................................................ 3
Getting Started...............................................................................................................3
General Guidelines .......................................................................................................9
Using the Countess™Automated Cell Counter .....................................................10
Cleaning and Maintenance........................................................................................19
Updating and Calibrating the Countess™Automated Cell Counter..................20
Troubleshooting ..........................................................................................................22
Countess™Automated Cell Counter Error Codes.................................................24
Appendix ............................................................................................ 25
Accessory Products.....................................................................................................25
Technical Support .......................................................................................................26
Purchaser Notification................................................................................................27

iv
Product Contents
Countess™
Automated Cell
Counter
The contents of the Countess™automated cell counter are listed
below:
Component Quantity
Countess™automated cell counter 1
Power Cord with 4 adaptor cords 1
(for U.S./Canada/Taiwan/Japan, Europe, or UK)
Countess™cell counting chamber slides 1 box of 50
Trypan blue stain (0.4%) 2 × 1 mL
Countess™USB drive 1
Quick reference card (QRC) 1
The complete instruction manual is available on our website at
www.invitrogen.com/countess.
See page vii for specifications and description of the Countess™
automated cell counter.
Upon
Receiving the
Instrument
Examine the instrument carefully for any damage incurred
during transit. Ensure that all parts of the instrument including
accessories listed above are included with the product. Any
damage claims must be filed with the carrier. The warranty does
not cover in-transit damage.
See page 3to install the instrument.
Upon receipt, store trypan blue stain at room temperature.

v
Safety Information
Safety
Precautions Review and follow the safety instructions below.
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. For operating environment, refer to
Product Specifications (page vii).
Do not touch the main plug or power cord with wet hands.
Always ensure that the power supply input voltage matches
the voltage available in your location.
This instrument is air-cooled so its surfaces may become hot
during operation. When installing the instrument, leave a
space of more than 10 cm (4 inches) around it and do not
place any objects between the instrument and the wall.
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 an
y
ob
j
ects into the air vents of the instrument as
this could result in electrical shock, personal injury, and
equipment damage.
Plug the power cord firmly into the wall outlet and AC
adapter.
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 off 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).
Continued on next page

Safety Information, Continued
Safety
Precautions,
continued
Do not use metal objects when assembling or dissembling a
battery.
Do not heat the battery or expose it to heat. For operating
environment, see Product Specifications (page vii).
If the instrument emits smoke, disconnect the power cord
from the wall outlet and contact a service person.
Symbols The symbols used on Countess™automated cell counter are
explained below.
Used on the instrument to indicate a warning. Consult the
manual to avoid possible personal injury or instrument damage.
Protective earth (Ground)
WEEE (Waste Electrical and Electronic Equipment) symbol
indicates that this product should not be disposed of in unsorted
municipal waste. Follow local municipal waste ordinances for
proper disposal provisions to reduce the environmental impact
of WEEE. Visit www.invitrogen.com/weee for collection and
recycling options.
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 Invitrogen.
This product conforms to UL61010-1/CSA C22.2 No. 61010-1
“Safety Requirements for Electrical Equipment for Measurement,
Control, and Laboratory Use, Part I: General Requirements.”
Instruments bearing the TUV symbol are certified by TUV
Product Services to be in conformance with the applicable safety
standards for the US and Canada.
vi

vii
Product Specifications
Countess™
Automated Cell
Counter
Specifications
Environmental Conditions
Operating Power 100–240 VAC, 1.5 A
Frequency 50/60 Hz
Electrical input 12 VDC, 1.0 A
Installation site: Indoor use only
Operating Temperature: 10–35°C
Maximum Relative Humidity: 20–80%
Altitude: <2,000 m
Transient Category: Installation categories II
Pollution Degree: 2
Degree of Protection: IPX0
Instrument Specifications
Instrument Type: Benchtop cell counter
Processing Time: <1 minute
Cell Sample Range: 1 × 104–1 × 107
Software: Countess™Software
www.invitrogen.com/countess
Instrument Dimensions: 27 cm (w) × 20 cm (d) ×
19 cm (h)
Weight: 2.1 kg (4.6 lbs)
The Countess™automated cell counter is equipped with a camera
(2.3X objective and 3.1 Mega pixel).
Countess™Cell Counting Chamber Slide Specifications
Material: Polymethyl methacrylate
(PMMA)
Dimensions: 75 mm (w) × 25 mm (d) ×
1.8 mm (h)
Chamber Depth: 100 μm
Chamber Volume: 10 μL
Countess™USB Drive: 1 Gigabyte

