The Experts below are selected from a list of 288 Experts worldwide ranked by ideXlab platform
Andrew L. Zydney - One of the best experts on this subject based on the ideXlab platform.
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Single-use, single-pass tangential Flow Filtration using low-cost hollow fiber modules
Journal of Membrane Science, 2020Co-Authors: Christopher J. Yehl, Andrew L. ZydneyAbstract:Abstract Increasing pressures on biomanufacturing costs have led to renewed interest in the development of single-use technologies that can be readily adapted to continuous processing. The objective of this study was to use commercially available hollow fiber membranes, originally designed for high-flux hemodialysis, for single pass tangential Flow Filtration with high conversions. Experiments were performed with solutions of Immunoglobulin G (IgG) using polysulfone hollow fiber membrane cartridges with a surface area of 1.9 m2. The hollow fiber modules were able to provide more than 10-fold concentration of IgG in a single-pass with exit concentrations greater than 200 g/L. Stable operation was achieved during a continuous run for over a 120-h period with feed side pressure drops less than 1 kPa and transmembrane pressure drops less than 60 kPa. A single dialyzer was able to process 1.4 kg of IgG per day with a total cost of less than $0.004 per g of IgG. Flux-pressure profiles were in good qualitative agreement with a simple model accounting for concentration polarization effects and the variation of flux and pressure with axial position in the module. These results clearly demonstrate the potential of using these low-cost single-use hollow fiber modules for single-pass tangential Flow Filtration in continuous biomanufacturing.
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High Performance Tangential Flow Filtration Using Charged Affinity Ligands
Separation Science and Technology, 2007Co-Authors: Suma Rao, Kimberly Ager, Andrew L. ZydneyAbstract:Abstract High performance tangential Flow Filtration is an emerging technology for protein separations. The objective of this work was to examine the use of small charged affinity ligands to control the protein charge and thus extend the potential applicability of this separation technique. Experiments were performed using binary mixtures of bovine serum albumin and ovalbumin, with Cibacron Blue added as an affinity ligand. The addition of Cibacron Blue caused a large increase in the selectivity due to the increase in BSA retention associated with the strong electrostatic repulsion between the negatively‐charged protein‐ligand complex and the negatively‐charged cellulosic membrane. Appropriate conditions for the separation were identified using total recycle experiments in a small‐scale tangential Flow Filtration device. A diaFiltration process was designed to separate BSA from ovalbumin, with the Cibacron Blue recovered in a second stage operated with a buffer containing NaSCN to displace the dye from th...
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high performance tangential Flow Filtration using charged membranes
Journal of Membrane Science, 1999Co-Authors: R Van Reis, J M Brake, John Charkoudian, Douglas B Burns, Andrew L. ZydneyAbstract:Abstract High-performance tangential Flow Filtration (HPTFF) is an emerging technology that enables the separation of proteins with similar size. HPTFF technology has become possible by exploiting several new discoveries. It has been demonstrated that optimum selectivity and throughput are obtained in the pressure-dependent flux regime. Selectivity and throughput can also be enhanced through module design and process configurations that reduce the transmembrane pressure gradient. Optimization of buffer pH and ionic strength have a significant impact on the sieving behavior of proteins in membrane systems. Finally, a novel set of design equations and diagrams have been derived which provide a rational means for determining the optimum combination of selectivity and throughput for any given process. The current study focused on exploring the effects of membrane charge, in combination with buffer pH, on protein separation using HPTFF. Order-of-magnitude improvements in both selectivity and throughput were obtained by selecting the appropriate membrane charge at an optimum pH. The high selectivity and throughput values enabled protein purification using a small number of diavolumes ( N = 4–12), reasonable membrane area per mass of product (17 m 2 kg −1 ), and short processing times (1–3 h). Purification factors up to 990-fold were obtained with yields of 94%. These results were obtained in linear scale-down systems representative of existing industrial scale systems, currently in use for ultraFiltration of human pharmaceuticals produced by recombinant DNA methods.
