Good column packing is essential for any chromatographic process and plays a key role in the large-scale commercial manufacture of biopharmaceuticals. A bed packed too densely might crack, which can lead to channeling and early breakthrough. A bed packed too loosely can further compress causing a liquid gap where mixing can occur.

Either instance will lead to costly process disruptions and loss of valuable product. Proper column packing eliminates such concerns and ensures a stable bed that performs according to expectations over many processing cycles.

This procedure describes packing of MabSelect PrismA resin in large-scale chromatography columns including AxiChrom, BPG, and Chromaflow columns. The procedure includes robust and verified packing methods that will help eliminate concerns and risks associated with poorly packed beds.

Chromatography resin and column characteristics

MabSelect PrismA chromatography resin

MabSelect PrismA is an affinity BioProcess resin for capturing monoclonal and bispecific antibodies (mAbs and BsAbs) as well as Fc-containing recombinant proteins. The combination of a high binding capacity and a high-flow base matrix gives excellent productivity.

General characteristics regarding flow velocity for MabSelect PrismA is at least 300 cm/h in an AxiChrom 300 column with 20 cm bed height at 20°C using process buffers with the same viscosity as water and pressure over bed < 2 bar (29 psi, 0.2 MPa).

The high-ligand stability of MabSelect PrismA reduces ligand leakage and offers exceptional alkali-tolerance. The protein A-derived ligand allows the use of 0.5 to 1.0 M sodium hydroxide (NaOH) for cleaning in place (CIP). The resin offers highly concentrated elution pools, which enables operating flexibility and small unit operation.

AxiChrom columns

AxiChrom columns are low-pressure, mechanical axial compression chromatography columns designed for process development and biopharmaceutical manufacturing environments (Fig 1). Mechanical axial compression enables accurate and reproducible control of the packing, even with large-diameter columns.

AxiChrom columns are available in many different configurations and materials. The columns are designed to be scalable and will give predictable results over the entire range of scales by enabling a uniform plug flow through the bed, irrespective of column size.

The columns feature Intelligent packing with preprogrammed methods that support all column sizes. Intelligent packing enables straightforward operation and high packing success rates. The chromatography column packing methods described here apply to bed heights up to 30 cm in AxiChrom columns up to 1000 mm i.d. A short guideline for larger columns is also provided.

Verified chromatography resin packing methods are available for AxiChrom columns

Fig 1. Verified chromatography resin packing methods are available for AxiChrom columns. These columns provide excellent packing control.

BPG columns

BPG columns are glass columns for process development and manufacturing. The single-screw adapter allows easy and efficient packing and running. The columns have diameters from 100 to 450 mm. The column packing methods described here are developed in BPG 300.

Chromaflow columns

Chromaflow columns are acrylic or stainless steel, pack-in-place columns for biopharmaceutical manufacturing. The columns have diameters ranging from 400 to 2000 mm. The chromatography column packing method described here applies to Chromaflow columns up to 800 mm i.d. A short guideline for columns larger than 800 mm is also provided.

Packing MabSelect PrismA resin

Definitions

The bed height of a gravity-settled bed differs from the bed height of a bed settled at low flow (consolidated). Therefore, the compression factor (CF) has to be separated from the packing factor (PF). In water, for example, the consolidated bed height (at 60 cm/h) is higher than the gravity settled bed height. For MabSelect PrismA resin the PF in this example would be 1.18 while the CF would be 1.1.

Equations to calculate CF, PF, and column volume (CV) are shown below:

Compression factor, CF = Lsettled / Lpacked

Packing factor, PF = Lcons / Lpacked

where

Lsettled = bed height measured after settling by gravity (cm)

Lcons = consolidated bed height, that is, bed height measured after settling the resin at a given flow velocity (cm)

Lpacked = packed bed height (cm)

CV = Lpacked × AC

where

AC = cross-sectional area of the column (cm2)

When packing BPG and AxiChrom columns, PF is used in the packing procedure to calculate the packed bed height after the consolidation step. CF is used in the resin preparation step to calculate the resin volume needed to pack a desired bed height. Because Chromaflow columns are pack-in-place columns, they have no registered consolidated bed heights and the CF is used throughout the packing process.

