Lonza Amaxa Nucleofector II User manual

Amaxa™ Nucleofector™ II Manual

For future protocol updates, also check www.lonza.com/cell-database
For more information and future program updates,
check www.lonza.com/nucleofection-program-update
Nucleofector™ II CD-ROM
Nucleofector™ II Chip Card

Contents
1
Contents
1.0 The Amaxa™ Nucleofector™ Technology 3
2.0 Operating Instructions 5
2.1 Restrictions 5
2.2 Maintenance 5
2.3 Safety Instructions–Please Read Carefully 6
2.4 Waste Disposal 7
2.5 Nucleofector™ II Device Components 7
2.6 Set-up Instructions 7
2.7 Instructions for Using the Nucleofector™ II Device 8
2 .7.1 Main menu and program execution 8
2.7.2 Selection of Nucleofector™ Programs by program code–“Free Program Choice” 8
2.7.3 Selection of Nucleofector™ Programs by name of cell type–“Cell Type List” 8
2 . 7. 4 Selection and storage of Nucleofector™ Custom Programs 9
2.7.5 Program execution 9
2.8 Additional Nucleofector™ II Settings and Options 10
2.8.1 Change of standard settings 10
2.8.2 Motor 10
2.8.3 Display contrast 10
2.8.4 Beep on/o 10
2.8.5 Change of start mode 10
2.8.6 Language 11
2.8.7 Software version 11
2.8.8 Upgrading program software 11
2.8.9 Serial number 11
2.8.10 96-well mode 11

2
3.0 Troubleshooting 12
4.0 Nucleofector™ II Device Error Codes 14
5.0 Appendix 16

3
The Amaxa™ Nucleofector™ Technology 1.0
1.0 The Amaxa™
Nucleofector™
Technology
Since its introduction in 2001, a constantly growing interest in
the scientic community and industrial markets established
the Amaxa™ Nucleofector™ Technology as a leading transfection
Technology all around the world. A continuously increasing number
of publications from many dierent research areas reect how the
Nucleofector™ Technology drives research in numerous dierent
cell types and applications.
The Nucleofector™ Technology oers you more opportunities than
any other transfection method to approach your research topics
from many dierent angles.
Transfection of virtually any cell
With over 100 protocols for cell lines optimized by Lonza’s R&D
team, more than 500 entries in the online cell data base and
almost 50 Nucleofector™ Kits for dierent primary cells, the
Nucleofector™ Technology is clearly the transfection method-of-
choice for virtually any dicult-to-transfect cell type.
Transfection of any substrate
The Nucleofector™ Technology oers high exibility with respect
to applications, since the same transfection parameters apply
for any nucleic acid substrate used. DNA vectors, e.g., expression
plasmids or shRNA vectors, and RNA, e.g., siRNA duplexes, can
be transfected using our transfection protocols. It is the ideal
tool to help you answer the scientic questions in your day-to-
day applications such as over-expression studies, gene silencing
approaches, protein expression, generation of stable clones and
many more.
Constant Innovation
At Lonza, we believe that listening to our customers needs is the
best way to develop innovative products that exactly meet your
demands. In line with this dedication, the Nucleofector™ II Device
was developed which oers a larger graphic display, a motor-
driven carousel, the possibility to store individual programs for
optimized cell lines and to perform bacterial transformations.
In addition, the Nucleofector™ II Device serves as the program
delivery unit for Lonza’s Nucleofector™ 96-well Shuttle™ System,
a 96-well transfection unit that allows e.g., high throughput
transfection of siRNA and cDNA libraries in virtually any cell,
including primary cells.
What has been our mission in the past will continue to be so in
the future–providing you with innovative transfection solutions to
help you thrive and excel in your research.

