Bio-Rad Bio-Scale Mini Bio-Gel P-6 Desalting Cartridges User Manual

Bio-Scale™Mini
Bio-Gel®P-6 Desalting
Cartridges, 10 and 50 ml

Instruction Manual

Catalog #
732-5304, 732-5312, 732-5314

Table of Contents

Section 1...Introduction ........................................1

Section 2 Connecting to Bio-Rad’s

Low-Pressure Chromatography

Instruments ......................................24

Section 3 Connecting to Other Liquid

Chromatography Systems..................9

3.1 Connecting to BioLogic
DuoFlow

™

Systems .................9

3.2 HPLC Systems ......................10

3.3 FPLC Systems .......................11

Section 4 Preparing a Cartridge for Use...........12

4.1 Sample Preparation ...............13

Section 5...Desalting and Buffer Exchange.........14

5.1 With a Liquid Chromatography

System ..................................15

Section 6 Technical Assistance ........................16

Section 7 Ordering Information.........................17

Section 8 References .......................................19

Section 1
Introduction

Bio-Scale Mini cartridges have a patent-pending
double-wall design that provides extra durability and
allows easy, reliable runs with aqueous buffers
commonly used for protein purification. The
polypropylene luer fittings and internal sealing
surfaces assure leak-free operation at pressures up
to 45 psi. Bio-Scale Mini cartridges are convenient,
disposable, and supplied ready for use. They are
easy-to-use and prepacked for fast, reproducible
chromatographic separations. Cartridges are
available for a variety of chromatographic techniques,
including desalting, ion exchange and affinity
chromatography. The patent-pending design of
Bio-Scale Mini cartridges offers:

• Supplied ready for use; simply equilibrate the
cartridge in the buffer of choice

1

• Luer fittings for convenient connection to any
chromatography system

Bio-Scale Mini Bio-Gel P-6 cartridges are packed
with Bio-Gel

®

P-6 gel. This gel consists of
hydrophilic spherical polyacrylamide beads
designed for the purification of proteins, nucleic
acids, viruses, plasmids, and other
macromolecules. The Bio-Gel P-6 beads are
designed to provide high-efficiency desalting or
buffer exchange at low backpressure. Detailed
product information is given in Table 1. See
Ordering Information for a listing of the complete
Bio-Scale Mini cartridge product line.

2

Table 1. Bio-Scale Mini Bio-Gel P-6
cartridge specifications.

Size 10 and 50 ml bed volume

Dimensions 10 ml: 80 mm length x 12.6 mm inner

diameter

50 ml: 80 mm length x 28.2 mm inner

diameter

Maximum pressure tolerance 10 ml: 45 psi

50 ml: 40 psi

Recommended flow rates:
at ambient temperature
(18–24°C) 10 ml: 0.5–4.0 ml/min (24–192 cm/hr)

50 ml: 2.5–20.0 ml/min (24–192 cm/hr)

at cold room temperature
(4–8°C) 10 ml: 0.5–2.0 ml/min (24–96 cm/hr)

50 ml: 2.5–10.0 ml/min (24–96 cm/hr)

Fittings Female luer-lock fitting inlet and

male luer fitting oulet

Column material Polypropylene

Frit material Polyethylene (HDPE)

Shipping conditions 20 mM Bis-Tris, pH 6.5 + 0.05% azide

Storage recommendation 20 mM Bis-Tris, pH 6.5 + 0.05% azide

Autoclavability Not autoclavable

3

Section 2
Connecting to Bio-Rad’s
Low-Pressure
Chromatography Instruments

Bio-Scale Mini cartridges are ideal for use with
Bio-Rad's BioLogic

™

LP chromatography system,

Econo

™

gradient pump, the patented* Model EP-1

Econo

™

pump, and all low-pressure chromatography
instruments. Bio-Scale Mini cartridges can be con­veniently connected directly to the system using the
luer fittings on the cartridge.

1. Install 1.6 mm inner diameter (ID) tubing in the

pumphead. Adjust platen pressure screw (on
pumphead) – using a screwdriver or coin, turn
the screw counterclockwise as far as it will go,
then turn clockwise three full turns. Assemble
with fittings and lock rings as shown in Figure 1.

*US Patent 5,135,658

4

(Use orange lock rings and medium size barb
fittings with 1.6 mm tubing.)

Fig. 1. BioLogic LP system setup.

5

Platen pressure

screw

See detail

Lock-ring

Tubing

Luer
fitting

Platen pressure

screw

See detail

23

1

9

456

7 8

0

C .

Alarm

Lock-ring

Tubing

Luer fittimg

2. To apply samples efficiently, install 1.6 mm ID

tubing from the pump to the MV-6 sample
inject valve (if available). If using the MV-6
sample inject valve, turn the knob
counterclockwise as far as it will go so it will
now correspond to the printed diagram on the
valve (see Figure 2).