Properties of MabSelect PrismA resin in various packing solutions

The MabSelect PrismA resin settles differently in different solutions. Adding NaCl to the packing solution slows the settling of the resin beads and allows them to settle looser. As little as 1 mM NaCl increases the consolidated bed height by 2% compared to consolidation in water. It is important to measure the chromatography resin in the correct solution to minimize the effect on settling.

Verification of the packing method for MabSelect PrismA in AxiChrom 50 to 1000 suggested that the preferred packing solution is water.

Table 1 shows typical packing factors (PFs) for MabSelect PrismA in different solutions if the recommended packing solution cannot be used. The listed PFs are also relevant for large-scale AxiChrom columns with an i.d. of 300 mm or larger, at a bed height of 20 cm.

Table 1. Typical packing factors for MabSelect PrismA resin in different solutions for optimal bed performance where the bed is consolidated at 60 cm/h.

SolutionPacking factor
Water1.18
20% ethanol1.18
10 mM NaCl1.20
0.4 M NaCl1.20
2% benzyl alcohol1.18*

* 2.0% v/v benzyl alcohol is compatible with the acrylic material used in AxiChrom columns at room temperature for up to one year.

To optimize the bed and flow performance of MabSelect PrismA in the larger AxiChrom columns ≥ 300 mm, a variable PF depending on bed height is needed. Figure 2 shows PFs for MabSelect PrismA packed in AxiChrom 300 to 1000 with a bed height of 10 to 30 cm. Water was used as packing buffer.

Graph showing packing factor interval for large-scale AxiChrom columns packed with MabSelect PrismA resin to different bed heights.

Fig 2.PF interval for AxiChrom 300 to 1600 columns packed with MabSelect PrismA resin to different bed heights in water with a consolidation velocity of 60 cm/h. The PF varies in relation to bed height. PF is calculated as y = −0.002x + 1.22 (where y is the calculated PF and x the packed bed height). For each bed height, the acceptable interval for the packing factor is ± 0.03.

Slurry preparation

Start by calculating the resin volume (V) needed to pack the desired bed height. In this step, use CF 1.10, which is the CF in 20% ethanol for the gravity-settled resin. This CF corresponds to the right packing factor in the method.

Measuring slurry concentration

To achieve the correct amount of chromatography resin for packing to target bed height or compression, it is important to measure the slurry concentration correctly. Measuring slurry concentration can be performed with a Tricorn 10/100 column. Cytiva offers a Slurry Concentration Kit with all the materials required for determination of slurry concentration.

Packing MabSelect PrismA in AxiChrom columns

Both 10 and 20 µm bed support (SS and PE) can be used in AxiChrom columns with MabSelect PrismA. If a crude feed is used it would be preferable to use a 20 µm bed support. When packing AxiChrom 50 to 200 columns with ÄKTA systems, Intelligent packing control is managed by the UNICORN system control software. For AxiChrom 300 to 1600 columns, Intelligent packing is performed by the AxiChrom Master, a separate unit that comprises a touchscreen-operated user interface, or from the UNICORN software on the ÄKTAprocess system.

Intelligent packing in AxiChrom columns—general considerations

Chromatography column packing methods are created by entering values for the packing variables (e.g., column, resin, slurry concentration, target bed height) in the Intelligent packing wizard. The packing factor given in the Intelligent packing wizard is dependent on entered packing variables and the packing solution. To pack the column, start the chosen method in the UNICORN software and follow the instructions.

When packing AxiChrom 50 to 200 columns, the slurry is introduced into the column by hand and adapter movement is driven by internal hydraulics. For ÄKTA chromatography systems with Intelligent packing support for AxiChrom 50 to 200, a slurry recipe can be generated with packing solution recommendations. The recipe is valid for slurry concentration measurements performed with resin in 20% ethanol that has settled by gravity. The method controls the flow rate of hydraulic fluid to drive the adapter and packing of the bed (Fig 3).

Intelligent packing in small AxiChrom columns.

Fig 3. Intelligent packing in small AxiChrom columns. The adapter is mounted to the column tube and the wizard is started (Start). The adapter moves down, forcing packing liquid out through the bottom bed support. The resin forms a consolidated bed (Consolidation). When the adapter comes in contact with the consolidated bed surface, the operator initiates bed compression in the UNICORN wizard (Compression start). Compression occurs according to a predetermined PF. The target bed height is attained (Packed).