4
Components of the Nucleofector™ Technology
The Nucleofector™ Technology is based on two unique components,
the Nucleofector™ Device that delivers the specically developed
electrical parameters and Nucleofector™ Kits that contain cell-
specic and optimized Nucleofector™ Solutions.
The Nucleofector™ II Device with Kit Components
As part of the Nucleofector™ Technology, Lonza provides cell-type
specic Amaxa™ Optimized Protocols for many dierent cell lines
and primary cells. An up-to-date database of the continuously
growing range of Amaxa™ Optimized Protocols can be found on our
homepage at www.lonza.com/cell-database.
If no Amaxa™ Optimized Protocol for your cell line of interest is
available, we oer a Cell Line Optimization Nucleofector™ Kit
enabling you to easily establish the Nucleofector™ Technology
for your specic cell line. For ne-tuning after the optimization
process, please contact our Scientic Support Teams.
Phone Europe/World: [phone]
USA: 800-521-0390
E-mail Europe/World: scienti[e-mail]
USA: scienti[e-mail]

Operating Instructions 2.0
5
2.1 Restrictions
Medical use restrictions
The Nucleofector™ Technology is intended for research and
investigational use by professionals only. Please note that Lonza’s
Nucleofector™ Technology is not intended to be used for diagnostic
purposes, for testing or treatment in humans.
License statement
Lonza Cologne GmbH is holder of various patents, patent
applications, copyrights and technical and scientic experience
with respect to the Nucleofector™ Technology. Use of Lonza’s
Nucleofector™ Technology and/or related software requires a
license from Lonza Cologne GmbH.
Purchasers are granted a non-exclusive, non-transferable
license for a limited use of Lonza’s Nucleofector™ Technology and
related software for research and development purposes, the
terms of which are disclosed in detail in the license agreement
accompanying the shipped Nucleofector™ Device. Commercial
application is allowed under Lonza’s license for for-prot-entities.
Both licenses exclude in particular any right to manufacture, copy,
reproduce, transmit, distribute, seIl, lease, transfer or sublicense
Lonza’s Nucleofector™ Technology and/or related software to any
third party. For license information contact Lonza Cologne GmbH
by phone [phone] or e-mail [e-mail].
2.2 Maintenance
The Nucleofector™ II Device requires very little maintenance to
assure reliable operation. To clean and disinfect the case, rst
unplug the power supply. Use a damp cloth to wipe down the
outer case (water or 70–80% ethanol). Avoid wetting the cuvette
holder within the cuvette carousel and the connectors located on
the rear of the device.
The Nucleofector™ II Device has been designed for use under
a sterile hood with or without UV radiation source. Prolonged
exposure of the outer casing to UV light will lead to discoloration
with no functional impairment of the Nucleofector™ Device. When
laminar ows are sterilized on a regular basis by UV radiation
overnight we recommend protection of the device by appropriate
shielding or removal during extended UV exposure.
The Nucleofector™ II Device is protected by two main fuses. Both
are inside a receptacle incorporated in the inner power socket (see
Figure 2, page 7). In case of a blown fuse, you can easily replace
it. Disconnect the Nucleofector™ II Device from power and insert
a small at screwdriver into the slot on the right-hand side of the
receptacle to pull it open. To function, both the upper and lower part
of the receptable must hold working fuses in the inner positions.
Blown fuses can usually be identied by molten interrupted
wires inside the glass tube. Only use T630mA or L250V fuses to
substitute blown fuses.
2.0 Operating Instructions

