VWR PerfectBlue Maxi S User manual

Instruction Manual – Bedienungsanleitung
PerfectBlue™
Horizontal Maxi Gel Systems /
Horizontale Maxigelsysteme
Maxi S, S Plus, M, L, xW,
Maxi M 'Revolution' & Maxi xW 'Revolution'

Instruction Manual PerfectBlue
™
Horizontal Maxi Gel Systems
VWR_v0617_ 1
CONT NTS
WARRANTY 2
PACKAGING LIST 2
SAF TY PR CAUTIONS 2
SYST M OV RVI W 3
Technical properties 3
G N RAL INSTRUCTIONS 4
Setting up the system and pouring the agarose gel 4
Loading of samples and electrophoresis 5
Visualisation 5
Cleaning 6
R QUIR D R AG NTS & R CIP S 7
lectrophoresis buffers 7
Agarose: Gel volumes and percentage 8
thidium bromide 8
Loading buffer/Sample buffer 9
Molecular weight marker 9
TROUBL SHOOTING 9
T CHNICAL SUPPORT AND ORD RING INFORMATION 11
PerfectBlue
™
Maxi S 11
PerfectBlue
™
Maxi M & Maxi M 'Revolution' 11
PerfectBlue
™
Maxi L 13
PerfectBlue
™
Maxi xW & Maxi xW 'Revolution' 13
Power Supplies 14
Agaroses 14
LIT RATUR 15

Instruction Manual PerfectBlue
™
Horizontal Maxi Gel Systems
VWR_v0617_ 2
WARRANTY
VWR guarantees that the horizontal electrophoresis system you have received has been thoroughly test-
ed and meets its published specification.
However, immediately upon arrival, please check carefully that the shipment is complete and has not
been damaged in transit. For missing parts or to report any kind of damage, please contact VWR (see
'T CHNICAL SUPPORT AND ORD RING INFORMATIONS'). Please retain all packaging materials until
the delivery has been completely checked since this will speed up the return of goods if required and
reduce environmental impact. Any form of returns, replacements or credit notes must be agreed in ad-
vance by VWR.
For the complete range of PerfectBlue
™
electrophoresis and blotting systems, VWR guarantees a warran-
ty period of 36 months if the products have been used solely according to the instruction manual unless
a different warranty has been offered in writing. No liability is accepted for loss or damage arising from
incorrect use. VWR's liability is limited to the repair or replacement of the unit or refund of the purchase
price, at VWR's discretion. VWR is not liable for any consequential damages. After the warranty period
has expired VWR can offer repairs.
VWR reserves the right to alter the technical specifications of the PerfectBlue
™
electrophoresis or blotting
systems without prior notice. This will enable us to implement developments as soon as they arise.
PACKAGING LIST
Unless requested otherwise, the following items are included in shipment for the models Maxi S, Maxi S
Plus, Maxi M, Maxi M 'Revolution', Maxi L, Maxi xW and Maxi xW 'Revolution':
one buffer chamber with corrosion-protected platinum electrodes
one safety lid with attached power cords
one UV-transmissible gel tray and two nd Gates
Maxi S: 3 combs, 1.5 mm thick, 12, 16 and 20 teeth
Maxi S Plus: 3 combs, 1.5 mm thick, 10, 17 and 34 teeth
Maxi M ('Revolution'): 3 combs, 1.5 mm thick, 16, 24 and 36 teeth
Maxi L: 4 combs, 1.5 mm tick, 2 each 25 and 50 teeth
Maxi xW ('Revolution'): 4 combs, 1.5 mm thick, 2 each 25 and 50 teeth
User Manual
SAF TY PR CAUTIONS
Please, read this Instruction Manual carefully before using the gel system.
Only use a C marked DC power supply.
Always disconnect the gel system from the power supply before removing the safety lid.
Always disconnect the gel system from the power supply when it is not in use or before moving it.
Running conditions for this unit should not exceed the maximum operating voltage or current.
Do not fill the chamber with running buffer above the maximum fill line.