Column packing in larger AxiChrom columns

In AxiChrom 300 to 1600 columns, slurry is introduced via a resin valve in the center of the bottom bed support and the adapter is driven by an electric servomotor. The two-position resin valve enables filling, packing, and unpacking without adjusting the assembled column.

After the column is primed, the adapter rises from its lowest position and the column fills with slurry via the resin valve. The slurry volume is calculated automatically from the target bed height, slurry concentration, and PF. As an electric servomotor controls the movement of the adapter, its position is monitored with millimeter accuracy.

When the correct slurry volume has been drawn into the column, the adapter starts to move downwards and packing buffer is forced out through the bottom bed support and bed consolidation starts. The time to complete consolidation (i.e., when the adapter reaches the bed) is automatically calculated (as for the AxiChrom 50 to 200 columns), allowing the operator to carry out other tasks in the meantime.

As the adapter meets the consolidated bed, it is detected by the Intelligent packing wizard. When this occurs, the operator confirms the bed detection.

The compression of the resin starts and a graphical interface is shown on the control screen of UNICORN or AxiChrom Master. This graphical interface assists the operator in finishing the packing, giving a well-packed bed. When the adapter symbol is within the range of approved packing factors and bed height limits, the operator can end the packing.

If selected in the UNICORN wizard, Intelligent packing will automatically run a packed bed evaluation test after the packing. For large AxiChrom columns, automatic methods for priming and unpacking can also be created with the Intelligent packing wizard.

Depending on the AxiChrom Master version and installed control software, there can be additional alternatives for packing the AxiChrom 300 to 1600 columns. The operator must select one of the alternatives for packing before the packing process can start.

The three alternatives that could be available for packing are:

  • Automatic bed detection confirmation and stop at the set packing factor
  • Automatic bed detection confirmation and stop at the set bed height
  • Confirm bed detection and manually stop packing at the preferred bed height or packing factor (described in this procedure)

Packing MabSelect PrismA resin in BPG columns

Summary of the verified packing method in BPG 300 at 20 cm bed height

Slurry and packing solution: 20% ethanol

Slurry concentration: 60% (±2)

Consolidation velocity: 30 cm/h (21.2 L/h)

Consolidation time: 50 min (±5)

Mechanical compression (packing factor): 1.13 (±0.01)

Column and system preparation

A detailed description of column preparation is available in the BPG column operating instructions. The packing pump should be as pulsation-free as possible. Screw or rotary lobe pumps are the most suitable for this task and multi-headed diaphragm pumps are satisfactory.

Note! Before starting the column packing, ensure that the chromatography column has no visible scratches in the glass tube and that the adapter moves smoothly in both upward and downward directions. In addition, there should be no difficulty in tightening the adapter O-ring to the column inner wall.

Follow the steps below to prepare the BPG column:

  1. Place a new 10 μm net on both adapter and bottom end piece.
  2. Level the column with the aid of a spirit level.
  3. A pressure relief valve should be used for safety reasons, especially against pressure spikes. Position this valve on the pump outlet and add a pressure gauge on the adapter.
  4. Mount one 4-port, 2-way valve on bottom inlet and one on top of the pressure gauge, 10 mm i.d. for BPG 300.

Resin preparation

Dilute the resin slurry to the desired concentration using 20% ethanol.

