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Shell 400/Shell 700
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  • Shell 400/Shell 700

Shell 400/Shell 700

Aplo Shell 400 and Aplo Shell 700 are multi-mode chromatography resin with molecular sieve, hydrophobic and ion exchange functions. There is an inert shell that does not contain any ligands on the surface of the medium microspheres. This shell has no adsorption effect on proteins. However, because the shell has a specific pore size, this shell can act as a molecular sieve, preventing large molecules or virus particles from entering the interior of the microsphere. The core of the microsphere contains ligands with dual functions of ion exchange and hydrophobicity, which can bind to most proteins. Aplo Shell 400 and Aplo Shell 700 use highly rigid agarose as the base frame, which has the characteristics of strong rigidity, high pressure resistance, and fast flow rate. It exhibits high dynamic binding capacity under high flow rate conditions and can process large volumes in a short time.
Brand Series Aplo
Product Type Prepacked columns, Resins
Pack Size 1mL 5mL 10mL 25mL 100mL 500mL 1L 5L 10L
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DetailOrder InformationDocuments

Product

Aplo Shell 400

Aplo Shell 700

Features

High rigidity, high flow rate, large sample loading capacity

High rigidity, high flow rate, large sample loading capacity

Application

Separation and purification of biological macromolecules such as viruses, exosomes and vaccines

Separation and purification of biological macromolecules such as viruses, exosomes and vaccines

Matrix

High-rigidity agarose

Particle size

70-120 μm

65-85 μm

Functional group

Octylamine

Shell exclusion molecular weight

400 kDa

700 kDa

Flow velocity

＜600 cm/h

＜600 cm/h

Max. pressure

＜0.5 MPa

pH stability

3-12(operational), 2-14(CIP, short term)

Chemical stability

Stable in commonly used buffers, such as 1 M NaCl, 1 M NaOH, 6 M guanidine hydrochloride, 8 M urea, 70% ethanol, 30% isopropyl alcohol, etc.

Storage conditions

20% ethanol; 4-30℃

Pack size

Cat. No.

Aplo Shell 400

Aplo Shell 700

1ml XTrap Prepacked columns

HC5080111

HC5080211

5ml XTrap Prepacked columns

HC5080112

HC5080212

5ml XScreen Prepacked columns

HC5080113

HC5080213

10ml

HC5080100

HC5080200

25ml

HC5080101

HC5080201

100ml

HC5080102

HC5080202

500ml

HC5080103

HC5080203

1L

HC5080104

HC5080204

5L

HC5080105

HC5080205

10L

HC5080106

HC5080206


Operating Instructions

1. Column Packing

1.1 Packing Buffer Preparation

Purified water; degas byultrasonication for 15 min.

1.2 Resin Preparation

Calculate the required amount ofresin using a compression factor of approximately 1.15, and measure out theresin. Exchange the storage solution for packing buffer using a vacuumfiltration flask. Add sufficient packing buffer to prepare a resin slurry ofapproximately 50%.

1.3 Column Preparation

Inspect the chromatography column toensure that all components are intact and clean. Install the bottom adaptor andtighten the O-ring. Secure the column vertically on a laboratory stand; checkand adjust its alignment with a spirit level. Fill a syringe with packingbuffer, connect it to the bottom outlet, and slowly expel the buffer to removeair bubbles from the bottom screen. Remove the syringe and install the bottomplug. Add packing buffer to a height of approximately 2 cm in the column.

1.4 Column Packing (example: 16 mm column diameter, 10 cm bedheight)

Mix the resin slurry thoroughly and pour it slowly into the columnalong a glass rod. If the liquid level remains below the top of the column, addpacking buffer to fill the column. Connect the adaptor to the chromatographysystem, start the pump, and remove air from the tubing and top screen at anappropriate flow rate; then pause the system. Insert the adaptor into thecolumn at a 45° angle, secure it, and tighten the sealing ring while preventingair from entering. Remove the bottom plug and direct the bottom tubing into awaste container. Pack at 60 cm/h until the resin-bed interface stabilizes.Increase the linear velocity to 600 cm/h, with the high-pressure alarm set to0.5 MPa, and maintain the final rate for 45 min. Mark the resin-bed interfaceand stop the system. Reinstall the bottom plug and disconnect the top of thecolumn from the pump. Loosen the adaptor sealing ring slightly, lower theadaptor to 2 mm below the resin surface, and retighten the seal. Connect thetop and bottom column tubing to the chromatography system and perform acolumn-efficiency test.[TR1]

2. Column Efficiency Test

Test column efficiency immediatelyafter packing. Column performance is typically assessed using the number oftheoretical plates per meter (N/m) and the peak asymmetry factor (As). Highercolumn efficiency indicates greater separation capability. For a more accurateassessment, use a test-sample volume of 1.0% of the column volume and a linearvelocity of 15-30 cm/h. Inject the sample as close to the column inlet aspossible to minimize dilution.