Description of Countess™Automated Cell
Counter
Front View of
Countess™
Automated
Cell Counter
The Countess™automated cell counter is a benchtop automated
cell counter that performs cell count and viability measurements
using trypan blue stain.
The front view showing various parts of the Countess™
automated cell counter is shown below.
Power button
The Power Button is used to turn the instrument on and off. The
red status light indicates that the instrument is off; the green
status light indicates that the instrument is on.
Touchscreen display located in the front of the instrument
contains buttons for all the functions needed and displays data
from the cell count.
Slide port
The Slide port is used to insert the Countess™cell counting
chamber slide containing the sample with trypan blue into the
counter for analysis.
USB port
The USB port allows you to transfer and save the cell count data
and image to your computer for record keeping and printing
purposes. The USB drive supplied with the instrument or any
other standard USB drive is inserted into the USB port for data
transfer. See page 16 for Transferring Data to a PC.
Slide
Port
USB port Power
button
Touchscreen
Continued on next page
viii

Description of Countess™Automated Cell
Counter, Continued
Rear and Side
View of
Countess™
Automated
Cell Counter
The rear and side view showing various parts of the Countess™
automated cell counter.
Power Inlet
Connect the counter to an electrical outlet using the supplied
power cord and the appropriate plug, based on the electrical outlet
configuration in your country.
Image Adjustment (Focus) Knob
The Image Adjustment (Focus) knob is used to adjust the image
quality to obtain better contrast between live (bright centers) and
dead (dark blue centers) cells. This is important to obtain accurate
cell counts and viability measurements. See page 14 for details.
Focus Lock Knob
The Focus lock knob is used to lock the Image Adjustment (Focus)
knob once the image is optimized. There is no need to use the
Focus lock knob, but is available for convenience, if you are
measuring multiple samples of the same cell type.
Rear View Side View
Focus
knob
Power
inlet Focus
lock
Continued on next page
ix

Description of Countess™Automated Cell
Counter, Continued
User Interface The touch screen user interface of the Countess™automated cell
counter is used to operate the instrument and consists of:
The touch screen buttons to operate the instrument.
The Digital Display shows the image of cells and sample
data.
Touch
Screen
Button
Countess™
Cell Counting
Chamber Slide
The Countess™cell counting chamber slides are plastic, disposable
enclosed chambers that hold the sample in two separate chambers
(A and B) for replicates. The cell counting occurs in the central
location of the counting chamber. The entire volume of cells
counted is 0.4 μL, the same as counting four (1 mm × 1 mm)
squares in a standard hemocytometer.
x

1
Introduction
Product Overview
Countess™
Automated
Cell Counter
The Countess™automated cell counter uses state-of-the-art optics
and image analysis to automate cell counting. The Countess™
automated cell counter is a benchtop counter designed to measure
cell count and viability (live, dead, and total cells) accurately and
precisely, using the standard trypan blue technique.
Using the same amount of sample that you currently use with the
hemocytometer, the Countess™automated cell counter takes less
than a minute per sample for a typical cell count and is compatible
with a wide variety of eukaryotic cells and provides information
on cell size.
The Countess™automated cell counter offers an intuitive user
interface, and provides the option to save and print cell count data
using the Countess™software (download from
www.invitrogen.com/countess) and USB drive supplied with the
instrument or available separately.
The Countess™automated cell counter is supplied with disposable
Countess™cell counting chamber slides that contain two enclosed
chambers to hold the sample to allow you to measure two different
samples or perform replicates of the same sample. The cell
counting occurs in the central location of the counting chamber
and the volume counted is 0.4 μL, the same as counting four
(1 mm × 1 mm) squares in a standard hemocytometer.
See page viii for details on various parts of the counter.
Continued on next page