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High performance tangential Flow Filtration.
Biotechnology and bioengineering, 1997Co-Authors: R. Van Reis, S Gadam, L.n. Frautschy, S Orlando, E.m. Goodrich, S. Saksena, Ralf Kuriyel, C M Simpson, Steven R. Pearl, Andrew L. ZydneyAbstract:Conventional tangential Flow Filtration (TFF) has traditionally been limited to separation of solutes that differ by about ten-fold in size. Wide pore-size distributions, membrane fouling, and concentration polarization phenomena have commonly been cited as reasons for this limitation. The use of TFF in the biotechnology industry has therefore been restricted to cell-protein, virus-protein, and protein-buffer separations. A multi-disciplinary team with industrial and academic members was formed to overcome these limitations and enable protein-protein separations using High Performance TFF (HPTFF) systems. Pore-size distributions have been improved with the development of new membrane formulation and casting techniques. Membrane fouling has been controlled by operating in the transmembrane pressure-dependent regime of the filtrate flux curve and by carefully controlling fluid dynamic start-up conditions. Concentration polarization was exploited to enhance, rather than limit, the resolution of solutes. Concentration polarization has also been controlled by operating a co-current filtrate stream that maintains transmembrane pressure constant along the length of the TFF module. High yields and purification factors were obtained even with small differences in protein sieving. IgG-BSA and BSA monomer-oligomer mixtures have successfully been separated with these systems. HPTFF technology provides a competitive purification tool to complement chromatographic processing of proteins.
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High performance tangential Flow Filtration
Biotechnology and Bioengineering, 1997Co-Authors: R. Van Reis, S Gadam, L.n. Frautschy, S Orlando, E.m. Goodrich, S. Saksena, Ralf Kuriyel, C M Simpson, Steven R. Pearl, Andrew L. ZydneyAbstract:Conventional tangential Flow Filtration (TFF) has traditionally been limited to separation of solutes that differ by about ten-fold in size. Wide pore-size distributions, membrane fouling, and concentration polarization phenomena have commonly been cited as reasons for this limitation. The use of TFF in the biotechnology industry has therefore been restricted to cell-protein, virus-protein, and protein-buffer separations. A multi-disciplinary team with industrial and academic members was formed to overcome these limitations and enable protein-protein separations using High Performance TFF (HPTFF) systems. Pore-size distributions have been improved with the development of new membrane formulation and casting techniques. Membrane fouling has been controlled by operating in the transmembrane pressure-dependent regime of the filtrate flux curve and by carefully controlling fluid dynamic start-up conditions. Concentration polarization was exploited to enhance, rather than limit, the resolution of solutes. Concentration polarization has also been controlled by operating a co-current filtrate stream that maintains transmembrane pressure constant along the length of the TFF module. High yields and purification factors were obtained even with small differences in protein sieving. IgG-BSA and BSA monomer-oligomer mixtures have successfully been separated with these systems. HPTFF technology provides a competitive purification tool to complement chromatographic processing of proteins. © 1997 John Wiley & Sons, Inc. Biotechnol Bioeng56: 71–82, 1997.
Lakshman Narasingha Rao Bhau - One of the best experts on this subject based on the ideXlab platform.
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Tangential Flow Filtration technology applicable to large scale recovery of diphtheria toxin.
Journal of bioscience and bioengineering, 2002Co-Authors: Bheeman Sundaran, Chitrambalam Palaniappan, Yarlagadda Udaya Bhaskara Rao, Ratnam Boopathy, Lakshman Narasingha Rao BhauAbstract:Abstract The commercial preparation of purified diphtheria toxoid primarily depends on the effective separation of the toxin from a large culture volume of Corynebacterium diphtheriae vaccine strain. Tangential Flow or cross-Flow Filtration is gaining momentum in the processing of biologicals. This study deals with the separation of toxins using a microporous membrane (0.45-μm pore size) with operational variables like transmembrane pressure (TMP), cross Flow rate and flux. Under the best conditions, ⪢98% recovery was achieved. The data and conclusions presented here have been drawn from process development work employing available equipment.
