Cleaver scientific Multisub midi96 User manual

multiSUB Midi96

Horizontal System

15/11/2018 Page 1

multiSUB®MIDI96 High

throughput horizontal

units

Instruction Manual

Catalogue Numbers

MSMIDI96

MSMIDI961.5

MSMIDI96/2M

MSMIDI96/1.5/2M

MSMIDI96ST

MSMIDI96ST1.5

MSMIDI96ST/2M

MSMIDI96ST/1.5/2M

Record the following for your records:

Model _____________________

Catalogue No. _____________________

Date of Delivery _____________________

Warranty Period _____________________

Serial No. _____________________

Invoice No. _____________________

Purchase Order No. _____________________

15/11/2018 Page 2

Contents

Instruction Manual 1

Catalogue Numbers 1

Contents 2

Safety Information 3

Packing List 4

Specifications 5

Operating Instructions 6

Usage Guidance and restrictions 6

Setting up the Horizontal Gel Tanks 6

Fitting Electrodes 6

Fitting Loading Guides 6

Gel Preparation 7

Gel Pouring 8

Casting Dams 8

Flexicaster 9

Tape 10

Running the Gel 10

Gel Staining and Viewing 11

Solutions 11

References 12

Troubleshooting 13

Care and Maintenance 14

Cleaning Horizontal Units 14

RNAse Decontamination 14

Ordering information 15

Combs 15

Related Products 16

Warranty 17

15/11/2018 Page 3

Safety Information

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 following European directives:

2006/95/CE Low Voltage Directive and 2014/30/UE (official Title 2004/108/EC)

EMC Electromagnetic Compatibility

By virtue of the following harmonised standards:

BS EN IEC 61010-1: 2010 Safety Testing of Lab Equipment

BS EN IEC 61326-1:2013 EMC Electro Magnetic Compatibility

15/11/2018 Page 4

Packing List

Each multiSUB unit includes a tank, wired electrodes, lid and the following

items:

Catalogue

No.

Tray Tray Dams Combs and

comb block

Loading Guides Cabl

MSMIDI96 MS10-UV96 10 x

12cm (W x L)

MS10-UVDAM,

Pack of 2

MSMIDI96-8-1-

MS10-LG – Strips,

MS10-WP – Platform

CSL-

CAB

MSMIDI961.

MS10-UV96 10 x

12cm (W x L)

MS10-UVDAM,

Pack of 2

MSMIDI96-8-

1.5-CB

MS10-LG – Strips,

MS10-WP – Platform

CSL-

CAB

MSMIDI96/

MS10-UV96 10 x

12cm (W x L)

MS10-UVDAM,

Pack of 2

MSMIDI96-8-

1/2M-CB

MS10-LG – Strips,

MS10-WP – Platform

CSL-

CAB

MSMIDI96/

1.5/2M

MS10-UV96 10 x

12cm (W x L)

MS10-UVDAM,

Pack of 2

MSMIDI96-8-

1.5/2M-CB

MS10-LG – Strips,

MS10-WP – Platform

CSL-

CAB

MSMIDI96S

MS10-UV96ST 10 x

24cm (W x L)

MS10-UVDAM,

Pack of 2

MSMIDI96ST-8-

1-CB

MS10-LG – Strips,

MS10-WP – Platform

CSL-

CAB

MSMIDI96S

T1.5

MS10-UV96ST 10 x

24cm (W x L)

MS10-UVDAM,

Pack of 2

MSMIDI96ST-8-

1.5-CB

MS10-LG – Strips,

MS10-WP – Platform

CSL-

CAB

MSMIDI96S

T/2M

MS10-UV96ST 10 x

24cm (W x L)

MS10-UVDAM,

Pack of 2

MSMIDI96ST-8-

1/2M-CB

MS10-LG – Strips,

MS10-WP – Platform

CSL-

CAB

MSMIDI96S

T/1.5/2M

MS10-UV96ST 10 x

24cm (W x L)

MS10-UVDAM,

Pack of 2

MSMIDI96ST-8-

1.5/2M-CB

MS10-LG – Strips,

MS10-WP – Platform

CSL-

CAB

Packing List Checked by: ________________________

Date: ________________________

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.

Cleaver Scientific is liable for all missing or damaged parts / accessories within

7 days after customers have received this instrument package. Please contact

Cleaver Scientific immediately regarding this issue. If no response within such

period is received from the customer, Cleaver Scientific will no longer be liable

for replacement/damaged parts.

Please contact your supplier if there are any problems or missing items.