Instruction Manual PerfectBlue
™
Horizontal Maxi Gel Systems
VWR_v0617_ 3
SYST M OV RVI W
The horizontal electrophoresis systems PerfectBlue™ Maxi S, Maxi S Plus, Maxi M ('Revolution'), Maxi L
and Maxi xW ('Revolution') are designed for the separation of medium to high sample numbers at
long distances and provide flat, even banding patterns and consistent results. The gel trays are sealed
inside the chamber by using nd Gates included in delivery. These gasketed nd Gates are specifically
engineered to fit accurately into the gel tray grooves eliminating the need to tape or seal the tray before
pouring gels. Utilizing the built-in leveling feature in all of the horizontal Maxi Gel Systems allows the
user to cast level gels quickly. Four gel sizes are available to provide maximum versatility. Depending
on the specific model, the UV-transmissible gel trays are equipped with 4 to 12 comb slots so that you
can run multiple sample sets to equal distances simultaneously.
VWR offers a wide variety of standard and microtiter combs in two different thicknesses (1 mm or 1.5
mm). Microtiter combs allow for speedy sample loading using a multi-channel pipette. Additional pre-
parative combs for large sample volumes and wall combs and casting dams to cast gels in smaller sizes
are available for increased versatility. For dividing up the gel tray, wall combs are put in the gel tray's
slots instead of a common comb but should get sealed using agarose of higher percentage before cast-
ing the gel. Casting dams can be inserted at any position into the gel tray. Due to the optimal fitting and
the heat conductance of the material neither adhesive tape nor concentrated agarose is required for
sealing by using the casting dams.
For detailed information on available accessories visit www.de.vwr.com or see 'T CHNICAL SUPPORT
AND ORD RING INFORMATION'.
The Maxi M 'Revolution' and Maxi xW
'Revolution' models are equipped with an inter-
nal buffer recirculation system. A trapping sys-
tem captures hydrogen bubbles which are pro-
duced at the cathode due to electrolysis, and
directs them through an ascending tube to the
opposing side of the buffer chamber where the
anode is located. During this hydrogen bubble
migration, the buffer circulates, preventing the
creation of detrimental pH or ion gradients.
Technical properties
PerfectBlue™ Cat. No. Gel size (W x
L)
Buffer volume
Voltage Current Time required
Maxi S P QL41-1325 13 x 25 cm 1600 ml 20 -
250 V
1 -
150 mA
60 -
120 min
Maxi S Plus P QL41-1525 15 x 25 cm 1800 ml 20 - 250
V
1 -
150 mA
60 -
120 min
Maxi M P QL41-2025 20 x 25 cm 2300 ml 20 -
250 V
1 -
150 mA
60 -
240 min
Maxi M 'Revolution'
P QL41-2025R 20 x 25 cm 2000 ml 20 -
250 V
1 -
150 mA
60 -
360 min
Maxi L P QL41-2340 23 x 40 cm 4500 ml 20 -
250 V
1 -
150 mA
60 -
360 min
Maxi xW P QL41-2325 23 x 25 cm 3000 ml 20 -
250 V
1 -
150 mA
60 -
240 min
Maxi xW
'Revolution' P QL41-2325R 23 x 25 cm 2800 ml 20 -
250 V
1 -
150 mA
60 -
360 min
Schematic drawing: 'Revolution'
-
Technology

Instruction Manual PerfectBlue
™
Horizontal Maxi Gel Systems
VWR_v0617_ 4
G N RAL INSTRUCTIONS
Setting up the system and pouring the agarose gel
1. Remove the lid from the gel box by holding the front of the buffer chamber with one hand and pull-
ing the lid off by holding the center of the back of the lid.
2. Slide the gasketed nd Gates into the outermost grooves on either side of the gel tray. The nd
Gates should be inserted tightly into the grooves with the gasket side facing out. Insert the sealed gel
tray into the buffer chamber and level the system by using the integrated three-point adjustable lev-
eling platform with bubble level.
Optional: The gel tray can be subdivided by a wall comb available as an accessory to pour shorter
gels. The wall comb should be sealed with 2 % agarose before pouring the real gel.
Alternatively, a casting dam available as accessory can be used for this purpose. This aluminum bar
can be inserted at any position into the gel tray. Due to the optimal fitting and the heat conductance
of the material neither adhesive tape nor concentrated agarose is required for sealing by using the
casting dams.
3. Use electrophoresis-grade agarose and compatible electrophoresis buffer to prepare the gel. The
percentage of agarose and the buffer to be used is determined by the size of the samples to be sep-
arated and further recovery of the samples (see 'R QUIR D R AG NTS & R CIP S'). The agarose
and buffer are mixed and heated over a heat plate by stirring or in a microwave oven until the
agarose is completely dissolved.
4. The prepared gel then must be cooled to below 60 °C before casting to avoid warping the UVT gel
tray due to excessive heat. If numerous gels are to be run in one day, a large volume of gel may be
prepared and be placed in a covered bottle stored 60 °C in a water bath.
5. Pour or pipette the measured amount (see 'Agarose: Gel volumes and percentage') of warm
agarose (< 60 °C) onto the UVT gel tray that has been placed into the correct position in the gel
box. Immediately after pouring, insert the desired comb or combs into the comb slots to form the
sample wells. Standard agarose should solidify completely in about 30 minutes. If low melting
agarose or a specialty agarose is used, please consult the instructions that came with the product.

