Downstream Protein Purification Unfortunately It Is Still Challenging for a User to Create a 100% Single-Use Process in Downstream Protein Purification

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turnover between processes. concerned about bioburden control concerned control about bioburden rather than sterility until the viral filtration step, the FDA still mandates that must sanitization standards be met. Single-use technologies solution to meet a practical are decrease these standards, contamination risk, costs, reduce speed and process increase

the target protein is harvested is harvested the target protein and purified. Single-use technologies have found their place during downstream a sterile providing purification protein without the environment processing cost and time of clean in place or steam in place sterilization (CIP/SIP). typically most although users are Downstream processing, specifically specifically processing, Downstream purification is protein downstream the final stage in bio-therapeutic manufacturing; the point when

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Heat Exchanger Heat Exchanger ™

single passsingle clarification, chromatography, chromatography, clarification, viral ultrafiltration/diafiltration, filtration and bulk drug substance equipment configuration equipment Heating/cooling efficienciesa in Heating/cooling Downstream purification protein Benefitsof using the DHX in harvest challenge that users must deal with challenge that users must deal with on a day-to-day basis. yields, decrease turnaround time and turnaround time and yields, decrease has never been greater cost reduce a presents and than it is today, yield and high quality products that that yield and high quality products for patient use. The pressure safe are on manufacturing facilities to increase precise control of process parameters parameters of process control precise high to ensure production throughout Introduction requires manufacturing Bio-therapeutic • • • and its applications: present the benefits of the benefits of present the Thermo Scientific DHX Scope: The purpose of Scope: The purpose of this technical note is to

Thermo DHX Heat Scientific Exchanger Integrate sterile and efficient temperature control with the with control andtemperature efficient sterile Integrate TECHNICAL NOTE TECHNICAL Downstream Protein Purification Unfortunately it is still challenging for a user to create a 100% single-use process in downstream protein purification. Several Typical downstream protein purification equipment configuration components of the manufacturing process including heat exchangers have not previously been available in a single-use format, requiring that users either insert stainless steel equipment into an otherwise single-use process flow, or use Figure 1. Typical downstream processing configuration. other single-use equipment such as Dashed boxes indicate where technical challenges exist. a jacketed mixing tote in place of a heat exchanger, a purpose for which While it is true that every step in the factor in determining the percent temperature control is not optimized. process generates heat that must be recovery from each pass. The process The ideal solution to this problem is to controlled, there are critical points solution and loading buffer must be at include a single-use heat exchanger where temperature control has a the same temperature during mixing at key points in the process to gain significant impact on the yield and and loading onto the column. Failure precise temperature control of the quality of final protein. to properly control the temperature of process solution without losing the either solution will lead to non-specific benefits of a single-use process Harvest Clarification binding and a reduced recovery. (see figure 1 to the right). Typically performed by means of centrifugation or filtration. Processing Ultrafiltration/Diafiltration Temperature control during large batch volumes through harvest Polishing via ultrafiltration/diafiltration downstream protein purification clarification can take a significant (UF/DF) is another critical step in Protein purification is technically period of time, which can lead to determining final yield. Recovery can challenging; proteins are heat labile, increased temperature. It is common be significantly improved through and most purification steps expose the practice to use a jacketed mixing proper temperature control of buffers process solution to unwanted heat. In tote or hold station after clarification and intermediates prior to loading, order to maximize quality and quantity and prior to column load to control and through rapid cooling of the of the final product, it is often desirable temperature. The DHX heat exchanger retentate or permeate immediately to cool the process solution after each provides an alternative solution with after filtration. step in order to prevent unwanted increased heat transfer rates, enabling binding or folding. more rapid cooling of the process Viral filtration and bulk drug solution, while maintaining the benefits substance Downstream processing is a of single-use. Final steps in downstream processing multi-step process that typically call for viral filtration and cooling of the includes the following steps: Chromatography bulk drug substance prior to freezing. Chromatography is one of the most Once viral filtration is complete, sterility 1. Harvest clarification to separate the important steps for determining final of the bulk drug substance is critical cells from media and cell debris yields. Proper optimization of the and single-use options for cooling 2. Chromatographic separation to chromatographic separation steps can expedite the freezing process. isolate and purify can have a significant impact on The DHX single-use heat exchanger 3. Ultrafiltration/diafiltration to further recovery rates. This is compounded provides an alternative; enabling sterile purify and concentrate by the fact that reaching the desired cooling in a single pass (depending on level of purity often necessitates temperature and flow rate of the bulk), 4. Viral filtration to leave a pure, clean, multiple rounds of chromatography. with heat transfer rates comparable to pharmaceutical grade product while there are always losses during a stainless steel heat exchanger, 5. Bulk drug substance freezing prior to chromatography (recovery is never while maintaining all the advantages shipment to Fill/Finish facility 100%), loading conditions are a critical of single-use. Given all the time and expense that has gone into getting the protein to this final stage it is critical to maximize the final yield. Proper set up of clarification, purification, and polishing techniques, and control of key parameters such as temperature, are all- important in the determination of final yield. The DHX single-use heat exchanger enables temperature Figure 2. Utilizing the DHX Heat Exchanger in downstream protein purification. control at critical points during downstream processing; increasing yields, driving down cost, reducing clean up and improving turnaround times of plant and equipment. • Reduced bioburden (See figure 2 to the right). The DHX heat exchanger creates a sterile, temperature controlled fluid Benefits path for process solution, buffers • Increased yields and any other process components. The DHX enables the user to The single-use benefits of the DHX accurately control the temperature addresses any weak points in the of their process solution before CIP or sanitization processes that and after critical filtration and typically arise when stainless steel chromatography steps without the equipment is used. need to incorporate stainless steel A plate and frame design promotes efficient equipment into the process flow. • Rapid turnaround of process heat transfer. Single-use BioProcess Precise temperature control is a key equipment Containers (BPCs) fit tightly between the determinant of recovery and yield, anytime stainless steel equipment stainless steel plates. especially during chromatography is used within a process, it creates and polishing steps. a requirement for time consuming CIP and sometimes SIP operations. • Reduced infrastructure These cleaning and sterilization requirements steps can increase the turnaround The DHX Heat Exchanger enables time from hours to days, locking up the user to cool or warm the process expensive capital equipment that fluid and buffers with the benefits could otherwise be put back into of single-use technology. The DHX service. The DHX Heat Exchanger eliminates the need for separate Serpentine flow paths of process fluid and addresses one critical point in the cold or warm storage rooms, which heating/cooling fluid maximize heat transfer. process where previously only greatly reduces the complexity and multi-use options were available. cost of the manufacturing facility requirements. Heat transfer rates in both heating and cooling mode are comparable with stainless steel heat exchangers