BPG column packing

  1. Set the pressure alarm or pressure relief valve according to the pressure specification (4 bar). Purge the system and tubing from air.
  2. Purge the net of trapped air by draining packing solution through the column outlet until about 2 cm of solution remains in the column. Close the bottom valve. If air is still trapped under the end-piece net, add more packing solution and connect a tube to the suction side of a pump. Start the pump and place the pump inlet tube on the bottom net and extract the remaining air.
  3. Add the slurry to the column and, if needed, additional packing solution to about 40 cm. Mix the resin and the packing solution to a homogeneous slurry. Note! The available height to allow the adapter to be inserted into a 50 cm column tube (for filling slurry) is 40 cm. Use a longer column tube when packing beds higher than 20 cm (75 cm and 95 cm tubes are available).
  4. Rinse the wall from particles and let the resin settle until there is about 1 cm of clear liquid on top of the slurry. This reduces the risk of particles sticking between the O-ring and the column wall, which can cause the column to leak.
  5. Insert the adapter and secure it to the column tube. Lower the adapter to the surface of the slurry and allow some clear liquid to pass the O-ring. Tighten the adapter O-ring.
  6. Make sure the column top valve is open. Slowly move the adapter down until no air bubbles can be seen leaving the top valve.
  7. Start the pump and adjust the settling velocity to 30 cm/h. Shift the top valve into the column and immediately open the bottom valve and lead the liquid to waste.
  8. Run the settling flow until the bed is completely consolidated. Note the consolidated bed height and calculate the packed bed height using the PF (1.13) in 20% ethanol. The packed bed height is the ratio between the consolidated bed height and the packing factor. Use a marker to indicate the packed bed height on the column.
  9. Stop the flow and close the top valve. Loosen the O-ring and lower the adapter down to 1 cm above the settled bed and seal the adapter O-ring. Shift the bottom valve to waste and use the adapter to mechanically compress the bed to the mark on the column (step 8). Excess packing solution is removed through the bottom of the column.

The column is now ready for use.

Packing MabSelect PrismA resin in Chromaflow columns

Summary of verified packing method in Chromaflow 600 at 20 cm bed height

Pressure: Chromaflow Packing Station Pack 100 start pressure = 5.5 bar. Pressure is kept on 3 bar once column pressure has reached that level.

Packing solution: purified water

Stop position: > 5 mm from adaptor screen (to eliminate nozzle imprint)

Slurry concentration: ~ 50%

Preparations

The method described below to pack a Chromaflow 600 column at 20 cm bed height is based on a standard Chromaflow procedure (i.e., via nozzles in the top and bottom bed supports, and without removing the lid or adaptor). This method makes use of the optional Chromaflow Packing Station, which includes a control panel with pneumatically-actuated diaphragm pumps and valves, and a tank to hold the slurry.

The MabSelect PrismA resin is delivered in a 20% ethanol solution. It is, however, preferable to perform the packing in water. Therefore, it is recommended to first do an equilibration packing in water before the final column packing.

To avoid introducing air into the column when packing, additional slurry is required for the extra volumes in tank and tubing. Add the slurry to the slurry tank and stir the resin. Dilute the suspension to about 50% slurry concentration.

Column preparation

A more detailed description about the column and pack station preparation can be found in the operating instructions for Chromaflow columns and Chromaflow Packing Stations. In this procedure, standard Chromaflow nomenclature is used for connections on the column and packing station.

Note! It is important that the supply air flow rate follows the specification of the Chromaflow Packing Station (1000 L/min for Pack 100) and that the supply air pressure into the packing station is 6 to 7 bar.

  1. Set up the column according to the Chromaflow columns operating instructions.
  2. A pressure relief valve should be used for safety reasons (see Chromaflow operating instructions and the column product documentation for pressure limitations). Position this valve on the slurry inlet top (SIT), with the waste tubing connected to the slurry tank. Place a pressure gauge on the mobile phase top (MPT) to record the pressure during packing. Mount one 3-port, 2-way valve on top of the pressure gauge and one on the mobile phase bottom (MPB). The top valve should lead in two directions: one side into the system and one to waste for purging the tubing. On the bottom valve, one side leads to the system and a 1.5” to 2” tubing leads to waste (for packing). Part of the MPB waste tubing should be placed above the outlet valve to prevent air from entering through the MPB.
  3. Connect appropriate tubing (i.d. 1” or 1.5”) and tanks to the column and packing station. If a flow meter is used, place it between the SIT and the packing station.
  4. Level the adapter to the desired bed height. Remember to loosen the nuts on the adapter rods to allow the adapter to be raised or lowered. Flush the adapter rods with 20% ethanol as lubrication.
  5. Prime the column, packing station, and tubing with water according to the operating instruction.

Chromaflow column packing

Note! Packing Chromaflow columns is a rapid procedure compared with other packing procedures and it is therefore important to thoroughly read the packing instructions and practice the packing steps in advance of the column packing.