15/11/2018 Page 5

Specifications

1Power supply cables 1500V rated with retractable 4mm connectors

2 Safety lid and viewing pane High impact moulded acrylic construction

3 Height adjustable combs Acrylic

4 UV transparent gel tray UV transparent above 300nm, Moulded acrylic

5 Comb Slots Allow multiple combs per tray

6 Casting dams Natural rubber

7 Colour-coded electrodes Moulded acrylic, 99.99% platinum wire, gold plated

4mm plugs

8 Gel platform Notched to fix gel position

9 Safety lid thumb locators For safe removal of lid

10 Moulded buffer tank High Impact moulded acrylic construction for chemical

compatibility and shock resistance

15/11/2018 Page 6

Operating Instructions

Further information (including videos) regarding setting up and running the

multiSUB®units can be found at www.cleaverscientific.com

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”.

Setting up the Horizontal Gel Tanks

Fitting Electrodes

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.

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.

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.

Refit the lid.

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.

15/11/2018 Page 7

2. Remove the tray.

3. Peel the back off the loading guide and carefully apply the loading

guide directly to the gel platform.

The unit is now ready to be used.

Gel Preparation

The Table 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 0.7 grams of agarose to 100 ml of 1x TAE or TBE solution. The same 1X

solution should be used in the tank buffer solution.

Gel volume for a 5mm thick gel

Multi Sub Midi 96 Multi Sub Midi 96 Stretch

60ml 120ml

1. Add the agarose powder to a conical flask.

2. 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.

3. 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 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.

15/11/2018 Page 8

Gel Pouring

The multiSUB®range of units allows three different methods of gel casting:

1. Casting Dams

2. Flexicaster

3. Traditional Tape

Casting Dams

1. To fit the casting dams, place one casting dam on the bench with the

groove facing upwards (1). Push the edge of the tray down firmly into

the groove (2). Repeat this for the other side (3). 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.

(1) (2) (3)

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Flexicaster

1. Level the Flexicaster base by adjusting the feet so that the bubble is

exactly central.

2. Insert the desired length tray into the Flexicaster such that one end of

the tray is pushed up and seals against the silicone mat of the

permanent end of the Flexicaster.

3. Position the movable end of the Flexicaster so that the silicone mat is

pushed against the other end of the tray.

4. Turn the cam so that the silicone mat tightly seals against the side of the

tray. 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.

5. Allow the agarose to set, ensuring that the gel remains undisturbed.

6. Carefully remove the gel casting gates and comb and transfer the gel

including tray to the main tank.

MS7-FC and MS20-FC Flexicasters

15/11/2018 Page 10

Tape

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 m1cm 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 buffer until the gel is just flooded with buffer. This will give

the fastest resolution times. For enhanced quality of resolution of

sample, fill the unit to 5mm above the gel.

3. Load the samples into the wells using pipettes. Multi-channel pipettes

can be used for loading samples with MC compatible combs, see

listing in accessories for identification of these.

4. Carefully place the lid on the tank and connect to a power supply.

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.

15/11/2018 Page 11

Gel Staining and Viewing

The Multi Sub trays 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 taken.

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.

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.

Solutions

0.5M EDTA stock (500mL) dissolve in 400 ml distilled water:

93.05g EDTA disodium salt

Fill to 500 ml litre final volume with distilled water

50X TAE stock (1L) dissolve in 750 ml distilled water:

242 g tris base (FW = 121)

57.1 ml glacial acetic acid

100 ml 0.5 M EDTA (pH 8.0).

Fill to 1 litre final volume with distilled water

15/11/2018 Page 12

10X TBE stock (1L) dissolve in 750 ml distilled water:

108 g tris base (FW = 121)

55 g boric acid (FW = 61.8)

40 ml 0.5 M EDTA (pH 8.0)

Fill to 1 litre final volume with distilled water

Loading Dye

10x sample buffer stock consists of 50% glycerol, 0.25% bromophenol blue,

and 0.25% xylene cyanole FF in 1x TAE buffer. Only 1–10 ml of the 10x loading

dye should be prepared.

Ethidium Bromide Solution

Add 10 mg of Ethidium Bromide to 1 ml distilled water.

References

1. Sambrook, Fritsch, and Maniatis, Molecular Cloning A Laboratory

Manual, Second Edition, Cold Spring Harbor Laboratory Press, 1989.

2. Current Protocols in Molecular Biology, Greene Publishing Associates

and Wiley-Interscience, 1989.

