ÄKTAcrossflow automated cross flow filtration for process development

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1 GE Healthcare Data File AB Membrane systems ÄKTAcrossflow automated cross flow filtration for process development ÄKTAcrossflow is the first fully automated system for cross flow filtration process development and is ideal for filter screening and process optimization at small-scale. ÄKTAcrossflow together with UNICORN control software puts you in control of your cross flow process to allow consistent simulation of large-scale conditions and provide data for comprehensive analysis of results. The benefits of working with ÄKTAcrossflow include: Broad range of applications that cover ultrafiltration and microfiltration Flexible operation of either hollow fiber cartridges or cross flow cassettes Thorough and efficient process development with full TMP and flux scouting Single familiar UNICORN interface for both chromatography and membrane separations No disruption to proteins or cells with low shear force pumps that require no cooling Minimum working volume of 25 ml ensures operation of complete processes using filters between 40 cm 2 and 150 cm 2 System design and description ÄKTAcrossflow consists of the separation unit, a computer with flat-screen monitor and UNICORN software (Fig 1). The compact, bench-top system is suitable for installation in laboratories, which reduces facility expenditure. The sanitary design means that it can formally qualify for use in current Good Manufacturing Practices (cgmp) processes. Fig 1. ÄKTAcrossflow has a compact and sanitary design. The system is built with ÄKTAdesign components for reliability, scalability and flexibility. ÄKTAcrossflow comes complete with UV, ph and conductivity detectors, as well as air, pressure and temperature sensors. The valves and the fittings are specially developed for hygienic, leak-free operation. Electrically actuated diaphragm valves diminish hold up volumes and prevent dead volumes. A wide range of cross flow devices include GE Healthcare hollow fiber cross flow cartridges for cell harvesting/ cell concentration and Kvick Start cassettes for product concentration/ diafiltration. ÄKTAcrossflow is designed to operate up to 150 cm 2 of membrane area and is suitable for processing small sample volumes.

2 Flow Restrictor Q T P T Air Valve Block 1 In 1 Air In 2 In 3 In 4 Valve Block 2 In 5 In 6 In 7 In 8 Pump (Module A) Level & Temperature L,T Vent Stirrer line Reservoir Recirculation line Feed Retentate Control Valve R-PCV Purge Valve Waste 1 P R Retentate Valve Block Out 2 Out 3 Retentate Permeate Permeate line Permeate Valve Block Permeate Control Valve Recycle Out 1 Out 2 Out 3 Fig 3. Sanitarily designed membrane valve. Fig 4. Sanitarily designed piston pump. Q F Feed Pump P F Out 1 Cartridge P P Cond UV Q P Permeate Pump (Module B) P-PCV ph Fig 2. The flow path. Fig 5. Retentate pressure control valve. The UNICORN control software allows quick, simple communication between systems and users. The software also meets the stringent control and data handling procedures required in modern production and laboratory facilities. Method wizards provide easy method generation. Optimized methods can be transferred from laboratory to production scale. Flow path Figure 2 shows the ÄKTAcrossflow flow path which comprises of a transfer line, a recirculation line, a permeate line, a reservoir plus the system pumps (Fig 3) and valves (Fig 4). The flow path components and sensors are sanitarily designed and are described in more detail below: line The transfer valve blocks select the appropriate inlet tubing among eight different inlets such as for sample, buffers, CIP (cleaning in place) and rinsing solutions. The transfer pump delivers liquid to the reservoir. Like all pumps in the ÄKTAcrossflow system, the transfer and permeate pumps are high precision metering pumps that act as flow meters by reporting accurate flow rates and volume information to the UNICORN control system. The transfer purge valve allows for a rinsing of the transfer line. A pressure sensor (P T ) measures the pressure in the transfer line for safety reasons. An air sensor can be used on the sample inlet tubing to allow for efficient use of the sample volume. The reservoir The reservoir contains the sample and retentate processed by the membrane. A magnetic stirrer in the bottom of the reservoir ensures uniform mixing between the bulk fluid, the retentate returned from the membrane and liquid added via the transfer line. An integrated level (L) and temperature sensor (T) at the bottom of the reservoir continuously reports the retentate liquid volume and the temperature of the liquid fed into the filter device. The level sensor can also be used to protect the filter against the introduction of air. Recirculation line The feed pump creates cross fl ow over the