Instruction Manual PerfectBlue
™
Horizontal Maxi Gel Systems
VWR_v0617_ 5
Loading of samples and electrophoresis
1. Once the gel is completely solidified, remove the nd Gates by carefully lifting out of the tray.
2. Pour enough compatible running buffer into the unit to fill chamber and completely cover and sub-
merge the gel. A 'Fill Line' is located on each unit to clearly mark the correct buffer level. See
'Technical properties' for approximate buffer volumes needed for your unit. Too little buffer may
cause the gel to dry out during the run, while excess buffer may slow DNA migration in the gel, in-
crease heat build-up and cause distorted bands.
3. Carefully remove the comb (or combs) by tapping lightly to loosen, and slowly lift straight up out of
the gel tray to avoid damage to the wells.
4. Load prepared samples into the wells. Samples should be mixed with a sample loading buffer (giv-
ing weight to the samples so that they drop evenly into the wells), and contain tracking dye to moni-
tor the gel run. For details on approximate well volumes see 'T CHNICAL SUPPORT & ORD RING
INFORMATION'. If microtiter combs have been used wells can be loaded by a multi channel pi-
pette. Wells that have been cast with microtiter combs of 26 teeth or less can be loaded 'directly',
i.e. the pipette tips fit sequentially into the gel wells. For gels prepared with microtiter combs with
more than 26 teeth the wells can be loaded 'shifted', i.e. the pipette tips fit only into every second
well.
NOT : It is wise to always run a sample lane of a known 'standard ladder' to determine concentra-
tion and size of separated fragments after the gel run, and to aid in photo documentation and anal-
ysis.
5. Carefully slide the lid with attached power cords onto the unit. This will connect the power cords to
the banana plugs to complete the circuit. Plug the other end of the power cords (4 mm, male) into
an appropriate power supply.
Take care to the proper orientation of the electrical field. Remember that nucleic acids are negatively
charged in an alkaline to neutral surrounding and therefore will migrate to the positively charged
anode. In general the color coding for positively charged electrodes is 'red'.
6. Turn on the power supply and run the gel at the appropriate voltage/current (see 'Technical proper-
ties'). You can observe the progress of electrophoresis by the visual migration of the loading dye.
Note that in 0.5 x TB gels, bromophenol blue co-migrates at 300 bp and xylene cyanol at 4 kbp
with the DNA fragments.

Instruction Manual PerfectBlue
™
Horizontal Maxi Gel Systems
VWR_v0617_ 6
Visualization
When the tracking dye has migrated as far through the gel as desired, or to the end of the gel, turn off
the power supply and slide off the lid to disconnect from the power source. Carefully remove the tray
containing the gel (wear gloves). The UV-transmissible gel tray makes for simple visualization and pho-
tography with a UV light source without the need to remove the gel from the tray. The gel tray may be
placed back into the casting chamber for convenient transport to the darkroom and to avoid damage to
the gel.
Cleaning
The buffer chamber, tray, combs and nd Gates should be rinsed under warm running water after each
use. Use a mild detergent to get rid of any debris. A following short rinse off with distilled water pre-
vents the formation of salt marks. It is recommended to allow the chamber to air dry rather than drying
with a towel to avoid damage to the electrode wires.
Do not use ethanol or other organic solvents to clean acrylic products, because organic solvents cause
acrylic to 'craze' or crack!

