Labnet ENDURO96 E1010-9611 Manuale utente
Lit M00203 May 2012
ENDURO96
HORIZONTAL GEL BOXES
User Manual
E1010-9611
E1010-9611.5
E1010-9621
E1010-9621.5
Lit M00203 May 2012
Contents:-
Page
1) Safety Instructions 1
2) Packing Lists 2
3) Usage Guidance and restrictions 3
4) Care and Maintenance 3
5) Setting Up 4
6) Gel Preparation 5
7) Gel Pouring 6
8) Gel Running 7
9) Gel Staining 8
10) Solutions 9
11) Combs 9
12) Symbols & Conventions 10
13) Equipment Disposal 10
14) Declaration of Conformity 11
15) Warranty 13
1
SAFETY PRECAUTION
WHEN USED CORRECTLY, THESE UNITS POSE NO HEALTH RISK.
HOWEVER, THESE UNITS CAN DELIVER DANGEROUS LEVELS OF
ELECTRICITY AND ARE TO BE OPERATED ONLY BY QUALIFIED
PERSONNEL FOLLOWING THE GUIDELINES LAID OUT IN THIS
INSTRUCTION MANUAL.
ANYONE INTENDING TO USE THIS EQUIPMENT SHOULD READ THE
COMPLETE MANUAL THOROUGHLY.
THE UNIT MUST NEVER BE USED WITHOUT THE SAFETY LID
CORRECTLY IN POSITION.
THE UNIT SHOULD NOT BE USED IF THERE IS ANY SIGN OF DAMAGE TO
THE EXTERNAL TANK OR LID.
THESE UNITS COMPLY WITH THE STATUTORY CE SAFETY DIRECTIVES:
73/23/EEC: LOW VOLTAGE DIRECTIVE: IEC 1010-1:1990 plus
AMENDMENT 1:1992
EN 61010-1:1993/BS EN 61010-1:1993
2
PACKING LISTS:
E1010-9611, E1010-9611.5, E1010-9621, E1010-9621.5
Units include tank, lid and electrodes and include the following accessories:-
Cat no.
Tray
Tray
Dams
Combs and
comb block
Cables
E1010-9611
E1010-96T 10 x
12cm (W x L)
E1010-CD
Pack of 2
E1010-9611-CB
E1107-EP
E1010-9611.5
E1010-96T 10 x
12cm (W x L)
E1010-CD
Pack of 2
E1010-9611.5-CB
E1107-EP
E1010-9621
E1010-96T 10 x
12cm (W x L)
E1010-CD
Pack of 2
E1010-9621-CB
E1107-EP
E1010-9621.5
E1010-96T 10 x
12cm (W x L)
E1010-CD
Pack of 2
E1010-9621.5-CB
E1107-EP
The packing lists should be referred to as soon as the units are received
to ensure that all components have been included. The unit should be
checked for damage when received.
Please contact your supplier if there are any problems or missing
items.
3
Usage Guidance and restrictions:
• Maximum altitude 2,000m.
• Temperature range between 4°C and 65°C.
• Maximum relative humidity 80% for temperatures up to 31OC decreasing
linearly to 50%relative humidity at 40OC.
• Not for outdoor Use.
This apparatus is rated POLLUTION DEGREE 2 in accordance with IEC 664.
POLLUTION DEGREE 2, states that: “Normally only non-conductive
pollution occurs.
Occasionally, however, a temporary conductivity caused by condensation
must be expected”.
Care and Maintenance:-
Cleaning Horizontal Units
Units are best cleaned using warm water and a mild detergent. Water at
temperatures above 600 C can cause damage to the unit and
components.
The tank should be thoroughly rinsed with warm water or distilled water to
prevent build up of salts but care should be taken not to damage the
enclosed electrode and vigorous cleaning is not necessary or advised.
Air drying is preferably before use.
The units should only be cleaned with the following:-
Warm water with a mild concentration of soap or other mild detergent.
Compatible detergents include dishwashing liquid, Hexane and Aliphatic
hydrocarbons
The units should not be left to in detergents for more than 30 minutes.
The units should never come into contact with the following cleaning
agents, these will cause irreversible and accumulative damage:-
4
Acetone, Phenol, Chloroform, Carbon tetrachloride, Methanol, Ethanol,
Isopropyl alcohol
Alkalis.
Rnase Decontamination
This can be performed using the following protocol:-
Clean the units with a mild detergent as described above.
Wash with 3% hydrogen peroxide (H2O2) for 10 minutes.
Rinsed with 0.1% DEPC- (diethyl pyrocarbonate) treated distilled water,
Caution: DEPC is a suspected carcinogen. Always take the necessary
precautions when using.
RNaseZAP™ (Ambion) can also be used. Please consult the instructions for
use with acrylic gel tanks.
Setting up the Horizontal Gel Tanks:-
Instructions for fitting Electrode Cables
1. Note the position of the lid on the unit. This shows the correct polarity
and the correct orientation of the cables, black is negative and red
positive.
2. Remove the lid from the unit. Note if the lid is not removed, fitting the
cables may result in un-tightening of the gold plug and damage to the
electrode.
3. Screw the cables into the tapped holes as fully as possible so that
there is no gap between the lid and the leading edge of the cable
fitting.
4. Refit the lid.
5
Instructions for fitting Loading Guides
These can be fitted to enhance visibility of the wells if desired. They
can be fitted to the white vinyl platform sheet or to the unit itself.