membrane. A valve (R-VB-Out1) in the retentate valve block before the cartridge allows liquid removal at high fl ow rates. (P F ) of the feed is measured at the inlet of the filter cartridge. At the retentate side of the cartridge, a pressure sensor (P R ) measures the pressure of the liquid rejected by the membrane that returns from the cartridge back to the reservoir. Figure 5 shows the retentate pressure control valve (R-PCV) is used to adjust the retentate pressure, for example under TMP (trans-membrane pressure) process control. The retentate valve block is comprised of three outlets; one pre-filter and two post-filter, and is used primarily for product recovery. Permeate line At the permeate side of the cartridge, a number of sensors monitor permeate pressure (P P ), conductivity (Cond), UV absorbance (UV) and ph (ph). The permeate pump is an AB

3 integral part of the permeate flow path, because it is used to control either a constant permeate flow or a constant permeate pressure. In TMP control, for example, the flow rate of the pump is regulated to maintain a constant permeate pressure set by UNICORN that continuously reports permeate flow rate. A pressure control valve (P-PCV) is situated after the permeate pump; this automatically raises the pressure to a level higher than the permeate pressure to assure flow rate accuracy by a proper function of the pump checkvalves. The flow path continues to the permeate valve block comprising four outlets. The fourth outlet is a preconfigured recycle line which is used for returning permeate back to the reservoir to allow steady state process analysis, such as, during process optimization with TMP scouting. Design for safety relief valves are incorporated in the retentate (R-VB-Out1) and permeate (R-VB-Out3) valve blocks for safety. The top cap of the reservoir relieves excess pressure in the event of malfunction due to a blocked vent filter. Furthermore, the system and filter cartridges are protected against overpressure by pressure alarms in the UNICORN control system. Reservoir and tubing ÄKTAcrossflow can be configured with reservoirs and tubing of different sizes to meet a broad range of application requirements regarding flow rate, sample and holdup volume. Each reservoir is equipped with a magnetic stir bar to ensure uniform mixing. A floating device in the reservoir prevents vortex and foam formation, to allow operation at with high recirculation flow rates with a minimum working volume. The standard reservoir size is 350 ml and a 1100 ml reservoir is available as an accessory. Two sanitarily designed tubing kits are available for the recirculation line to customize the ÄKTAcrossflow system to the specific application: 1.7 mm ID PVDF tubing and 2.9 mm ID EFTE tubing. The tube fittings have prefabricated molded tube ends and o-ring seals to prevent leakage in or out of the flow path. The small size of the tubings enables a compact system design. Pumps and valves The ÄKTAcrossflow system has high precision metering pumps. The pump heads are sanitarily designed with a selfcontained rinsing system to prevent contamination and pump damage. A verified low shear design ensures that sensitive cellular material is not damaged during operation. Furthermore, this design assures negligible heat transfer from the pump heads to the process fluid. The feed pump is equipped with four pump heads for gentle and low pulsation circulation of sensitive process fluids up to 600 ml/min. The transfer and permeate pump allow for 200 ml/min low pulsation flow. The precision of the pumps over the entire flow/viscosity range enables the system to operate without the need for flowmeters. The flow in the ÄKTAcrossflow system is controlled by four blocks of stepper motor actuated membrane valves and three lever actuated valve units. All valve units are sanitarily designed with EPDM membranes for high chemical resistance. The valve blocks comprise multiple valves to minimize hold up volumes. They have different numbers of inlet and outlet ports depending on their location in the flow path. The lever actuated valve units are configured either as three-way valve (transfer purge valve) or as pressure control valve (R-PCV and P-PCV). In both configurations, the EPDM encapsulated lever is actuated by a solenoid to open or close a flow path. For the control valves, the solenoid is adjusting the closing force of the level against the inlet. This novel design allows for a robust control of pressure and filtration process. UNICORN a fast and secure way to work UNICORN software controls ÄKTAcrossflow. UNICORN has the benefits of one familiar control platform and user interface for all scales of operation in both filtration and chromatography. The scouting feature gives automatic support to process development and optimization. Method wizards and pre-programmed cleaning methods provide a