Table 2. Column EfficiencyTest Methods

Item

Contents

Test Method

Acetone Method

NaCl Method

Sample

1.0% (v/v) acetone in water

0.8 M NaCl solution

Sample Volume

1.0%CV*

1.0%CV*

Mobile Phase

Water

0.4 M NaCl solution

Flow Rate

30 cm/h

30 cm/h

Detection

UV 280

Conductivity

Note*: CV (Column Volume) refers tocolumn volume and is abbreviated as CV below.

Column Efficiency Calculation

Calculate N/m and As from the UVcurve (or conductivity curve) using the following equations:

N/m = N/L

N = 5.54(VR/Wh)2

As=b/a

Note: In the equations, VR is theretention volume; Wh is the peak width at half height; L is the bed height inmeters; and N is the number of theoretical plates. VR and Wh must use the sameunits. In Figure 1, a is the leading half-width of the peak at 10% peak height,and b is the trailing half-width at 10% peak height.

Figure 1. SchematicDiagram for Column Efficiency Calculation

Column efficiency test chromatogram showing retention volume and peak widths

The peak should be symmetrical, withan asymmetry factor (As) as close to 1.0 as possible; values between 0.8 and1.8 are generally acceptable. N/m > 4,000.

3. Separation and Purification

3.1 Column Equilibration

Connect thechromatography column to the purification system. Equilibrate the column withat least 3-5 CV of equilibration buffer until the pH and conductivity of thecolumn effluent match those of the binding buffer. After equilibration, zerothe UV detector.[TR2]

3.2 Sample Loading

Load the viral-particle-containingsample onto the column. Sample volume depends on resin binding capacity, theconcentration of resin-binding molecules in the sample, and otherchromatography conditions. Collect the flow-through containing the purified viralparticles.

3.3 Column Washing

Wash with 2-3 CV of equilibrationbuffer to recover the entire flow-through fraction. Continue collecting theflow-through.

3.4 Cleaning and Regeneration

Wash the column with2-3 CV of 1 M NaOH containing 27% propanol. Equilibrate with at least 3-5 CV ofequilibration buffer until the effluent pH and conductivity match those of thebinding buffer.[TR3]

3.5 Storage

Wash with purified water until theconductivity of the column effluent reaches 0, followed by 2-3 CV of 20%ethanol. Store the column at 4-30°C.

Note: Select the buffer typeaccording to the pH required for purification of the target molecule. Filterthrough a 0.22 μm membrane.

4. Cleaning in Place

Cleaning in place (CIP) removesstrongly bound, precipitated, or denatured material from the resin. Residualimpurities can impair column performance; substantial accumulation can clog thecolumn, increase backpressure, and reduce flow rate. Regular CIP preventscontaminants from accumulating in the bed and helps maintain resin bindingcapacity and flow properties. A cleaning solution of 1 M NaOH containing 27%propanol is typically used.

If the column will be usedimmediately after cleaning, wash with 3-10 CV of binding buffer.

5. Sanitization

Sanitization minimizes microbialcontamination in the chromatography column. Aplo Shell 400/Aplo Shell 700chromatography resin can be sanitized with NaOH. NaOH effectively removesviruses, bacteria, yeasts, and endotoxins at very low operating cost. Followthe procedure below:

5.1 Wash the column with 3-5 CV ofbinding buffer.

5.2 Wash the column with 2-5 CV of0.5 M NaOH.

5.3 Soak the column in 0.5-1 M NaOHfor 1 h.

5.4 Complete sanitization by washingthe column with 5-10 CV of binding buffer at pH 7-8.

Precautions:

High NaOH concentrations orprolonged exposure may reduce resin binding capacity. Carefully control theconcentration and contact time during sanitization.

6. Storage

Store unused resin at 4-30°C in adry, well-ventilated, clean area. Keep the container tightly closed. Do notfreeze.

Store packed columns sealed andimmersed in 20% ethanol to prevent microbial contamination.

Note: Before using stored resin,perform CIP and equilibrate with at least 5 CV of binding buffer.

7. Linear Scale-up

A purification process optimized atlaboratory scale can be scaled linearly to pilot or production scale. Considerthe following during scale-up:

Keep the residence time constant tomaintain stable dynamic binding capacity.

Select the column volume based onthe required binding capacity. If the bed height changes, assess the effect onthe purification step.

Determine the column diameter fromthe required flow rate and the bed height from the established residence time.A bed height of 10-25 cm is generally recommended.

Ensure uniform sample concentrationand consistent elution conditions.

Troubleshooting

Table 3. TroubleshootingRecommendations

Issue

Possible Cause and   Recommended Solution

Excessive pressure during use

Replace the inline filter.

The chromatography column is  clogged; perform CIP cleaning.

Unstable pressure curve

Remove air bubbles from the sample  loading pump.

Use a bubble trap to remove air  bubbles from the sample.