English - Merck Millipore Auto2D Plus Quick Start Manual

English

Intended Use
The Auto2D Electrophoresis Device fully
®
automates the process of 2D electrophoresis,
separating proteins first by isoelectric point and
second by molecular weight. Complete protein
separation is achieved in just over one hour.
Contents
• Auto2D Electrophoresis Device
®
• Electrical Power Cables:
• Power cable for JP
and North America
• 3P-2P Conversion Plug
for JP and North America
• Power cable for Europe
• Power cable for China
• Power cable for UK
Also Needed
(not included)
See Product Ordering, p.52 for catalogue
numbers.
• Auto2D Electrode Chip Plus
®
• Auto2D PAGE Chip
®
• Auto2D ® IEF Chip
• Auto2D Solution Chip Plus
®
• Auto2D Tris-Glycine or Tris-Tricine
®
Reagent Kit
• Ampholyte (choose appropriate
ampholyte based on IEF Chip pH range)
• Distilled water
• Filter paper 0.3 mm thick
User Guide Online
The complete User Guide includes detailed
instructions and recipes. It can be downloaded
from the Auto2D ® Device product page at
SigmaAldrich.com.
Storage and Stability
The Auto2D device should be used/stored
®
indoors only. See each component's product
label for storage requirements.
20424816 Rev 06/21
Y Y Precautions
For research use only.
Please read the Safety Sheet (enclosed) and
the complete User Guide at SigmaAldrich.com
carefully before using this product.
Auto2D Plus Workflow
®
Total process: 66–138 minutes
(depending on the recipe)
Sample Preparation
Application of Reagents
Gel Rehydration 3 Minutes
Sample Loading to Gel 10 Minutes
First Dimension:
17 Minutes
Isoelectric focusing
SDS Equilibration 3 Minutes
Second Dimension:
33 Minutes
SDS-PAGE
Detection
2
Preparation
Reagents
1. Prepare and store Reagent Kits as
directed in the literature provided with the
kits, or the complete User Guide (online).
2. Allow the following products stored at
low temperatures to equilibrate to room
temperature (20–25 °C approximately
10 minutes).
• IEF Chip
• PAGE Chip
• Rehydration Solution
• DTT Solution
• Ampholyte
3. Prepare Working Rehydration Solution for
rehydration of IEF Chip and extraction/
dilution of protein sample:
Reagents
Rehydration
Solution
DTT Solution
(1M)
Ampholyte*
Total
* Select ampholyte according to the range
of IEF Chip to use. For ampholytes at
40% stock concentration, add 1 µL. For
ampholytes at 100X, add 2 µL.
4. Prepare Working Equilibration Buffer for
equilibration of focused proteins before
SDS-PAGE.
Reagents
Equilibration
buffer premix
DTT Solution
(1M)
Total
Sample Preparation
Dissolve the protein sample in the Working
Rehydration Solution, prepared according to
step 3. Sample may be diluted 2-fold or more
with Working Rehydration Solution to reach
desired protein concentration and decrease
salt concentration.
English
20424816 Rev 06/21
A high salt concentration can affect protein
separation during isoelectric focusing and
cause the current to exceed 100 µA. Samples
with a high salt concentration should be
desalted by either:
TCA/acetone precipitation and resuspension
of proteins in working rehydration solution
OR
Buffer exchange using a spin column
OR
Desalting protocol using Auto2D
(See complete User Guide online, for more
information.)
NOTE: During protein resuspension, avoid
heating proteins over 37 °C.
Quantify proteins. Between 0.1–100 µg of
protein can be loaded for 2D electrophoresis.
The optimum amount of protein will depend on
detection method and sample complexity. The
Final
following protein amounts should be used as a
Volume Concentration
starting point, users may need to optimize the
loading amount for their particular sample.
189 µL
• Coomassie Brilliant Blue staining: 50 µg
10 µL 50 mM
• Fluorescent staining: 10 µg
1–2 µL 0.5–1% v/v
• Silver staining: 5 µg
200 µL
• Fluorescent pre-labeling: 3 µg
Use the amount of protein loaded into Solution
Chip to determine whether to use S, M, or L
recipe type for protein separation.
The sample volume applied to the device is
13–15 µL. Sample can be diluted with Working
Rehydration Solution, as necessary.
Power the Auto2D
Final
Volume Concentration
Select the appropriate power cord (provided
with the device) that is compatible with the
760 µL
power outlets in your country. Firmly push the
matching end into the back of the Auto2D
40 µL 50 mM Device, and the other end into the power outlet.
The power switch is located next to the AC plug
800 µL
port on the back of the Auto2D
the switch up to turn on.
The application will be automatically launched.
From the screen, select Auto2D
3
Device
®
Device
®
®
® Device. Press
Plus mode.
®
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