6
2.3 Safety Instructions–
Please Read Carefully
This symbol means that there is a risk of electric shock. An electric
shock could cause death or personal injury.
The Nucleofector™ II Device has been certied by international
safety standards and is safe to use when operated in accordance
with this manual.
This device is designed to deliver variable high voltage electrical
impulses for the purpose of introducing nucleic acids into
eukaryotic and prokaryotic cells.
These electrical impulses can be DEADLY.
Therefore, use this device with care and take the following
precautions:
— Only use the device once you have read and understood the
Nucleofector™ II Manual. The manual should be accessible
for all users. Make sure that each potential user reads and
understands it.
— Do not open the device. The device does not contain user-
serviceable parts. Under no circumstances should circuit
components be interfered with, as they can deliver an electric
shock even when system is not in operation.
— Do not alter the device in any manner.
— Do not obstruct the movement of the carousel of the
Nucleofector™ Device.
— Do not plug any non-Lonza-certified connectors into the
external electrodesockets attherearside oftheNucleofector™
Device. The sockets are high-voltage outputs that can deliver
electric shocks via any non-Lonza device that is connected to
the Nucleofector™ Device. They are designed to connect Lonza
certified add-on devices to the Nucleofector™ II Device, e.g.,
the 96-well Shuttle™ Device.
— Only use the device when it is set on top of a safe, plain and
stable table or bench.
— Place the device such that easy removal of the power cord is
possible at any time.
— Do not expose the device to a humid environment.
— The device shall not be exposed to direct sunlight nor be
placed in a hot environment.
— The device is not approved for use in fire or explosion
endangered areas, nor for use with inflammable or explosive
media.
— Employ precautions against great impact and vibration in
moving and transporting the Nucleofector™ II Device.
— Use the device with Lonza’s certified Nucleofector™ Solutions
and Lonza certified cuvettes only. Use of any other solution
or cuvettes from any other source than Lonza will preclude all
warranty and liability claims.
— Standard Nucleofector™ Solutions and 96-well Nucleofector™
Solutions (used for the Nucleofector™ 96-well Shuttle™
System) are not compatible. Please do not use the
Nucleofector™ II Device with the 96-well Nucleofector™
Solutions.
— Unpack the cuvettes just prior to the experiment. Make sure
that the outer contact areas are dry.
— If any fluid has been spilled in the close vicinity of the
Nucleofector™ II Device, safety may be compromised. Ensure
that no fluid has been in contact with or entered the device.
— If any fluid has been spilled at the device, the safety may be
compromised. To confirm that the use of the device is safe,
contact Lonza Scientific Support for actions or precautions.
— Never place any foreign object on the device to avoid the risk
of equipment damage.
— Do not enter or place foreign objects in the carousel area of
the Nucleofector™ II Device.
— If any foreign object has entered the Nucleofector™ II Device,
safety may be compromised. To confirm that the use of the
device is safe, contact Lonza Scientific Support for actions or
precautions.
— If the Nucleofector™ II Device has been damaged, ensure that
the device can not be used by any personnel and contact
Lonza Scientific Support for assistance.
— All service shall be performed by Lonza authorized personnel
only.
— Handling of device parts that has the possible risk of sample
contamination shall always be performed with protective
gloves and any disposal of such parts must be according to
Federal, state or local procedures for clinical waste handling
and disposal. Use secure leak proof containers and avoid
unprotected handling of such parts.
Lonza Cologne GmbH disclaims all warranties and shall in no
event be liable for any kind of damages caused by or arising out of
any operation or use in violation of the above safety and handling
instructions.

7
Operating Instructions 2.0
2.4 Waste Disposal
Disposal of consumables from Nucleofector™ Kits (cuvettes,
pipettes and Nucleofector™ Solutions): Discard cuvettes, pipettes
and expired Nucleofector™ Solution residuals in a biohazard
container. Refer to your local waste management organization and
to the relevant laboratory safety instructions for proper disposal
practices.
2.5 Nucleofector™ II Device
Components
The Nucleofector™ II Device is delivered with the following
components:
— 1 Nucleofector™ II Device
— 1 Power cord
— 1 Nucleofector™ II Manual
Front panel of the Nucleofector™ II Device
1 Power button
2 Graphic display
3 “Up” button
4 “Down” button
5 Functional button “menu”
6 Functional button “options”
7 “X” button with start, exit or enter function
8 Automatic cuvette carousel
9 Cuvette holder
10 Chipcard slot
Rear side of Nucleofector™ II Device
Figure 2
11 Power cord receptacle
12 Spare fuse
13 External electrode sockets
For use within the Nucleofector™ 96-well Shuttle™ System
14 Receptacle for USB connection to Laptop
15 Receptacle for Serial connection to 96-well Shuttle™
The Nucleofector™ II Device comes with default programs and
contains a chipcard reader (10) below the front handle for program
updates.
2.6 Set-up Instructions
1. Remove all packing material.
2. Attach the power cord to the receptacle on the rear side of the
Nucleofector™ II Device (11) and plug it into an appropriate
electrical outlet. The device will automatically be in the stand-
by mode. Turn on the device by pressing the orange power
button in the upper left corner of the front panel (1). The
device performs a self test accompanied by a short intro in
the front graphic display (2). After powering up, the display
shows the main menu “Free Program Choice” (Figure 3.).
The start mode can be changed to the last program used as
described in chapter 2.8.5.
13 1112
14 15
1
510 7
6 4 9
2 3 8
Figure 3
Figure 1
Amaxa® Nucleofector® II Device
Amaxa® Nucleofector® II Device
Amaxa® Nucleofector® II Device
Amaxa® Nucleofector® II Device