high degree of efficiency in scale-up and production work. UNICORN fully conforms to all applicable regulations including 21 CFR Part 11 (Fig 6). Complete documentation and protection Method Editor Method Controlled user access System Control Main Menu Fig 6. UNICORN fulfils stringent control and data handling procedures. Cross flow process control Logbook Result Report Validation Support Evaluation The ÄKTAcrossflow system with UNICORN control software supports the process control modes commonly used in ultrafiltration and microfiltration applications such as, TMP control and flux control. These control modes can be combined with selectable feed pump instructions, such as feed flow rate, feed pressure, Delta P, retentate flow rate or shear rate. UNICORN also reports real-time process parameters such as retentate volume, concentration factor, diafiltration exchange factor, and accumulated permeate volume AB 3

4 Fig 7. Product step selection in the method wizard. Fig 8. The operations avaialble in the evaluation module. Fig 9. Raw data recorded from process optimisation for BSA concentration/ diafiltration using TMP scouting with 100g/l and 20g/l BSA solutions. Fig 10. Curves generated in the evaluation module from the raw data (Fig 9). The lower three curves show the results for 100g/l BSA, the upper three curves show the results for 20g/l BSA. Method wizard With the method wizard, you can easily program your filtration methods for all the steps in a typical run. For example, it is possible to rinse new filters; CIP used filters and test water flux to check filter quality and status before each run. A system sanitization method is also provided in the method wizard. The water flux test is a typical example of the benefits when using an automated system. Water flux tests are easily included in the method, both before and after the filtration steps. From the evaluation wizard, you can gather the data for a certain filter to check its membrane flux recovery over multiple cycles. For product steps, up to three concentration or diafiltration/wash steps can be selected (Fig 7). It is also possible to run in fed batch or tank batch mode depending on your total initial processing volume. A special option is the process optimization method, where you select a certain recirculation condition and scout a series of TMP values to find the optimal TMP for your run. The method is linked to the evaluation module, where you can analyze your results. Evaluation module The evaluation module saves you time by eliminating manual transfer of data to spreadsheets. The module allows flexible and direct presentation of optimization results with five different evaluation operations (Fig 8). The operations include, process optimization, normalized water flux, diafiltration time optimization, capacity plots and any vs. any. Process optimization is used to analyze process characterization experiments where a series of set points are tested (Fig 9). The most common experiments are excursions of TMP at different retentate flow rates and protein concentrations. Process optimization makes a new plot from user-identified points along original data curves, for example, flux vs. TMP (Fig 10). Process optimization also allows the user to overlay multiple plots (i.e. flux vs. TMP at different retentate flow rates or protein concentrations). This capability can be used for any process parameter. The any vs. any evaluation operation is used to analyze results from routine concentration, diafiltration and cell processing runs. It allows you to plot any process parameter captured as a curve in a given result file on either the X-axis or the Y-axis. Filtration devices The system is designed for use with filtration cassettes and hollow fibers. A wide range of cross flow devices include GE Healthcare hollow fiber cross flow cartridges for cell harvesting/ cell concentration and Kvick Start cassettes for product concentration/ diafiltration. ÄKTAcrossflow is designed to operate up to 150 cm 2 of membrane area for processing small sample volumes AB

5 Sanitary design ÄKTAcrossflow has a design that allows effective sanitization using 1 M sodium hydroxide (NaOH) as a sanitizing agent. Sanitization is the use of a chemical agent to reduce a microbial population to acceptable, predetermined level. Microbial challenge tests evaluate the efficiency of the sanitizing agent. In a study the system has been subjected to high level (1x10 6 Colony Forming Units CFU/ml) of microbial challenge. Two strains of bacteria that are recommended by the United States Pharmacopeia (USP 27) for antimicrobial testing (Table 1) were used. The results show that the provided method is sufficient for sanitization and that the numbers of viable organisms were efficiency reduced. Table 1. The organisms used in the microbial challenge tests. Escherichia coli ATCC 8739 Gram negative