2
Product Overview, Continued
System
Overview The Countess™automated cell counter performs viability and cell
counting measurements using the trypan blue method of dead-cell
staining combined with advanced image analysis.
Cell sample is mixed with trypan blue and loaded into a Countess™
cell counting chamber slide. The camera acquires cell images from
the sample on the slide and the image analysis software
automatically analyzes acquired cell images, and measures cell
count and viability using the trypan blue stain.
A single, sample measurement within a minute provides the
following data:
Live and dead cell concentration/mL
Total cell concentration/mL
Viability (% live cells to total cells)
Mean diameter
Cell images
Graphical data representation
Features Important features of the Countess™automatic cell counter are:
User-friendly, benchtop design for simple, fast, automated cell
count and viability measurements within a minute
Provides data on cell size and is compatible with a wide variety
of eukaryotic cells without the need for any special changes
between large or small sizes
Measures cell concentrations ranging from 1 × 104to 1 × 107
cells/mL and cells with sizes ranging from 5 μm to 60 μm
Uses dis
p
osable countin
g
chamber slides that eliminate washin
g
steps and cross contamination between samples
Saves and prints cell count data including images using the
Countess™software and Countess™USB drive
Presents comprehensive data with graphical reports and as a
.CSV (comma separated value) file for sample comparisons
Allows you to gate and analyze cells based on cell size

3
Methods
Getting Started
Installing the
Countess™
Automated
Cell Counter
1. After unpacking the instrument, place the instrument on a flat,
level, dry surface.
2. Plug one end of the supplied power cord into the Countess™
instrument. To the other end attach the appropriate plug
adaptor, based on the electrical outlet configuration in your
country.
3. 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.
To register your Countess™automated cell counter, visit
www.invitrogen.com/countess. Enter the instrument serial
number, your name, and your contact details. Registering your
instrument ensures that you receive notifications of software
upgrades and fast service.
4. When you are ready to use, start the Countess™instrument by
pressing the Power button.
Continued on next page

Getting Started, Continued
Optional:
Installing the
Countess™
Software
The Countess™instrument is designed for stand-alone use and does
not require the use of an external computer.
If you wish to archive data and generate reports, you must transfer
data to your computer, and use the Countess™software to generate
and
p
rint re
p
orts (see instructions, below). Alternativel
y
, data stored
in the .CSV file may be transferred to your computer by the USB
drive and imported into any spreadsheet pro
g
ram, without the need
for Countess™software.
Computer requirements (Countess™is not compatible with
Macintosh Operating systems)
USB port (1.1 or later, 2.0 is recommended)
Windows XP/2000/Vista
1. Visit www.invitrogen.com/countess and follow instructions
on the page to install the Countess™software.
2. When the software is installed, the Countess™automated cell
counting software icon appears on your desktop.
3. To download data, use any USB drive to capture data from
your experiment and transfer data to your computer as
described in Transferring Data to a PC (page 16).
Start-Up
Screen When the instrument is turned on, the Start Up screen is
displayed. Here you can proceed immediately to cell counting
(page 9), set up the instrument for cell or bead count, or adjust the
screen brightness.
Start Up Screen
Screen Brightness
Use the scroll buttons to adjust the screen brightness.
Continued on next page
4

Getting Started, Continued
Settings
Screen Press Settings from the Start-Up screen to display Settings.
The Settings menu allows you to set up the following:
Parameters (see below and next page for details)
Calibration to calibrate the instrument (page 20)
Update to install new firmware versions as they become
available
Date and Time to set up date and time (page 8)
Count mode to operate the instrument for cell counting
(choose Cells) or bead counting (choose Beads)
Cell Mode
Parameters
Screen
Press Parameters from the Settings screen to display Cell Mode
Parameters Screen.
The Countess™instrument comes with pre-set parameters to match
the majority of cultured cell types which can be accurately counted
without changing any of the default parameters. The Parameters
function allows you to change the image analysis algorithm for
specific or mixed cell types, and the specific parameters must be
determined empirically.
Continued on next page
5

6
Getting Started, Continued
Cell Mode
Parameters
Screen,
continued
The Cell Mode Parameters are described below:
Sensitivity (refers to the contrast of the objects from the
background). Adjusting the sensitivity higher makes the
instrument more sensitive to objects; useful for cells that do not
stain well with trypan blue while adjusting the sensitivity
lower makes the instrument less sensitive and is useful if there
is a lot of background.
Minimum cell size is used to select the lowest particle size to
include in the count. The size is set in 1 micrometer
increments. The lower range of the Countess™instrument is
5 μm. However, you can select a minimum cell size as low as
2 μm; when a cell size outside of the usual range is selected, the
numbers indicating particle size are shown in red.
Maximum cell size is used to select the maximum particle size
to include in the count. The size is set in 1 micrometer
increments. The maximum size that can be set is 60 μm.
Circularity is used to determine the objects to include in the
measurement based on roundness. Increasing the value from
80% requires objects to be more round for inclusion in the
measurement. Decreasing the value from 80% allows objects to
be less round. Adjusting this may be useful if the cell type is
not particularly circular or perhaps oddly shaped.
After modifying any parameters:
Press Apply to make the changes or,
Press Default to restore default parameters or,
Press Save to create a protocol. Save the protocol name and
user name in the appropriate fields, and press Save again. The
protocol is now saved and appears highlighted in the protocol
menu. To use the highlighted protocol, press Apply.
Continued on next page