Fig. 2. Connecting to an MV-6 valve.

6

SAMPLE LOOP

INJECT
PORT

MV-6

BIOLOGIC LP

SYSTEM OR ECONO

GRADIENT PUMP

TO

INJECT

VALVE

FILL

TO

COLUMN

WASTE

SYSTEM OR ECONO

GRADIENT PUMP

BIOLOGIC LP

SAMPLE LOOP

INJECT
PORT

FILL

MV-6

INJECT

TO

VALVE

"INJECT" POSITION"FILL" POSITION

TO

COLUMN

WASTE

3. Connect the inlet of the cartridge to the male

luer fitting on the MV-6 sample inject valve (see
Figure 2). If not using the MV-6 sample inject
valve, connect a barb to male luer fitting on the

1.6 mm ID tubing, then connect to the top of the
female luer on the Bio-Scale Mini cartridge. For
optimum performance, a cartridge should be
mounted vertically with the arrow on the
cartridge pointing downward (see Figure 3).

4. Connect the cartridge outlet to the 1.6 mm ID

tubing leading to the BioLogic LP system optics
module or to the Model EM-1 Econo

™

UV
monitor. It is recommended to use the shortest
length (approximately 10 cm) of 1.6 mm ID
tubing. Connect a barb to female luer to the

1.6 mm ID tubing, then connect to the bottom
of the male luer on the Bio-Scale Mini
cartridge.

7

Fig. 3. Cartridge and fittings.

8

Section 3
Connecting to Other Liquid
Chromatography Systems

Bio-Scale Mini cartridges can be connected to any liquid
chromotography system, provieded that the maximum
pressure limits for the 10 ml (3 bar, 45 psi, or 300 kPa) or
for the 50 ml (2.6 bar, 40 psi, or 260 kPa) cartridges are
not exceeded. It is recommended that the system
pressure limit be set according to the cartridge pressure
limit. Pressures in excess of 3 bar are usually caused by
restrictions in tubing or detector cells downstream from
the cartridge. Bio-Rad offers two fittings kits for easy
connection of a Bio-Scale Mini cartridge to a BioLogic
DuoFlow, HPLC, or FPLC-type system.

3.1 Connecting to a BioLogic DuoFlow
System

The luer to BioLogic system fittings kit (catalog
#732-0113) includes 1/4-28 female to male luer
and 1/4-28 female to female luer to connect one
Bio-Scale Mini cartridge to the BioLogic DuoFlow
system, see Figure 4.

9

Fig. 4. Luer to 1/4-28 adaptor.

3.2 HPLC Systems

The luer to 10-32 adaptor fittings kit (catalog
#732-0112), provides fittings necessary to connect
the Bio-Scale Mini cartridge to nut and ferrule type
fittings found on most HPLC systems. Alternatively,
the cartridge can be connected to HPLC systems
via a low dead-volume 1/16 inch union with a new
piece of stainless steel tubing attached to the union.
Simply slip a short length of the 0.8 mm ID tubing
over 1/16 inch OD stainless-steel tubing to a
distance of 1 cm.

10

3.3 FPLC Systems

The luer to M6 adaptor fittings kit (catalog
#732-0111) provides fittings necessary to connect
the Bio-Scale Mini cartridge to the M6 fittings found
on FPLC or related systems. Alternatively, connection
can be made by using one GE Healthcare Union
luer lock female to M6 female fitting (GE 18-1027-12)
and one female luer lock to M6 male fitting
(Upchurch P-686 or GE 18-1027-62). To prevent
tubing or cartridge failure, do not exceed the
maximum recommended flow rate of the cartridge.

11

Section 4
Preparing a Cartridge
For Use

Bio-Scale Mini Bio-Gel P-6 cartridges contain
20 mM Bis-Tris, pH 6.5 + 0.05% sodium azide as
the storage solution. The fully hydrated support is
ready to use after equilibrating the cartridge in the
buffer of choice. To perform a buffer exchange,
connect the cartridge to a liquid chromatography
system or peristaltic pump and condition it as
instructed below:

1. Set the pump flow rate for the 10 ml cartridge
to 4.0 ml/min (192 cm/hr) for room temperature
use or 2.0 ml/min (96 cm/hr) for cold room
temperature use.

2. Set the pump flow rate for the 50 ml cartridge to

20.0 ml/min (192 cm/hr) for room temperature
use or 10.0 ml/min (96 cm/hr) for cold room
temperature use.

12

3. Equilibrate the cartridge with 2.2 column volumes
of degassed equilibration buffer: 6 minutes at the
room temperature flow rate setting or 12 minutes
at the cold room temperature setting.