Table 1. Cooling efficiencies Table 2. Heating efficiencies DHX cooling efficiencies measured by temperature vs. time. DHX heating efficiencies measured by temperature vs. time. DHX COOLING EFFICIENCES DHX HEATING EFFICIENCIES 1 L/min 3 L/min 5 L/min 10 L/min 15 L/min 1 L/min 3 L/min 5 L/min 10 L/min 15 L/min 40 45

35 40 30 1 LPM 1 LPM 25 3 LPM 35 3 LPM Temp (oC) TEMPTemp (oC) (oC) TEMP (oC) 20 5 LPM 5 LPM 10 LPM 30 10 LPM 15 15 LPM 15 LPM 10 25 5 20 0 0 5 10 15 20 25 30 0 5 10 15 20 25 30 TimeTIME (min)(min) TimeTIME (min)(min)

Conclusion References There are several reasons to choose the DHX 1. State-of-the-art in downstream processing of monoclonal antibodies; process trends in design and validation. Marichal-gallardo, P.A., and Alvarez, M.M. Biotechnology Heat Exchanger, but the most compelling argument Progress. Volume 28, Issue 4. 899-916, June 2012 is the increased quality and improved yields that can 2. Single-use disposable technologies for biopharmaceutical manufacturing. Shukla A, be achieved by controlling the temperature of the process gottschalk U. Trends in Biotechnology, Volume 31, Issue 3, 147-154, 26 November 2012 solution before and after critical steps such as filtration and chromatography. The cost and time required to take a modern bio-therapeutic all the way through upstream and downstream processing is immense. Any tool or technique that can increase the final yield will positively impact the process; reducing cost and increasing profit.

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