Modeling hindered diffusion of antibodies in agarose beads considering pore size reduction due to adsorption

Hagemann, F; Adametz, Patrick; Weßling, Matthias; Thom, Volkmar

doi:10.1016/j.chroma.2020.461319

Cited by 10 publications

(2 citation statements)

References 43 publications

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“…Previous studies have shown that when agarose forms a gel, it has a spatial network structure and uniform pore size distribution, while the size of the pores is related to the agarose concentration. The higher the agarose concentration, the smaller the pore size [16]. However, we found an inconsistent result with it when different concentrations of agarose were observed under the electron microscope (ESM Fig.…”

Section: Discussionmentioning

confidence: 61%

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Low bias multiple displacement amplification with confinement effect based on agarose gel

et al. 2021

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Multiple displacement amplification (MDA) is a popular single-cell whole-genome amplification (WGA) technique that can greatly improve the amplification efficiency of single-cell genomes. However, there is an inherent problem that cannot be completely solved, that is, the amplification bias. We here propose an improved MDA method based on low melting agarose gel, named gelMDA. Firstly, the agarose gel and solution were characterized with SEM and fluorescent reagent. Then, we used gelMDA for cDNA amplification in library preparation of RNA-seq, and conventional MDA was used as a comparison. The sensitivity, efficiency of gelMDA, and amplification bias were evaluated with fluorescence curve, product yield, and the sequencing results. Finally, gelMDA was used for single-cell transcriptome sequencing. The results showed that the sensitivity and product yield of gelMDA were significantly higher than those of conventional MDA. A lower coefficient of variation (CV) and a higher reproducibility were obtained from gelMDA sequencing results. A region of 30 μm in diameter was amplified from the tissue sections and successfully sequenced. In conclusion, gelMDA obtained higher amplification efficiency and lower amplification bias in the present study. It suggested the great potential in single-cell RNA amplification and sequencing.

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Section: Discussionmentioning

confidence: 61%

“…1e, its pore size is mostly between 200 and 300 μm, which is similar to the hundred-microns space of a capillary. Agarose gel has a spatial network structure with uniform pore size distribution [16]. In the reaction system, there are only a few molecules in the space of about 40 nL units at a time.…”

Section: Discussionmentioning

confidence: 99%

Low bias multiple displacement amplification with confinement effect based on agarose gel

et al. 2021

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From protein structure to an optimized chromatographic capture step using multiscale modeling

Keulen,

Neijenhuis,

Lazopoulou

et al. 2024

Biotechnology Progress

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Optimizing a biopharmaceutical chromatographic purification process is currently the greatest challenge during process development. A lack of process understanding calls for extensive experimental efforts in pursuit of an optimal process. In silico techniques, such as mechanistic or data driven modeling, enhance the understanding, allowing more cost‐effective and time efficient process optimization. This work presents a modeling strategy integrating quantitative structure property relationship (QSPR) models and chromatographic mechanistic models (MM) to optimize a cation exchange (CEX) capture step, limiting experiments. In QSPR, structural characteristics obtained from the protein structure are used to describe physicochemical behavior. This QSPR information can be applied in MM to predict the chromatogram and optimize the entire process. To validate this approach, retention profiles of six proteins were determined experimentally from mixtures, at different pH (3.5, 4.3, 5.0, and 7.0). Four proteins at different pH's were used to train QSPR models predicting the retention volumes and characteristic charge, subsequently the equilibrium constant was determined. For an unseen protein knowing only the protein structure, the retention peak difference between the modeled and experimental peaks was 0.2% relative to the gradient length (60 column volume). Next, the CEX capture step was optimized, demonstrating a consistent result in both the experimental and QSPR‐based methods. The impact of model parameter confidence on the final optimization revealed two viable process conditions, one of which is similar to the optimization achieved using experimentally obtained parameters. The multiscale modeling approach reduces the required experimental effort by identification of initial process conditions, which can be optimized.

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Progress of ligand-modified agarose microspheres for protein isolation and purification

Qi,

Chen

2024

Microchim Acta

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Modeling hindered diffusion of antibodies in agarose beads considering pore size reduction due to adsorption

Cited by 10 publications

References 43 publications

Low bias multiple displacement amplification with confinement effect based on agarose gel

Low bias multiple displacement amplification with confinement effect based on agarose gel

From protein structure to an optimized chromatographic capture step using multiscale modeling

Progress of ligand-modified agarose microspheres for protein isolation and purification

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