Bio-Rad Mini-PROTEAN Tetra Cell User Manual

Mini-PROTEAN
Tetra Cell

Instruction Manual

Catalog Numbers

165-8000 165-8004
165-8001 165-8005
165-8002 165-8006
165-8003 165-8007

®

Table of Contents

Section 1 General Information 1

1.1 Introduction 1

1.2 Components 1

1.3 Specifications 4

1.4 Safety 5

Section 2 Setup and Basic Operation 6

2.1 Gel Cassette Preparation 6

2.2 Electophoresis Module Assembly and
Sample Loading 9

Section 3 Separation Theory and Optimization 15

3.1 Introduction 15

3.2 SDS-PAGE (Laemmli) Buffer System 16

3.3 Native PAGE 17

Section 4 Reagent Preparation and Stock Solutions 19

4.1 Volumes Required per Gel 19

4.2 SDS-PAGE (Laemmli) Buffer System 19

4.3 Discontinuous Native PAGE (Ornstein-Davis) 22

4.4 Continuous Native PAGE 24

Section 5 References 27

Section 6 Maintenance 27

Section 7 Troubleshooting Guide 28

Section 8 Product Information and Accessories 31

Section 9 Warranty Information 35

Section 1
General Information

1.1 Introduction

The Mini-PROTEAN® Tetra cell runs both handcast gels and Ready
Gel® precast gels interchangeably. The Mini-PROTEAN Tetra
system includes a casting stand and glass plates with permanently
bonded gel spacers that simplify handcasting and eliminate leaking
during casting. The cell can run one or four gels, and the mini
tank is compatible with other Bio-Rad electrode modules for tank
blotting, 2-D electrophoresis, and electroelution.

1.2 Components

To get the best performance from your Mini-PROTEAN Tetra
Cell, familiarize yourself with the components by assembling and
disassembling the cell before using it (refer to Figures 1 and 2).

Spacer Plate The spacer plate is the taller glass plate

Short Plate The short plate is the shorter, flat glass

Casting Frame The casting frame, when placed on the

Gel Cassette Assembly One casting frame, a spacer plate, and a

Casting Stand The casting stand secures the gel cassette

Gel Cassette Sandwich A spacer plate and short plate with

Buffer Dam The molded, one-piece buffer dam is used

with permanently bonded gel spacers.
Spacer plates are available in 0.75 mm, 1.0
mm, and 1.5 mm thicknesses, which are
marked directly on each spacer plate.

plate that combines with the spacer plate
to form the gel cassette sandwich.

benchtop, evenly aligns and secures the
spacer plate and the short plate together
to form the gel cassette sandwich prior to
casting.

short plate form one gel cassette assembly.

assembly during gel casting. It contains
pressure levers that seal the gel cassette
assembly against the casting gaskets.

polymerized gel form a gel sandwich.

when running only one or three gels.

1

Electrode Assembly The electrode assembly holds the gel

Companion Assembly The companion assembly allows you to run

Mini Tank and Lid The mini tank and lid combine to fully

sandwich. It houses the sealing gasket,
the upper and lower electrodes, and the
connecting banana plugs. The anode
(lower electrode) banana plug is identified
with a red marker and the cathode (upper
electrode) banana plug with a black marker.

gels 3 and 4. It holds the gel sandwich and
houses the sealing gasket.

enclose the inner chamber during
electrophoresis. The lid cannot be removed
without disrupting the electrical circuit.
The mini tank and lid are also compatible
with other Bio-Rad electrode modules
for blotting, first-dimension of 2-D
electrophoresis, and electroelution.

2

Lid

Electrode
assembly

Banana plug jacks

Gel cassette

Notch on

U-shaped gasket

Mini tank

Fig. 1. Assembling the Mini-PROTEAN Tetra Cell.

Fig. 2. Assembling the Mini-PROTEAN Tetra Cell casting frame and
casting stand

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1.3 Specifications

Casting Stand* Polycarbonate

Pin, retaining ring, and spring Stainless steel

Casting Frames* Polysulfone

Gray gaskets Thermoplastic rubber (gray)

Electrode Assembly Glass-filled polybutylene

terephthalate

Electrodes Platinum wire, 0.010 inches diameter

Gasket, electrode inner core Silicone rubber (green)

Mini Tank and Lid Polycarbonate

Sample Loading Guides** Delrin

Combs* Polycarbonate

Overall Size (W x L x H, cm) 12 x 16 x 18

Precast Gel Compatibility Ready Gel and Mini-PROTEAN

precast gels (for more information,
go to www.bio-rad.com/mpgels)