  1. Set both nozzles in run position to prime the top nozzle tubing with slurry. Lead the slurry outlet top (SOT) tubing back to the slurry tank and secure it. Stir the slurry to keep it homogeneous, select slurry and SIT on the packing station, open the slurry tank, and start the packing pump.
  2. As the aim of this procedure is to prime the tubing and allow the pump speed to be set, the chromatography column is bypassed at this stage. Increase the pump flow until the pressure on Chromaflow Packing Station Pack 100 equals 5.5 bar (or about 101 L/min). Figure 4 shows the increase in pressure and decrease in flow velocity when packing a 20 cm bed height in Chromaflow 600.
  3. When the tubing is primed and the flow rate set, set the SIT/slurry inlet bottom (SIB) valve to the position between SIT and SIB on the Chromaflow Packing Station to block the flow during step 4, while maintaining the pump pressure/correct flow rate for the next step. If a flow meter with flow totalizer function is used, reset the flow totalizer.
  4. Move the top nozzle down into the packing position.
  5. Simultaneously open the bottom mobile phase valve to waste and turn the SIT/SIB valve to SIT on the packing station, this is preferably done by two operators. The column then starts to fill with slurry and the bed builds up slowly from the bottom as excess liquid exits via the MPB. The start condition for the packing station should be 5.5 bar.
  6. Note! Column pressure should not exceed the operating pressure limit of the column (i.e., 3 bar). If this pressure is reached, gently decrease the packing flow so that the pressure remains just below 3 bar.

  7. Stop the packing pump when the bed is somewhere between 1 and 0.5 cm from the top bed support by setting the SIT/SIB to the position between SIT and SIB, as described in step 3.
  8. Note! If a non-transparent (e.g., stainless steel) column tube is used, stop the packing flow when the calculated volume of slurry, has been introduced into the column. Check the volume in the slurry tank or use a volume totalizator.

  9. Immediately move the top nozzle back to the run position. The bed will then expand until it reaches the adapter.
  10. Close the MPB valve when the pressure in the column is between 0.3 and 0.1 bar. If a flow totalizer is used, register the total slurry volume pumped through the column.
  11. Use packing solution to rinse residual resin from the tubing and the top nozzle. Pump the packing solution through the top nozzle back into the slurry tank.
  12. Close the slurry tank and empty the tubing between the tank and packing station.
  13. Equilibrate the packed bed up flow to remove any trapped air in the column.

Graph showing column pressure and flow velocity during packing of MabSelect PrismA resin to a 20 cm bed height in a Chromaflow 600 column.

Fig 4. Column pressure and flow velocity during packing of a 20 cm bed height in Chromaflow 600 using Chromaflow Packing Station Pack 100. The graph shows that the pressure is kept constant at 3 bar after 60 seconds.

Testing the performance of the packed column

Process-scale chromatography columns must perform with a high degree of efficiency over many processing cycles (i.e., display high stability). The efficiency of a packed column can be expressed in terms of height equivalent to a theoretical plate (HETP) and asymmetry factor (As). This test should be repeated regularly to monitor the state of the bed throughout the working life of the column.

If the test results are to be comparable over time, conditions such as fluid velocity (cm/h), liquid pathway, sample composition, and elution buffer should be kept constant. The requirements for the test have to be set in accordance with test conditions and the goal of the purification. The test is further described in the application note Column efficiency testing.

Test conditions used in this study

Sample: 2% v/v acetone

Sample volume: 1% of the CV

Test velocity: 30 cm/h for AxiChrom, BPG, and Chromaflow columns

Eluent: water

To compare the performance of columns packed with chromatography resin of different particle diameters, the reduced plate height (h = HETP/average bead diameter [dp]) is typically used. As a guideline, a value of h < 3 is considered good at optimal test conditions.

Efficiency tests at different flow velocities

Efficiency tests were run at different flow velocities. Figure 5 shows a Van Deemter plot of the results. The asymmetry factor is stable at the different flow velocities. The reduced plate height increases with the flow velocity, which is in accordance with the van Deemter equation. In this study, optimal results were achieved at 20 to 40 cm/h.

When running at higher flow velocities, the asymmetry factor and reduced plate height continue to show a linear correlation, indicating that the efficiency test can be run at any flow velocity. However, the expectation of the result needs to be adjusted based on the test flow velocity used.

Van Deemter plot for an AxiChrom 300 column packed with MabSelect PrismA resin at 20 cm bed height.

Fig 5. Van Deemter plot for AxiChrom 300 packed with MabSelect PrismA resin at 20 cm bed height.