Andre F Palmer - One of the best experts on this subject based on the ideXlab platform.
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Tangential Flow Filtration of haptoglobin.
Biotechnology progress, 2020Co-Authors: Ivan S Pires, Andre F PalmerAbstract:Haptoglobin (Hp) is a plasma glycoprotein that scavenges cell-free hemoglobin (Hb). Hp has various potential therapeutic applications, but it has been mainly studied for treatment of acute hemolytic conditions that can arise from situations such as massive blood transfusion, infusion of stored red blood cells, severe burns, trauma, sepsis, radiation injury, and others. Therefore, Hp may also be beneficial during chronic hemolytic disease states such as hereditary spherocytosis, nocturnal hemoglobinuria, sickle-cell anemia, and malaria. Various methods have been developed to purify Hp from plasma or plasma fractions. However, none of these methods have exploited the large molecular weight (MW) range distribution of Hp polymers to easily isolate Hp from other plasma proteins. The present study used tangential Flow Filtration (TFF) to isolate polymeric Hp from plasma proteins using human Fraction IV (FIV) as the starting material. After removal of insoluble material from a suspension of FIV paste, the protein mixture was clarified on a 0.2 μm hollow fiber (HF) TFF filter. The clarified protein solution was then bracketed based on protein MW using HF filters with MW cut-offs (MWCOs) of 750, 500, and 100 kDa. Using untreated FIV, the Hp purity of the main bracket was ~75% with a total Hb binding capacity (HbBC) yield of 1.2 g starting from 500 g of FIV paste. However, pretreatment of FIV with fumed silica to remove lipoproteins increased Hp purity to >95% with a HbBC yield of 1.7 g per 500 g of FIV. Taken together this study provides a novel and scalable method to purify Hp from plasma or plasma fractions.
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purification of hemoglobin by tangential Flow Filtration with diaFiltration
Biotechnology Progress, 2009Co-Authors: Jacob Elmer, David R Harris, Andre F PalmerAbstract:A recent study by Palmer, Sun, and Harris (Biotechnol. Prog., 25:189–199, 2009) demonstrated that tangential Flow Filtration (TFF) can be used to produce HPLC-grade bovine and human hemoglobin (Hb). In this current study, we assessed the quality of bovine Hb (bHb) purified by introducing a 10 L batch-mode diaFiltration step to the previously mentioned TFF Hb purification process. The bHb was purified from bovine red blood cells (RBCs) by filtering clarified RBC lysate through 50 nm (stage I) and 500 kDa (stage II) hollow fiber (HF) membranes. The filtrate was then passed through a 100 kDa (stage III) HF membrane with or without an additional 10 L diaFiltration step to potentially remove additional small molecular weight impurities. Protein assays, SDS-PAGE, and LC-MS of the purified bHb (stage III retentate) reveal that addition of a diaFiltration step has no effect on bHb purity or yield; however, it does increase the methemoglobin level and oxygen affinity of purified bHb. Therefore, we conclude that no additional benefit is gained from diaFiltration at stage III and a three stage TFF process is sufficient to produce HPLC-grade bHb. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009
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purification of hemoglobin by tangential Flow Filtration with diaFiltration
Biotechnology Progress, 2009Co-Authors: Jacob Elmer, David R Harris, Guoyong Sun, Andre F PalmerAbstract:A recent study by Palmer, Sun, and Harris (Biotechnol. Prog., 25:189-199, 2009) demonstrated that tangential Flow Filtration (TFF) can be used to produce HPLC-grade bovine and human hemoglobin (Hb). In this current study, we assessed the quality of bovine Hb (bHb) purified by introducing a 10 L batch-mode diaFiltration step to the previously mentioned TFF Hb purification process. The bHb was purified from bovine red blood cells (RBCs) by filtering clarified RBC lysate through 50 nm (stage I) and 500 kDa (stage II) hollow fiber (HF) membranes. The filtrate was then passed through a 100 kDa (stage III) HF membrane with or without an additional 10 L diaFiltration step to potentially remove additional small molecular weight impurities. Protein assays, SDS-PAGE, and LC-MS of the purified bHb (stage III retentate) reveal that addition of a diaFiltration step has no effect on bHb purity or yield; however, it does increase the methemoglobin level and oxygen affinity of purified bHb. Therefore, we conclude that no additional benefit is gained from diaFiltration at stage III and a three stage TFF process is sufficient to produce HPLC-grade bHb.
