AI Article Synopsis
- The study explored using protein A affinity precipitation as a new method for purifying antibodies, specifically human immunoglobulin G (hIgG), instead of traditional techniques.
- The researchers linked recombinant protein A to a pH-responsive polymer called Eudragit S-100, achieving a high binding capacity and rapid adsorption of hIgG.
- The results showed a successful purification process with a 65% yield, 96% purity, and the potential for reusability, suggesting this method could be a viable alternative to protein A chromatography.
Article Abstract
The potential of protein A affinity precipitation as an alternative method for traditional antibody purification techniques was investigated. Recombinant produced protein A from Staphylococcus aureus (SpA) was covalently linked to the pH-responsive copolymer Eudragit(®) S-100 and used for purification of human immunoglobulin G (hIgG). The Eudragit-SpA conjugate had a static binding capacity of 93.9 ± 2.8 mg hIgG per g conjugate and a dissociation constant of 787 ± 67 nM at 7 ± 1°C. The antibody was adsorbed rapidly onto Eudragit-SpA and reached equilibrium within 5 min. An excess of hIgG binding sites, provided by the conjugate, as well as adjusted elution conditions resulted in an appropriate hIgG purification performance. In summary, Eudragit-SpA was successfully applied to capture hIgG from a protein mixture with 65% antibody yield in the elution step. Nearly 96% purity and a purification factor of 12.4 were achieved. The Eudragit-SpA conjugate showed a stable ligand density over several cycles, which enabled reusability for repeated precipitation of hIgG. According to this, pH induced affinity precipitation can be seen as a potential alternative for protein A chromatography in antibody purification processes.
Download full-text PDF |
Source |
|---|---|
| http://dx.doi.org/10.1016/j.jchromb.2014.06.011 | DOI Listing |
Publication Analysis
Top Keywords
Similar Publications
Competing addition processes give distinct growth regimes in the assembly of 1D filaments.
Biophys J
January 2025
Department of Physics, Kansas State University, Manhattan, KS 66506, USA. Electronic address:
We present a model to describe the concentration-dependent growth of protein filaments. Our model contains two states, a low entropy/high affinity ordered state and a high entropy/low affinity disordered state. Consistent with experiments, our model shows a diffusion-limited linear growth regime at low concentration, followed by a concentration-independent plateau at intermediate concentrations, and rapid disordered precipitation at the highest concentrations.
View Article and Find Full Text PDFLarge library docking of tangible molecules has revealed potent ligands across many targets. While make-on-demand libraries now exceed 75 billion enumerated molecules, their synthetic routes are dominated by a few reaction types, reducing diversity and inevitably leaving many interesting bioactive-like chemotypes unexplored. Here, we investigate the large-scale enumeration and targeted docking of isoquinuclidines.
View Article and Find Full Text PDF[A novel carbonyl reductase for the synthesis of ()-tolvaptan].
Sheng Wu Gong Cheng Xue Bao
January 2025
Key Laboratory of Medicinal Chemistry and Molecular Diagnosis of the Ministry of Education, College of Chemistry and Materials Sciences, Hebei University, Baoding 071002, Hebei, China.
Screening carbonyl reductases with the ability to catalyze the reduction of complex carbonyl compounds is of great significance for the biosynthesis of -tolvaptan(-TVP). In this study, the target carbonyl reductase in the crude enzyme extract of rabbit liver was separated, purified, and identified by ammonium sulfate precipitation, gel-filtration chromatography, ion exchange chromatography, affinity chromatography, and protein mass spectrometry. With the rabbit liver genome as the template, the gene encoding the carbonyl reductase was amplified by PCR and the recombinant strain was successfully constructed.
View Article and Find Full Text PDFPreparation of molecularly imprinted polymer nanoparticles for the extraction and purification of progesterone combined with UPLC-ESI-MS mass spectrometry detection in royal jelly.
J Chromatogr B Analyt Technol Biomed Life Sci
January 2025
Jilin Ginseng Academy, Changchun University of Chinese Medicine, Changchun 130117 China. Electronic address:
This study aimed to develop molecularly imprinted polymer (MIP) nanoparticles specifically for the selective extraction and enrichment of progesterone (P) from royal jelly (RJ), and quantitatively analyzed them by ultra-performance-liquid chromatography electrospray ionization mass spectrometry (UPLC-ESI-MS). Gaussian software-based theoretical calculations identified methacrylic acid (MAA) as the optimal functional monomer for its strong binding affinity to P. MIP was synthesized by precipitation polymerization, and the preparation process of MIP was optimized by one-way variance design and response surface methodology.
View Article and Find Full Text PDFNi-induced selective precipitation of His-tagged recombinant proteins shortens purification time while maintaining high yield.
J Biotechnol
January 2025
Department of Biotechnology and Life Science, Faculty of Engineering, Tokyo University of Agriculture and Technology, 2-24-16 Nakamachi, Koganei-shi, Tokyo 184-8588, Japan; Institute of Global Innovation Research, Tokyo University of Agriculture and Technology, 3-8-1 Harumi-cho, Fuchu-shi, Tokyo 183-8538, Japan. Electronic address:
Nickel-NTA affinity chromatography is the current standard method for purifying His-tagged recombinant proteins. However, this process involves repetitive tasks, can be time-consuming, and reduces protein yield. Here, we present a simple, fast, and handy method for purifying His-tagged proteins using free Ni²⁺.
View Article and Find Full Text PDFWant AI Summaries of new PubMed Abstracts delivered to your In-box?
Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!