8
2.7 Instructions for Using the
Nucleofector™ II Device
2 .7. 1 Main menu and program execution
The Nucleofector™ II Device oers two separate ways for program
selection. Programs can either be chosen by direct selection of
the program code, e.g., A-023, in the “Free Program Choice”
mode (Figure 3), which is comparable to program selection in
Nucleofector™ I Device. As a convenient alternative, programs
can also be selected by cell type code using the “Cell Type List”
(Figure 4). This list is divided in three consecutive sections:
1. All programs from currently available Amaxa™ Optimized
Protocols are led under the specic cell type name in
alphabetical order (Figure 4).
2. This alphabetical list for eukaryotic cells is followed by a list
of programs for the transformation of bacteria (for details
please contact Lonza’s Scientic Support Teams (Europe/
World: [phone]; USA: 800-521-0390) and ask
for the latest developments and experimental support).
3. The last section comprises custom programs (see 2.7.4.)
in chronological order. In addition, custom programs can
be found in the “Custom Programs” mode. You can switch
program modes by pressing the functional button “menu” (5)
several times until the required program mode appears and
conrm by pressing the functional button “go” (6).
Note
Only custom programs can be stored or deleted by the user. The
program codes of the Nucleofector™ II Device consist of two parts:
a one-digit alphabetical code followed by a three-digit numerical
code, e.g., A-023. This corresponds to the program code A-23 of
the Nucleofector™ I Device.
2.7.2 Selection of Nucleofector™ Programs by
program code–“Free Program Choice”
1. Turn on the Nucleofector™ II Device by using the power button
(1). Nucleofector™ II Device starts in the “Free Program
Choice” mode (Figure 3). If it does not start in the “Free
Program Choice” press the functional button “menu” (5)
repeatedly until “Go to Free Program Choice” appears and
conrm this pressing the functional button “go” (6).
2. The alphabetical rst digit blinks. Choose the appropriate
letter of the program code by using the “Up” and “Down”
buttons (3,4). Then move the cursor with the functional
buttons “<<” and “>>” (5,6) to the numerical three-digit code.
Select the appropriate number of the program code with the
“Up” and “Down” buttons (3,4). Continual pressing of the “Up”
and “Down” buttons allows for fast scrolling of choices. Your
Nucleofector™ II Device is now ready for program execution
(see 2.7.5).
2.7.3 Selection of Nucleofector™ Programs by
program code–“Free Program Choice”
1. Turn on the Nucleofector™ II Device by using the power button
(1). If the Nucleofector™ II Device starts in the “Free Program
Choice” mode (Figure 3), exit this mode by pressing the “X”
button (7). Press the functional button “menu” (5) several
times until “Go to Cell Type List” appears and conrm this
choice by pressing the functional button “go” (6).
2. Choose the appropriate program for your cell type (Figure
4) by navigating through the pre-programed list using the
“Up” and “Down” buttons (3, 4). The programs are displayed
in alphabetical order. The list is organized by rst the cell
name (e.g., T cells), descriptions of the species and further
details of the cell type are given in line two. Line four shows
the corresponding program itself. In cases where two or more
programs are available, the third line indicates the specicity
of the program such as for higher transfection eciency or for
higher cell viability. Your Nucleofector™ II Device is now ready
for program execution (see 2.7.5).
Mode Program can be
edited deleted
“Free Program Choice” Yes No
“Cell Type List” (pre-programmed) No No
“Custom programs” Yes Yes
Figure 4