4. Reduce the flow rate to that which will be used
in the buffer exchange or desalting cycle.

4.1 Sample Preparation

If the sample contains insoluble material, filter it with
a 0.45 µm filter before applying it to the cartridge.

13

Section 5
Desalting and Buffer
Exchange

Samples can be desalted or the sample buffer
exchanged by connecting the cartridge to a liquid
chromatography system. The void volume for the 10
ml Bio-Scale Mini Bio-Gel P-6 Desalting cartridge is

3.6 ml and for the 50 ml Bio-Scale Mini Bio-Gel P-6

Desalting cartridge is 18 ml. With the 10 ml
cartridge the desalted sample, the sample, or the
sample in its new buffer is contained within the
fraction between 3 ml and 9 ml with the highest
concentration of sample in the 3.6 to 6.0 ml fraction.
With the 50 ml cartridge the desalted sample or the
sample or the sample in its new buffer is contained
within the fraction between 15 ml and 45 ml with
the highest concentration of sample in the 18 to
30 ml fraction.

14

5.1 With a Liquid Chromatography System

1. Set the flow rate to the desired flow.
a. 10 ml cartridge: 0.5–4.0 ml/min at room

temperature, 0.5–2.0 ml/min at cold room
temperature.

b. 50 ml cartridge: 2.5–20 ml/min at room

temperature, 2.5–10 ml/min at cold room
temperature.

2. Wash the prepared cartridge with degassed
buffer as indicated below.
a. 10 ml cartridge: 10 ml
b. 50 ml cartridge: 50 ml

3. Inject up to the amount of sample indicated
below, discarding the first amount of effluent
indicated below.
a. 10 ml cartridge: 6 ml of sample, discarding

the first 3 ml of effluent

b. 50 ml cartridge: 30 ml of sample, discarding

first 15 ml of effluent

4. Monitor the cartridge effluent with a UV monitor
or conductivity monitor.

15

Section 6
Technical Assistance

For additional information and technical assistance,
contact your local Bio-Rad representative as listed
on the back cover of our catalog, or in the US, call
Technical Support at 1-800-4BIORAD.

16

Section 7
Ordering Information

Bio-Scale Mini Cartridges*

Catalog #s

Description 5 x 1 ml 1 x 5 ml 5 x 5 ml

UNOsphere

™

Q Support 732-4100 731-4102 731-4104
UNOsphere S Support 732-4110 731-4112 731-4114
Macro-Prep

™

High Q Support 732-4120 732-4122 732-4124
Macro-Prep High S Support 732-4130 732-4132 732-4134
Macro-Prep DEAE Support 732-4140 732-4142 732-4144
Bio-Gel P-6 Support — 732-4502 732-4504
Affi-Prep

®

Protein A Support 732-4600 732-4602 —

Profinity

™

IMAC Support 732-4610 732-4612 732-4614
Profinity GST Support 732-4620 732-4622 732-4624
Affi-Gel

®

DEAE Blue Support — 732-4632 732-4634

Affi-Gel Blue Support — 732-4642 732-4644

1x 5x

Bio-Gel P-6 Desalting, 10 ml — — 732-5304
Bio-Gel P-6 Desalting, 50 ml — 732-5312 732-5314
* Visit www.bio-rad.com/cartridges/ for current information on

prepacked cartridges.

• Larger package sizes of media are available for process-scale

chromatography. Inquire with your local Bio-Rad representative.

17

Fittings Kits

18

Catalog # Description

732-0111 Luer to M6 Adaptor Fittings Kit, includes luer to

732-0112 Luer to 10-32 Adaptor Fittings Kit, includes luer

732-0113 Luer BioLogic System Fittings Kit, includes

M6 fitting to connect to an FPLC system

to polypropylene/PTFE 10-32 fittings to connect
1 cartridge to an HPLC system

1/4-28 female to male luer and 1/4-28 female to
female luer to connect 1 cartridge to the
BioLogic DuoFlow system

Section 8

19

References

Gagnon P, Avoiding Instrument Associated Variation
in Purification Scale-up and Scale-down, BioPharm,
10 (3) 42–45, 1997

Harris ELV and Angal S, Protein Purification Methods,
A Practical Approach, IRL Press, Oxford, 1989

Scopes RK, Protein Purification, Principles and
Practice, (2nd Edition), Springer-Verlag, New York,
1987

Snyder LR and Kirkland JJ, Introduction to Modern
Liquid Chromatography, (2nd Edition), John Wiley &
Sons, Inc., New York, 1979

FPLC is trademark of GE Healthcare. Luer-Lok is trademark of
Becton, Dickinson and Co. Triton is a trademark of Union Carbide.

Bio-Rad Laboratories, Inc.
2000 Alfred Nobel Dr.
Hercules, CA 94547 USA
510-741-1000
1-800-424-6723

10006995 Rev C