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Tangential Flow Filtration of hemoglobin
Biotechnology progress, 2009Co-Authors: Andre F Palmer, Guoyong Sun, David R HarrisAbstract:Bovine and human hemoglobin (bHb and hHb, respectively) was purified from bovine and human red blood cells via tangential Flow Filtration (TFF) in four successive stages. TFF is a fast and simple method to purify Hb from RBCs using Filtration through hollow fiber (HF) membranes. Most of the Hb was retained in stage III (100 kDa HF membrane) and displayed methemoglobin levels less than 1%, yielding final concentrations of 318 and 300 mg/mL for bHb and hHb, respectively. Purified Hb exhibited much lower endotoxin levels than their respective RBCs. The purity of Hb was initially assessed via SDS-PAGE, and showed tiny impurity bands for the stage III retentate. The oxygen affinity (P(50)) and cooperativity coefficient (n) were regressed from the measured oxygen-RBC/Hb equilibrium curves of RBCs and purified Hb. These results suggest that TFF yielded oxygen affinities of bHb and hHb that are comparable to values in the literature. LC-MS was used to measure the molecular weight of the alpha (alpha) and beta (beta) globin chains of purified Hb. No impurity peaks were present in the HPLC chromatograms of purified Hb. The mass of the molecular ions corresponding to the alpha and beta globin chains agreed well with the calculated theoretical mass of the alpha- and beta- globin chains. Taken together, our results demonstrate that HPLC-grade Hb can be generated via TFF. In general, this method can be more broadly applied to purify Hb from any source of RBCs. This work is significant, since it outlines a simple method for generating Hb for synthesis and/or formulation of Hb-based oxygen carriers.
Bheeman Sundaran - One of the best experts on this subject based on the ideXlab platform.
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Tangential Flow Filtration technology applicable to large scale recovery of diphtheria toxin.
Journal of bioscience and bioengineering, 2002Co-Authors: Bheeman Sundaran, Chitrambalam Palaniappan, Yarlagadda Udaya Bhaskara Rao, Ratnam Boopathy, Lakshman Narasingha Rao BhauAbstract:Abstract The commercial preparation of purified diphtheria toxoid primarily depends on the effective separation of the toxin from a large culture volume of Corynebacterium diphtheriae vaccine strain. Tangential Flow or cross-Flow Filtration is gaining momentum in the processing of biologicals. This study deals with the separation of toxins using a microporous membrane (0.45-μm pore size) with operational variables like transmembrane pressure (TMP), cross Flow rate and flux. Under the best conditions, ⪢98% recovery was achieved. The data and conclusions presented here have been drawn from process development work employing available equipment.
György Marko-varga - One of the best experts on this subject based on the ideXlab platform.
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Characterization of a sampling unit based on tangential Flow Filtration for on-line bioprocess monitoring
Analytica Chimica Acta, 1993Co-Authors: Torbjörn Buttler, Lo Gorton, György Marko-vargaAbstract:The performance of a sampling unit (Waters Filter/Acquisition Module) based on tangential Flow Filtration, in conjuction with column liquid chromatography, is described. Filtration characteristics of this module were investigated. Two membrane types and pore sizes and their influence on the sampling were determined. The influence of feed rate on the Flow-rate of the filtrate for model solutions and real fermentation substrates and broths was investigated. Membrane recovery values for carbohydrates in solutions varying in complexity were determined. The contents of amino acids and organic acids in these substrates and broths and their recovery values are presented.