Examples of results for packed columns

Columns packed with the methods outlined above were tested for plate number, asymmetry, pressure-flow properties, and stability.

AxiChrom column results

Table 3 provides examples of efficiency and stability results for MabSelect PrismA packed in AxiChrom columns ranging from 50 to 1000 mm i.d. These results show that the verified packing methods used give excellent packing in AxiChrom 50 to 1000. The stability test showed that the beds were stable when running in water at the velocities given in Table 3.

Pressure-flow curves provide a simple, yet effective illustration of column performance in terms of the maximum operating velocity at which the purification process can be run. These curves also show the magnitude of the back pressure in the system at a certain liquid velocity. AxiChrom columns can use the full flow potential of MabSelect PrismA, even at large column diameters (Figs 6 to 11).

Table 3. Column efficiency data for different packings of MabSelect PrismA in different AxiChrom columns at different bed heights

ColumnBed height (cm)Average plates/m1Reduced plate height (h) range1Asymmetry factor (As) range1Flow velocity for stability test (cm/h)2Change after stability test (%)
      hAs
AxiChrom 501010 9001.4–1.61.1–1.1600-20
2011 0001.5–1.51.0–1.030071
4011 4001.4–1.50.8–0.915052
AxiChrom 1001012 0001.3–1.41.1–1.160074
2012 1001.3–1.41.0–1.130053
4010 9001.4–1.61.0–1.115032
AxiChrom 1402012 3001.3–1.41.0–1.130034
AxiChrom 2001011 2001.4–1.51.2–1.360056
2011 7001.4–1.41.1–1.130045
4011 1001.4–1.51.0–1.1150-24
AxiChrom 3001010 0001.6–1.71.3–1.4600-3-6
2011 0001.5–1.61.1–1.2300-3-4
3010 8001.4–1.61.0–1.32007-7
AxiChrom 60032010 3001.5–1.51.1–1.1300-23
3010 1001.5–1.61.1–1.120054
AxiChrom 10003 10 11700 1.4−1.5 1.2−1.4 NA NA NA
20 11100 1.5−1.6 1.1−1.2 300 3 3
30 10200 1.5−1.8 0.9−1.1 NA NA NA
AxiChrom 16003 10 10900 1.4−1.7 1.2−1.4 550 -2 -1.5
20 10400 1.5−1.7 1.1−1.2 300 0 6
30 10000 1.6−1.8 1.1−1.2 200 -2 0

1 Test performed at near optimal test conditions. Average and ranges of upflow and downflow tests for at least three packings.

2 Stability tests were run once for each bed height/resin/column combination in water for 16 h at the given velocity.

3 Results from studies performed on MabSelect PrismA base matrix.

Pressure flow curves for MabSelect PrismA resin packed in an AxiChrom 100 column at different bed heights.

Fig 6. Pressure flow curves for MabSelect PrismA resin packed in AxiChrom 100 at different bed heights. The results are normalized to 10, 20, and 40 cm bed height in water at 20°C. Tubing and system pressure are excluded.

Pressure flow curves for MabSelect PrismA resin packed in AxiChrom 50, 100, 140, and 200 columns at 20 cm bed height.

Fig 7. Pressure flow curves for MabSelect PrismA resin in AxiChrom 50, 100, 140, and 200 columns at 20 cm bed height. The results are normalized to 20 cm bed height in water at 20°C. Tubing and system pressure are excluded.

Pressure flow curves for MabSelect PrismA resin packed in a AxiChrom 300 column at different bed heights.

Fig 8. Pressure flow curves for MabSelect PrismA resin packed in AxiChrom 300 at different bed heights. The results are normalized to 10, 20, and 30 cm bed height in water at 20°C. Tubing and system pressure are excluded.

Pressure-flow curve for the MabSelect PrismA base matrix at 20 cm bed height in AxiChrom 600 and 1000 columns.

Fig 9. Pressure-flow curve for the MabSelect PrismA base matrix at 20 cm bed height in AxiChrom 600 and 1000. The results are normalized to 20 cm bed height in water at 20°C. Tubing and system pressure are excluded.

Pressure-flow curve for the MabSelect PrismA base matrix at 20 cm bed height in AxiChrom 600 and 1000 columns.