bacteria Pseudomonas aeruginosa ATCC 9027 Gram negative bacteria Validation documentation & services Regulatory authorities expect companies that produce therapeutic or diagnostic products to qualify equipment before use in production or analysis. Fast Trak Validation is GE Healthcare s specialist service that provides validation documentation on request after delivery of ÄKTAcrossflow. Installation and Operational Qualifications (IQ/OQ) of production and analysis equipment are part of the formal qualification required by current Good Manufacturing Practices (cgmp). As such, this documentation is subject to inspection by regulatory authorities. Fast Trak IQ/OQ packages consist of professionally developed templates that will facilitate equipment qualification. The templates have help texts and are easy to complete or if assistance is required, Fast Trak can complete the qualification for you. Technical specifications Operating range Feed flow rate ml/min accuracy < ± 2% flow rate ml/min accuracy < ± 0.5% Permeate flow rate ml/min accuracy < ± 0.5% Max. system pressure 5.2 bar (75.4 psi) Min. recirculation volume < 25 ml (with 1.7mm ID tubing) Holdup volume, recirculation line < 20 ml (with 1.7 mm ID tubing kit, empty reservoir) Reservoir max. holdup volume 350 ml (375 ml without floater) 1100 ml (1200 ml without floater), optional Recommended membrane area cm 2 System specifications Dimensions (W L H): mm Weight, approximate: 70 kg Operating temperature: 4-40 C (50 C during membrane cleaning) Detection and control transducers at cartridge accuracy TMP control accuracy UV measurement ph measurement Conductivity measurement Ordering information Product Feed, retentate and permeate 0.01 bar (0.15 psi) 0.05 bar (0.73 psi) Hg lamp: 254 and 280 nm (other wavelengths optional) Zn lamp: 214 nm Range: 0 14 (spec. valid between 2 and 12) Range: 1 μs/cm ms/cm Code Number ÄKTAcrossflow Accessories 1100 ml reservoir Flat sheet membrane cassettes Item Quantity/pack Code no. Kvick Start, 45 cm 2, 5 kd, PES UFEST ST Kvick Start, 45 cm 2, 10 kds, PES UFEST SE Kvick Start, 45 cm 2, 10 kd, PES UFEST ST Kvick Start, 45 cm 2, 30 kd, PES UFEST ST Kvick Start, 45 cm 2, 50 kd, PES UFEST ST Kvick Start, 45 cm 2, 100 kd, PES UFEST ST Kvick Start PACK, 45 cm kd, 30 kd, 50 kd, 100 kd UFESTCPAK045ST AB 5

6 Hollow fiber cartridges HF Start AXM, 50 cm 2, 750 kd, 1 mm UFP-750-E-2U HF Start AXM, 50 cm 2, 0.1 μm, 1 mm CFP-1-E-2U HF Start AXM, 50 cm 2, 0.2 μm, 1 mm CFP-2-E-2U HF Start AXM, 50 cm 2, 0.45 μm, 1 mm CFP-4-E-2U HF Start AXM, 50 cm 2, 0.65 μm, 0.75 mm CFP-6-D-2U HF Start AXM PACK, 50 cm kd, 0.1 μm, 0.2 μm, 0.45 μm, 0.65 μm Hollow fiber hoop cartridges HF Start AXH, 40 cm 2, 3 kd, 0.5 mm UFP-3-C-H24U HF Start AXH, 40 cm 2, 10 kd, 0.5 mm UFP-10-C-H24U HF Start AXH, 40 cm 2, 30 kd, 0.5 mm UFP-30-C-H24U HF Start AXH, 40 cm 2, 100 kd, 0.5 mm UFP-100-C-H24U HF Start AXH, 40 cm 2, 300 kd, 0.5 mm UFP-300-C-H24U HF Start AXH, 40 cm 2, 500 kd, 0.5 mm UFP-500-C-H24U Related literature Code Number UNICORN Control System Further information For the latest news and product information visit: or contact your local GE Healthcare representative. GE Healthcare Björkgatan Uppsala Sweden ÄKTAcrossflow, ÄKTAdesign, Fast Trak Validation, Kvick, and UNICORN are trademarks of GE Healthcare Limited a General Electric Company. All goods and services are sold subject to the terms and conditions of sale of the company within GE Healthcare which supplies them. GE Healthcare reserves the right, subject to any regulatory and contractual approval, if required, to make changes in specifications and features shown herein, or discontinue the product described at any time without notice or obligation General Electric Company All rights reserved. Amersham Biosciences AB, a General Electric Company going to market as GE Healthcare. GE Healthcare Amersham Biosciences AB Björkgatan 30, Uppsala, Sweden GE Healthcare Amersham Biosciences Europe GmbH Munzinger Strasse 9, D Freiburg, Germany GE Healthcare Amersham Biosciences UK Ltd Amersham Place, Little Chalfont, Buckinghamshire, HP7 9NA, UK GE Healthcare Amersham Biosciences Corp 800 Centennial Avenue, P.O. Box 1327, Piscataway, NJ , USA GE Healthcare Amersham Biosciences KK Sanken Bldg , Hyakunincho, Shinjuku-ku, Tokyo , Japan Asia Pacifi c Tel: Fax: Australasia Tel: Fax: Austria Tel: 01/ Fax: 01/ Belgium Tel: Fax: Canada Tel: Fax: Central, East, & South East Europe Tel: Fax: Denmark Tel: Fax: Finland & Baltics Tel: +358-(0) Fax: +358 (0) France Tel: Fax: Germany Tel: 0761/ Fax: 0761/ Italy Tel: Fax: Japan Tel: Fax: Latin America Tel: Fax: Middle East & Africa Tel: Fax: Netherlands Tel: Fax: Norway Tel: Fax: Portugal Tel: Fax: Russia & other C.I.S. & N.I.S Tel: +7 (095) , Fax: +7 (095) South East Asia Tel: Fax: Spain Tel: Fax: Sweden Tel: Fax: Switzerland Tel: Fax: UK Tel: Fax: USA Tel: Fax: imagination at work AB