Voltage Limit 600 V DC and 500 W

Shipping Weight 2.0 kg

Maximum Sample Volume per Well

# Wells Well Width 0.75 mm 1.0 mm 1.5 mm

5 12.7 mm 20 µl 105 µl 160 µl

9 5.08 mm 33 µl 44 µl 66 µl

10 5.08 mm 33 µl 44 µl 66 µl

15 3.35 mm 20 µl 26 µl 40 µl

IPG 6.2 mm – 420 µl 730 µl

Prep/2-D

Reference well 3.1 mm 13 µl 17 µl 30 µl

Sample well 71.1 mm 310 µl 400 µl 680 µl

* US patent No. 6,162,342
** US patent No. 5,656,145

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Chemical Compatibility

Mini-PROTEAN Tetra cell components are not compatible with
acetone or ethanol. Use of organic solvents voids all warranties.
Call 1-800-4-BIORAD (US) or your local Bio-Rad representative for
technical information regarding chemical compatibility of the
Mini-PROTEAN Tetra cell with various laboratory reagents.
The Mini-PROTEAN are not compatible with repeated exposure to
100% TEMED. Rubbing the combs with TEMED prior to casting
will destroy the structural integrity of the combs over time.

1.4 Safety

Power to the Mini-PROTEAN Tetra cell is supplied by an external
DC voltage power supply (not included). The output of this power
supply must be isolated from external ground to ensure that the
DC voltage output floats with respect to ground. All Bio-Rad power
supplies meet this important safety requirement. Regardless of the
power supply used, the maximum specified operating parameters
for the Mini-PROTEAN Tetra cell are as follows

• 600 V DC maximum voltage limit

• 500 W maximum power limit

• 40°C maximum ambient temperature limit

The current to the cell enters the unit through the lid assembly,
which provides a safety interlock to the user. The current to the
cell is broken when the lid is removed. Always turn off the power
supply before removing the lid. Do not attempt to use the cell

without the safety lid.
Important: This Bio-Rad product is designed and certified to meet

IEC61010-1 and EN61010-1* safety standards. Certified products
are safe to use when operated in accordance with the instruction
manual. This instrument should not be modified or altered in any
way. Alteration of this instrument will

• Void the warranty

• Void the IEC61010-1 and EN61010-1 certifications, and

• Create a potential safety hazard

Bio-Rad is not responsible for any injury or damage caused by
use of this instrument for purposes other than those for which it is
intended or by modifications of the instrument not performed by
Bio-Rad or an authorized agent.

*IEC61010-1 and EN61010-1 are internationally accepted electrical safety
standards for laboratory instruments.

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Section 2
Setup and Basic Operation

2.1 Gel Cassette Preparation

Handcast Gels

1. Glass Cassette and Casting Stand Assembly

Note: All glass plates should be clean and dry.

a. Place the casting frame upright with the pressure cams in

the open position and facing forward on a flat surface.

b. Select a spacer plate of the desired gel thickness and place
a short plate on top of it (see Figure 3a).

c. Orient the spacer plate so that the labeling is up. Slide the
two glass plates into the casting frame, keeping the short
plate facing the front of the frame (side with pressure cams)
(see Figure 3b).

Note: Ensure that both plates are flush on a level surface and
that the labels on the spacer plate are oriented correctly. Leaking
may occur if the plates are misaligned or oriented incorrectly.

d. When the glass plates are in place, engage the pressure
cams to secure the glass cassette sandwich in the casting
frame (see Figure 3c). Check that both plates are flush at
the bottom.

e. Place the casting frame into the casting stand by positioning
the casting frame (with the locked pressure cams facing out)
onto the casting gasket while engaging the spring-loaded
lever of the casting stand onto the spacer plate (see Figure
3d).

Note: The gray casting stand gaskets must be clean and dry.
The casting stand gaskets are made of a special thermoplastic
material that swells when soaked in water, so we recommend
that you do not soak the gaskets for prolonged periods prior to
casting. If the gaskets do get accidentally soaked and display
swelling and/or deformation, just allow them to air dry and they
will regain their original shape, size and performance.

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f. Repeat steps a–e for additional gels.

3a 3b

3c 3d

Fig. 3. Assembling the Mini-PROTEAN casting stand and frame.

2.0 Gel Casting

a. Discontinuous Polyacrylamide Gels

i. Place a comb completely into the assembled gel cassette.
Mark the glass plate 1 cm below the comb teeth. This is the
level to which the resolving gel is poured. Remove the comb.

ii. Prepare the resolving gel monomer solution by combining all
reagents except APS and TEMED. (Refer to section 4 for
gel formulations.) Degas the solution under vacuum for at
least 15 min. Do not use a sink water aspirator.

iii. Add APS and TEMED to the degassed monomer solution
and pour to the mark using a glass or disposable plastic
pipet. Pour the solution smoothly to prevent it from mixing
with air.

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iv. Immediately overlay the monomer solution with water or
t-amyl alcohol.