Fig 10. Pressure-flow curve for the MabSelect PrismA base matrix in AxiChrom 1000 at 10, 20, and 30 cm bed height at the optimal compression for each bed height according to Fig. 2. The results are normalized to 20 cm bed height in water at 20°C. Tubing and system pressure are excluded.

Pressure-flow curve for the MabSelect PrismA base matrix at 20 cm bed height in AxiChrom 600 and 1000 columns.

Fig 11. Pressure-flow curve for the MabSelect PrismA base matrix in AxiChrom 1600 at 10, 20, and 30 cm bed height at the optimal compression for each bed height according to Fig. 2. The results are normalized to 20 cm bed height in water at 20°C. Tubing and system pressure are excluded.

BPG column results

Examples of efficiency and stability results for MabSelectPrismA are shown in Table 4. Good plate numbers and asymmetry factors were achieved. The pressure-flow curve for the resin in BPG 300 column is shown in Figure 10. The pressure-flow profile for MabSelect PrismA in BPG 300, at 20 cm bed height, shows that the resin can be run at 300 cm/h with a pressure of less than 2.0 bar over the bed.

Table 4. Column efficiency data for MabSelect PrismA packed in a BPG column with 300 mm i.d.

ColumnBed height (cm)Average plates/m1Reduced plate height (h) range1Asymmetry factor (As) range1Flow velocity for stability test (cm/h)2Change after stability test (%)
      hAs
BPG 3002080001.7–2.81.2–1.530063

1 Test performed at near optimal test conditions. Average and ranges of upflow and downflow tests for at least three packings.

2 Stability tests were run once for each bed height/resin/column combination in water for 16 h at the given velocity.

Pressure/flow rate study on MabSelect PrismA resin packed at 20 cm bed height in a BPG 300 column.

Fig 10. Pressure/flow rate study on MabSelect PrismA resin packed at 20 cm bed height in a BPG 300 column. The results are normalized to 20 cm bed height in water at 20°C. Tubing and system pressure are excluded.

Chromaflow column results

The efficiency results for MabSelect PrismA packed in Chromaflow 600 are shown in Table 5. Good plate numbers and asymmetry factors were achieved.

The pressure-flow curve for Chromaflow 600 is shown in Figure 11. The pressure-flow profile for MabSelect PrismA in Chromaflow 600, at 20 cm bed height, shows that the resin can be run at 300 cm/h with a pressure of less than 2.0 bar over the bed.

Due to column design and flow restrictions, there is a possibility that large (> 800 mm i.d.) Chromaflow columns cannot be packed at sufficient packing velocity and for this reason cannot fully use the flow potential of MabSelect PrismA.

Table 5. Column efficiency data for different packings of MabSelect PrismA base matrix in a Chromaflow column with 600 mm i.d.

ColumnBed height (cm)Average plates/m1Reduced plate height (h) range1Asymmetry factor (As) range1Flow velocity for stability test (cm/h)2Change after stability test (%)
      hAs
Chromaflow 6002080001.8–2.30.9–1.1300-96

1 Test performed at near optimal test conditions. Average and ranges of upflow and downflow tests for at least three packings.

2 Stability tests were run once for each bed height/resin/column combination in water for 16 h at the given velocity.

Pressure-flow curve for the MabSelect PrismA base matrix at 20 cm bed height in a Chromaflow 600 column.

Fig 11. Pressure-flow curve for the MabSelect PrismA base matrix at 20 cm bed height in Chromaflow 600. The packed bed compression was 1.08. The results are normalized to 20 cm bed height in water at 20°C. Tubing and system pressure are excluded.

Conclusions

This procedure describes packing of MabSelect PrismA resin in AxiChrom, BPG, and Chromaflow columns. AxiChrom columns enable the full use of the resin’s high fluid velocity capabilities. In addition, these columns are available in a range of diameters and bed heights, allowing for more flexible and rapid processes. It is important to remember that each chromatography column packing method is related to a specific packing solution. Changes in packing solution can have a significant impact on the PF and subsequently on the packing results. To utilize the full flow potential of MabSelect PrismA, AxiChrom columns are recommended.

Ordering information

ProductQuantityProduct code
MabSelect PrismA bulk resin25 mL, 200 mL, 1 L, 5 L, and 10 L17549801 to 17549805
Media Wand 50128922764
Media Wand 100128922767
Media Handling Unit128922769
Slurry Concentration Kit129096100