Getting Started, Continued
Cell Mode
Parameters
Screen,
continued
Note: Once a protocol is saved, it is available for use at any time. To access
saved protocols, press Settings > Parameters > Load Protocol. Use the up
and down arrows to find your saved protocol. Once the protocol is located,
use the touch screen to highlight the protocol and press Apply. Once the
protocol of interest is selected, count cells using the standard procedure.
7

Getting Started, Continued
Date and
Time Setup The date and time is already preset when you receive the
instrument. To reset the date and time, follow these steps:
1. Turn on the Countess™automated cell counter by pressing the
Power button. The Start up screen is displayed after a few
seconds.
2. Press Settings and then press Date and Time.
3. The Date/Time Properties screen is displayed. To select the
month and day, scroll to the appropriate month usin
g
the arrow
keys and then press the day on the calendar.
Note: Use a pointed object, like a stylus or pipette tip, to push the small
buttons on the calendar.
4. To select the time, scroll to the appropriate time and select
Automatically adjust clock for daylight saving, if needed.
Press Apply or OK to make the date/time changes. The
updated date/time is displayed on the top of the window.
Once the date/time is set, there is no need to set it each time the
instrument is turned on.
5. Press Close to exit the screen.
8

General Guidelines
Countess™
Cell Counting
Chamber
Slides
The Countess™cell counting chamber slides supplied with the
counter or available separately from Invitrogen (page 25) are
specially designed for use exclusively with the Countess™
automated cell counter. Use of other slides result in inaccurate cell
counts and can damage the Countess™instrument.
R
E
C
O
M
M
E
N
D
A
T
I
O
N
To obtain the best results, follow these recommendations:
Wear gloves during sample handling.
Do not touch the optical surfaces of the Countess™cell
counting chamber slides. Hold the chamber slides by the
edges.
Use the Countess™automated cell counter at room
temperature only (10–35ºC).
For accurate viability count results, ensure the counting area is
covered with cell suspension and count cells within 3 minutes
of mixing the cells with trypan blue solution as trypan blue is
toxic to cells. For best data with biological samples, we
recommend counting at least two samples and taking an
average.
The Countess™automated cell counter is supplied pre-
calibrated. To recalibrate your instrument, see page 20.
The Countess™automated cell counter memory holds one set
of data. Save your data to the USB drive after each reading.
You may transfer the data to your PC, using the USB drive
immediately as described in Transferring Data to a PC (page
16).
After using Countess™cell counting chamber slides,
appropriately dispose slides as biohazardous waste. Do not
reuse the chamber slides.
9

Using the Countess™Automated Cell Counter
The Countess™automated cell counter is supplied
precalibrated. However, if you wish to calibrate the Countess™
automated cell counter, see page 20.
Instructions are provided in this section for preparing the cell
sample with trypan blue dye for use with disposable
Countess™cell counting chamber slides for automated cell
count using the Countess™counter.
Visit www.invitrogen.com/countess for additional application
notes, technical notes, and a list of validated cell lines
including adherent, primary, and suspension cells that have
been tested using the Countess™Automated Cell Counter.
Materials
Needed
Cell sample
Countess™cell counting chamber slides (supplied with the
instrument or available separately, page 25)
Trypan blue stain (0.4%); supplied with the instrument or
available separately, page 25
Optional: Countess™ USB drive for data transfer, supplied with
the instrument or available separately (page 25)
Cell Counting
Procedure 1. Push the Power button to start the instrument. The Start-up
screen is displayed.
2. The Countess™automated cell counter is preset to Cell mode.
If you have changed the mode to bead for bead counting (page
17) or calibration (page 21), be sure to change the mode to
Cells Mode prior to counting cells by pressing Settings and
then Cells.
Continued on next page
10