Instruction Manual PerfectBlue
™
Horizontal Maxi Gel Systems
VWR_v0617_
7
R QUIR D R AG NTS & R CIP S
lectrophoresis buffers
lectrophoresis buffers supply the ions necessary for electrophoresis and establishing a certain pH value
in which the target molecule adapts to its the required electric charge. Nucleic acids for example will be
negatively charged in an alkaline to neutral surrounding. Additionally, electrophoresis buffers often
contain reagents which protect the target molecule from degradation (e.g. DTA, which complexes biva-
lent cations and therefore inhibits DNases). If electrophoresis under denaturing conditions is desired
(like for the electrophoresis of RNA), electrophoresis buffers will additionally contain reagents that elimi-
nate the formation of secondary structures.
You will find recipes below for TA and TB , two of the most commonly used buffers for the electropho-
resis of DNA. If the intention is to eventually isolate DNA from the gel, TA buffer should be chosen. In
comparison to TB , migration will be faster and a better resolution of supercoiled DNA will be achieved
when using TA . However, because of TA 's limited buffering capacity, TB should be selected for per-
forming extended electrophoresis separations and if the electrophoresis chamber does not possess a
system for buffer recirculation. VWR's PerfectBlue 'Revolution' Systems are equipped with an internal
buffer recirculation system that prevents the formation of pH and ion gradients during extended runs.
Since agarose tends to create finer pore sizes and a more solid matrix in TB , diffusion of DNA will be
reduced and a more discrete band pattern will be achieved.
TA (Tris-Acetate- DTA) Buffer
1 x working solution: 40 mM Tris-acetate, 1 mM DTA
50 x stock solution (1 L): 242 g Tris-Base
57.1 ml Glacial acetic acid
100 ml 0.5 M DTA (pH 8.0)
Adjust volume to 1 L using distilled H
2
O
TB (Tris-Borate- DTA) Buffer
0.5 x working solution*: 45 mM Tris-Borate, 1 mM DTA
5 x stock solution (1 L)**: 54 g Tris-Base
27.5 g Boric acid
20 ml 0.5 M DTA (pH 8.0)
Adjust volume to 1 L using distilled H
2
O
* 0.5 x TB is sufficient for agarose gel electrophoresis. For vertical electrophoresis in polyacrylamide gels,
1 x TB is often applied due to the comparatively smaller buffer reservoirs of vertical electrophoresis chambers.
** 5 x TB stock solutions tend to precipitate during long storage periods and should get remade. Because of this
property, higher concentrations of TB stock solutions should be avoided.

Instruction Manual PerfectBlue
™
Horizontal Maxi Gel Systems
VWR_v0617_ 8
Agarose: Gel volumes and percentage
VWR offers an extensive range of high quality agaroses, for many specific applications (see
'T CHNICAL SUPPORT AND ORD RING INFORMATION').
The required volume of the gel is calculated using the following formula.
gel width (cm) x gel length (cm) x gel thickness (cm) = required volume agarose solution (ml)
The following volumes will result:
PerfectBlue™ Gel size (cm) Gel thickness (cm)
0.25 0.5 0.75 1.0
Maxi S 13 x 25 (W x L) 81 ml 162 ml 243 ml 325 ml
Maxi S Plus 15 x 25 (W x L) 95 ml 190 ml 285 ml 375 ml
Maxi M 20 x 25 (W x L) 125 ml 250 ml 375 ml 500 ml
Maxi M 'Revolution' 20 x 25 (W x L) 125 ml 250 ml 375 ml 500 ml
Maxi L 23 x 40 (W x L) 230 ml 460 ml 690 ml 920 ml
Maxi xW 23 x 25 (W x L) 144 ml 288 ml 432 ml 575 ml
Maxi xW 'Revolution' 23 x 25 (W x L) 144 ml 288 ml 432 ml 575 ml
The optimal range of DNA fragment sizes separated by any electrophoresis experiment is dependent on
the agarose concentration of the gel. The higher the agarose concentration, the better small fragments
are separated from each other and vice versa. However, for the smallest or largest fragment lengths, the
usage of specialized agaroses or polyacrylamide gels should be considered (see table below) since a
3 % agarose solution solidifies rapidly and a 0.3 % agarose gel is very soft and difficult to handle.
Agarose content (w/v) Agarose (g) Buffer (ml) optimal separation range (kb)
0.3 % 0.3 100 5 - 30
0.5 % 0.5 100 1 - 15
0.7 % 0.7 100 0.8 - 10
1.0 % 1.0 100 0.5 - 7
1.2 % 1.2 100 0.3 - 6
1.5 % 1.5 100 0.2 - 4
2.0 % 2.0 100 0.1 - 3
3.0 % 3.0 100 < 0.1
thidium bromide
The gel may be stained during or following the run with a variety of stains for photo documentation. The
most common stain for DNA is ethidium bromide. Because of its capacity to intercalate into double
stranded nucleic acids and alter the conformation of DNA, ethidium bromide is judged to be highly
mutagenic. Therefore appropriate safety measures must be applied.
thidium bromide may be added directly to the gel before pouring it at a concentration of 0.1 to
0.5 µg/ml. However, being positively charged, ethidium bromide will migrate to the cathode during the
electrophoresis leading to uneven staining. Improved results can be obtained by incubating the gel after
the electrophoresis is finished in electrophoresis buffer containing 0.5 µg/ml ethidium bromide for 5 to
20 min. Subsequently the gel should get rinsed in electrophoresis buffer without ethidium bromide for up
to 20 min in order to reduce background signal.