A novel method to evaluate protein solubility using a high throughput screening approach

Wiendahl, Matthias; Völker, Christiane; Husemann, Ilka; Krarup, Janus; Staby, Arne; Scholl, Stephan; Hubbuch, Jürgen

doi:10.1016/j.ces.2009.05.029

Cited by 25 publications

(19 citation statements)

References 53 publications

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“…In contrast to previous 96-well approaches used to determine protein solubility, where protein stocks are progressively diluted into precipitating buffers or solutions are slowly evaporated (Wiendahl et al, 2009), the protein concentration is here kept constant and the ammonium sulfate salt concentration incremented to obtain, for the first time in microplates, complete and accurate sigmoidal transition-curves for salt-induced reversible protein precipitation at equilibrium. The monomeric hen egg-white lysozyme (HEWL; EC 3.2.1.17) and tetrameric alcohol dehydrogenase (ADH; EC 1.1.1.1) from Saccharomyces cerevisiae, were used as two model proteins as they have been used in previous protein solubility studies (Curtis et al, 1998;Curtis and Lue, 2006;Richardson et al, 1990;Timasheff and Arakawa, 1988) and characterized extensively for structure, function, stability, and folding (Bachali et al, 2002;Clark, 1998;Dill, 1990;Dobson, 2003;Duester, 1996;Eklund et al, 1976;Mannall et al, 2006;Ramaswamy et al, 1994;Sarcar et al, 1992;Yoshimura et al, 1988).…”

Section: Introductionmentioning

confidence: 99%

Thermodynamic parameters for salt‐induced reversible protein precipitation from automated microscale experiments

Ahmad

Dalby

2010

Biotech & Bioengineering

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Reversible precipitation can be used as an efficient purification tool for proteins. In addition, identifying conditions under which precipitation or aggregation occurs is of key importance in the bioprocessing and pharmaceutical industry, as this can aid in better formulations and hinder aggregation in chromatography. We have evaluated the precipitation of proteins as determined by light scattering in microplates as a tool for the high-throughput determination of thermodynamic parameters for protein precipitation, with the potential for screening of formulation additives and relevant bioprocess conditions such as pH. This provides a useful complementary technique to existing microplate-based protein thermostability measurements. Using hen egg-white lysozyme and alcohol dehydrogenase as model proteins we have determined the extent of reversible precipitation as a function of ammonium sulfate and sodium chloride concentrations, and also demonstrated global fitting of the data to generate a model where the fraction precipitated can be predicted for any given condition. The global fit provided thermodynamic parameters, including the free energy for protein precipitation, and also allowed an approximate determination of the average size of the structural nucleus that contributes to the free energy of precipitation for each protein. The rapid collection of thermodynamic parameters for protein precipitation, in parallel with protein thermostability measurements, will provide a powerful platform for protein formulation, and also lead to datasets useful for testing theoretical predictions of reversible precipitation based on the molecular modeling of specific protein structure interactions.

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Section: Introductionmentioning

confidence: 99%

Thermodynamic parameters for salt‐induced reversible protein precipitation from automated microscale experiments

Ahmad

Dalby

2010

Biotech & Bioengineering

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“…The present method for determining the solubility curve is similar to the method employed by Kramarczyk et al . More detailed study on the solubility curve measurement with the 96‐well microplate system has been reported . Compared with the conventional method for measuring the solubility curve , the MP‐based method is easier and quicker .…”

Section: Discussionmentioning

confidence: 95%

High‐throughput process development methods for chromatography and precipitation of proteins: Advantages and precautions

Yoshimoto

Minakuchi

Itoh

et al. 2013

Engineering in Life Sciences

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Although high‐throughput process development methods using 96‐well microplate (MP) systems are promising and attractive, they still have several unclear problems in the practical applications. Several case studies for the purification process development by using manually operated 96‐well MP systems are presented. In the first example, the MPs with ion‐exchange monolith disks or ion‐exchange membranes were employed for linear gradient elution and breakthrough curve experiments. The data such as the peak salt concentration and dynamic binding capacity values obtained with the MP system were in good agreement with those by the LC system. In the second example, batch adsorption experiments with a 96‐well filter MP were performed for a protein A resin (particle)–antibody system in order to obtain the adsorption isotherms. The static binding capacities calculated from the isotherms were similar to those calculated from the LC system. In the last example, protein precipitation experiments by PEG were carried out for determining the solubility curve. It was found that the presence of dimer and aggregates affect the solubility curve measurement significantly. The proposed methods suggest powerful practical applications of high‐throughput process development for chromatography and precipitation process when the experiments are carefully designed and carried out.

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“…Different HT screening methods for investigating colloidal stability and protein phase behavior can be found in the literature. Wiendahl et al [95] developed an MTP-based method for protein precipitation based on fast liquid evaporation in micro-scale format. In case studies for lysozyme and insulin variants, systems of different protein and precipitant concentrations were pipetted on a fully automated liquid handling station.…”

Section: Ht Solubility Screeningsmentioning

confidence: 99%

Downstream process development strategies for effective bioprocesses: Trends, progress, and combinatorial approaches

Baumann

Hubbuch

2016

Engineering in Life Sciences

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The biopharmaceutical industry is at a turning point moving toward a more customized and patient-oriented medicine (precision medicine). Straightforward routines such as the antibody platform process are extended to production processes for a new portfolio of molecules. As a consequence, individual and tailored productions require generic approaches for a fast and dedicated purification process development. In this article, different effective strategies in biopharmaceutical purification process development are reviewed that can analogously be used for the new generation of antibodies. Conventional approaches based on heuristics and high-throughput process development are discussed and compared to modern technologies such as multivariate calibration and mechanistic modeling tools. Such approaches constitute a good foundation for fast and effective process development for new products and processes, but their full potential becomes obvious in a correlated combination. Thus, different combinatorial approaches are presented, which might become future directions in the biopharmaceutical industry.

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A novel method to evaluate protein solubility using a high throughput screening approach

Cited by 25 publications

References 53 publications

Thermodynamic parameters for salt‐induced reversible protein precipitation from automated microscale experiments

Thermodynamic parameters for salt‐induced reversible protein precipitation from automated microscale experiments

High‐throughput process development methods for chromatography and precipitation of proteins: Advantages and precautions

Downstream process development strategies for effective bioprocesses: Trends, progress, and combinatorial approaches

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