15/11/2018 Page 13

Troubleshooting

Problem Cause Solution

Bands sharp but not enough

bands seen

Gel agarose percentage too

high

Incomplete digestion

Decrease agarose percentage.

Review enzyme activity, digest

further.

Band smearing and streaking

Agarose has improper

endosmosis

Salt concentration in sample too

high

Excessive power and heating

Sample spilled out of well

Incomplete digestion, nuclease

contamination, bad enzyme

Sample wells cast through the

gel. Sample leaks along bottom

of running surface

Consult Cleaver Scientific about

agarose.

Reduce salt concentration to

≤0.1M.

Reduce voltage. See

electrophoresis instructions.

Apply sample carefully. Increase

gel thickness for large sample

volumes. Adjust comb height.

Heat sample. Review enzyme

activity. Digest sample further.

Comb should be placed to 1 to 2

mm above the base of the

running surface.

Curved line or distortion of bands Bubbles in sample wells Remove bubbles prior to

electrophoresis.

Curved bands, smiles Sample overload Reduce load.

Differential relative mobilities

Sample spilled out of wells

Unit not levelled

Samples should have proper

density. Apply carefully.

Level unit. Use a steady work

bench.

Gels crack

Too high voltage gradient,

especially with low melting

temperature agarose or low gel

strength gels

Reduce voltage. Run gel at lower

temperature.

High MW bands sharp; low MW

bands smeared Gel agarose percentage too low Increase agarose percentage.

Switch to polyacrylamide.

Ragged bands

Sample density incorrect

Sample well deformed

Excessive power or heating

See sample application

instructions.

Carefully remove comb,

especially from soft gels. Make

sure gel has solidified.

Cooling soft gels aids in comb

removal.

Reduce voltage. See

electrophoresis instructions.

Slanted lanes (bands) Gel not fully solidified

Comb warped or at an angle

Gel to solidify for at least 30-

45min.

Check alignment of comb.

15/11/2018 Page 14

Care and Maintenance

Cleaning Horizontal Units

Units are best cleaned using warm water and a mild detergent. Water at

temperatures above 60°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:

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 pyro carbonate) 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.

15/11/2018 Page 15

Ordering information

Combs

Catalogue No. Description

MSMIDI96-8-1-CB MSMIDI 96 Comb 8 sample MC + 1 Marker, 1mm thick

COMB BLOCK

MSMIDI96-8-1.5-CB MSMIDI 96 Comb 8 sample MC + 1 Marker, 1.5mm thick

COMB BLOCK

MSMIDI96-8-1/2M-CB MSMIDI 96 Comb 8 sample MC + 2 Marker, 1mm thick

COMB BLOCK

MSMIDI96-8-1.5/2M-CB MSMIDI 96 Comb 8 sample MC + 2 Marker, 1.5mm thick

COMB BLOCK

MSMIDI96-8-1 MSMIDI 96 Comb 8 sample MC, 1mm thick. one marker

lane.

MSMIDI96-8-1.5 MSMIDI 96 Comb 8 sample MC, 1.5mm thick. one marker

lane.

MSMIDI96-8-1/2M MSMIDI 96 Comb 8 sample MC, 1mm thick. two marker

lanes.

MSMIDI96-8-1.5/2M MSMIDI 96 Comb 8 sample MC, 1.5mm thick. two marker

lanes.

MSMIDI96ST-8-1-CB MSMIDI 96 STRETCH Comb 8 sample MC + 1 Marker, 1mm

thick COMB BLOCK

MSMIDI96ST-8-1.5-CB MSMIDI 96 STRETCH Comb 8 sample MC + 1 Marker, 1.5mm

thick COMB BLOCK

MSMIDI96ST-8-1/2M-CB MSMIDI 96 STRETCH Comb 8 sample MC + 2 Marker, 1mm

thick COMB BLOCK

MSMIDI96ST-8-1.5/2M-

MSMIDI 96 STRETCH Comb 8 sample MC + 2 Marker, 1.5mm

thick COMB BLOCK

MSMIDI96-8-1 MSMIDI 96 Comb 8 sample MC, 1mm thick. one marker

lane.

MSMIDI96-8-1.5 MSMIDI 96 Comb 8 sample MC, 1.5mm thick. one marker

lane.

MSMIDI96-8-1/2M MSMIDI 96 Comb 8 sample MC, 1mm thick. two marker

lanes.

MSMIDI96-8-1.5/2M MSMIDI 96 Comb 8 sample MC, 1.5mm thick. two marker

lanes.

15/11/2018 Page 16

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