1. Seat the tray in the unit and note the position of the comb grooves.
The samples run black to red but the trays can be used frontward or
backwards so ensure that the comb grooves closest to the black
electrode are marked.
2. Remove the tray.
3. Peel the back off of the loading guide and carefully apply the loading
guide directly to the gel platform.
The unit is now ready to be used.
Gel Preparation:-
1. Table 1 below shows the volume of agarose solution required to make the
desired agarose gel for each unit tray size. For a standard 0.7% agarose gel,
add 0.7grammes of agarose to 100 ml of 1x TAE or TBE solution. The same
1 x solution should be used in the tank buffer solution.
E1010-9611 & E1010-9611.5
Gel volume for a 5mm thick gel
60ml
2. Add the agarose powder to a conical flask.
3. Add the appropriate amount of 1x TAE or TBE solution from the table
above. To prevent evaporation during the dissolving steps below, the conical
flask should be covered with parafilm.
4. Dissolve the agarose powder by heating the agarose either on a magnetic
hot plate with stirring bar or in a microwave oven. If using the microwave
method, the microwave should be set at around a 400 watt or medium
setting and the flask swirled every minute. The solution should be heated
6
until all crystals are dissolved. This is best viewed against a light
background. Crystals appear as translucent crystals. These will interfere
with sample migration if not completely dissolved.
The gel must be cooled to between 50°C and 60°C degrees before
pouring.
Gel Pouring:-
The range of units allows three different methods of gel casting:-
Casting Dams
Flexicaster
Traditional Tape
Using trays with Casting Dams:
1. Fit the casting dams over each end of the tray and place onto a level
surface. The dams should be fitted so that there is no gap between the
sides of the tray and the groove in the dams. This will ensure that
there is no possibility of gel leakage.
2. Place the comb(s) in the grooves. Each tray has more than one comb
grove so that multiple combs can be used. Using multiple combs
increases sample number available per gel but decreases run length
and care must be taken to ensure that samples from the first wells do
not migrate into the lanes of the second comb wells.
3. Pour in the agarose carefully so as not to generate bubbles. Any
bubbles that do occur can be smoothed to the edge of the gel and
dispersed using a pipette tip.
4. Allow the agarose to set, ensuring that the gel remains undisturbed.
5. Carefully remove the gel casting gates and comb and transfer the gel
including tray to the main tank.
7
Using Traditional tape method:-
1. Autoclave or plastic backed general tape should be used. A length
5cm longer than the width of each end of the tray should be cut. One
length should be placed over one end of the tray and stuck 1cm in
from the tray edge. This should then be folded and the edges sealed
securely. Repeat for the other end and place onto a level surface for
gel pouring.
2. Place the comb(s) in the grooves. Each tray has more than one comb
grove so that multiple combs can be used. Using multiple combs
increases sample number available per gel but decreases run length
and care must be taken to ensure that samples from the first wells do
not migrate into the lanes of the second comb wells.
3. Pour in the agarose carefully so as not to generate bubbles. Any
bubbles that do occur can be smoothed to the edge of the gel and
dispersed using a pipette tip.
4. Allow the agarose to set, ensuring that the gel remains undisturbed.
5. Carefully remove the gel casting gates and comb and transfer the gel
including tray to the main tank.
Running the Gel:-
1. Mix the sample to be loaded with sample buffer –see solutions for
common sample buffers. Usually 3ul of sample buffer is adequate but less
may be used with sample volumes of less than 10ul.
2. Fill the unit with enough buffer so that it will just cover the gels when
they are immersed. This will give the fastest resolution times. For enhanced
quality of resolution of sample, fill the unit to 5mm above the gel.
3. On the bench surface, load a small amount of running buffer to flood the
wells. Load the samples into the wells using a pipette. Multi-channel
pipettes can be used forloading samples into wells formed by MC compatible
combs, see listing in accessories for identification of these.
8
4. Once loaded, gently immerse the gels within the buffer, stacking them
carefully on top of each other. Replace lid and connect unit to the power
supply using cables.
5. Typically gels are run at between 90 and 150 volts. However, maximum
voltages are indicated on the serial badge of each unit. It should be noted
that higher voltages generally give faster but poorer quality sample
resolution.
Gel Staining and Viewing:-
The Multi Sub trays and the Mini Fast unit allow staining to be performed
without removing the gel from the tray if this is preferred.
1. Transfer the gel to a vessel containing the appropriate volume of 0.5
µg/ml ethidium bromide stain for 15–30 minutes, see solutions for stock
stain concentration and adjust to the volume used accordingly. The entire
gel should be covered.
NOTE:- Ethidium bromide is a suspected carcinogen and the necessary
safety precautions should be undertaken.
2. De-stain the gel for 10–30 minutes in distilled water again ensuring the
gel is completely immersed.
3. Rinse the gel twice for a couple of seconds with distilled water.
4. Transfer the gel to a UV Transilluminator.
5. The samples will often appear as brighter, clearer bands when
photographed or viewed using a gel documentation system. However if the
gel bands are too faint then the staining procedure should be adjusted so
that there is less de-staining. If there is too much background then the
staining procedure should be adjusted so that there is more de-staining.
References
1. Sambrook, Fritsch, and Maniatis, Molecular Cloning A Laboratory
Manual, Second Edition,
Cold Spring Harbor Laboratory Press, 1989.
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