Note: If water is used, add it slowly and evenly to prevent mixing.
Do not overlay with butanol or isobutanol.

v. Allow the gel to polymerize for 45 min to 1 hr. Rinse
the gel surface completely with distilled water. Do not leave
the alcohol overlay on the gel for more than 1 hr because
it will dehydrate the top of the gel.

Note: At this point the resolving gel can be stored at room
temperature overnight. Add 5 ml of 1:4 dilution of 1.5 M
Tris-HCl, pH 8.8 buffer (for Laemmli system) to the resolving gel
to keep it hydrated. If using another buffer system, add 5
ml 1x resolving gel buffer for storage.

vi. Prepare the stacking gel monomer solution. Combine all
reagents except APS and TEMED. Degas under vacuum for
at least 15 min.

vii. Dry the top of the resolving gel with filter paper before
pouring the stacking gel.

viii. Add APS and TEMED to the degassed stacking gel
monomer solution and pour the solution between the glass
plates. Continue to pour until the top of the short plate is
reached.

b. Continuous Polyacrylamide Gels

i. Prepare the monomer solution by combining all reagents
except the APS and the TEMED. Degas under vacuum for
15 min (refer to section 4 for gel formulations).

ii. Add APS and TEMED to the degassed monomer solution
and pour the solution between the glass plates. Continue to
pour until the top of the short plate is reached.

iii. Insert the desired comb between the spacers starting at the
top of the spacer plate, making sure that the tabs at the
ends of each comb are guided between the spacers. Seat
the comb in the gel cassette by aligning the comb ridge with
the top of the short plate.

iv. Rinse the casting frame(s) and stand with distilled,
deionized water after use.

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®

Ready Gel

Precast Gels

1. Ready Gel Cassette Preparation

Note: The Mini-PROTEAN Tetra cell is guaranteed for use with

Bio-Rad’s Ready Gel and Mini-PROTEAN® precast gels. For
more information, go to www.bio-rad.com/mpgels.

a. Remove the Ready Gel from the storage pouch.

b. Gently remove the comb and rinse the wells thoroughly with
distilled water or running buffer.

c. Cut along the dotted line at the bottom of the Ready Gel
cassette with a razor blade.

d. Pull the clear tape at the bottom of the Ready Gel cassette
to expose the bottom edge of the gel.

e. Repeat for second Ready Gel.

Note: If only one or three gels are to be run, use the mini cell
buffer dam.

2.2 Electrophoresis Module Assembly and Sample Loading

Required materials:

• Clean and dry Mini-PROTEAN Tetra cell tank

• Electrophoresis module (electrode assembly module only for

1 or 2 gels; for 3 or 4 gels also use the companion running
module)

• Running buffer (700 ml for 2 gels; 1000 ml for 4 gels)

• Ready Gel precast gels or hand-cast gels

• PowerPac™ Basic power supply

1. Assembly

Note: When running 2 gels only, use the electrode assembly (the

one with the banana plugs), not the companion running module
(the one without the banana plugs). When running 4 gels, both
the electrode assembly and the companion running module
must be used, for a total of 4 gels (2 gels per assembly).

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a. Set the clamping frame to the open position on a clean flat
surface (see Figure 4a).

b. Place the first gel sandwich or gel cassette (with the short
plate facing inward) onto the gel supports; gel supports are
molded into the bottom of the clamping frame assembly;
there are two supports in each side of the assembly. Note
that the gel will now rest at a 30° angle, tilting away from the
center of the clamping frame.Please use caution when

placing the first gel, making sure that the clamping frame
remains balanced and does not tip over. Now, place the

second gel on the other side of the clamping frame, again by
resting the gel onto the supports. At this point there will be
two gels resting at an angle, one on either side of the
clamping frame, tilting away from the center of the frame (see
Figure 4b).

Note: It is critical that gel cassettes are placed into the clamping
frame with the short plate facing inward. Also, the clamping
frame requires 2 gels to create a functioning assembly. If an odd
number of gels (1 or 3) is being run, you must use the buffer dam
(see Figure 4b).

c. Using one hand, gently pull both gels towards each other,
making sure that they rest firmly and squarely against the
green gaskets that are built into the clamping frame; make
certain that the short plates sit just below the notch at the
top of the green gasket.

d. While gently squeezing the gel sandwiches or cassettes
against the green gaskets with one hand (keeping constant
pressure and both gels firmly held in place), slide the green
arms of the clamping frame over the gels, locking them
into place. Alternatively, you may choose to pickup the
entire assembly with both hands, making sure that the
gels do not shift, and simultaneously sliding both arms of
the clamping frame into place (see Figure 4c).

The arms of the clamping frame push the short plates of each
gel cassette up against the notch in the green gasket, creating a
leak-proof seal (check again to make certain that the short plates
sit just below the notch at the top of the green gasket). At this
point, the sample wells can be washedout with running buffer,
and sample can be loaded (Figure 4d).

Note: If running more than 2 gels, repeat steps 1a–d with the
companion running module

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