Activation of resin with controllable ligand density via catalytic oxa-Michael addition and application in antibody purification

Cheng, Fang; Li, Mingyang; He, Wei; Sun, Bingbing; Qin, Jinyan; Qü, Jingping

doi:10.1016/j.chroma.2018.07.032

Cited by 15 publications

(8 citation statements)

References 29 publications

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“…Our previous study showed that the traditional DVS activation method in alkaline aqueous solution leads to low reactivity and hydrolysis of the VS groups, resulting in a low surface VS density. 36 The zeta potentials of the Si NPs, PEG-Si NPs, and three kinds of VS-Si NPs were all negative, which is consistent with previous reports. 34,37 After anti-HER2 Fab-6His immobilization, the zeta potentials of all three VS-Si NPs became positive because the isoelectric point of anti-HER2 Fab-6His is estimated to be 8.5.…”

Section: ■ Results and Discussionsupporting

confidence: 91%

“…This indicates that the VS density can be adjusted by catalysts and reaction conditions, which is in good agreement with our previous study. 32,36 The Fab densities were assessed using the BCA method and were calculated according to eqs 2−4. According to Table 2, the Fab densities of the three NP samples are calculated to be 51.6 ± 17.2, 97.3 ± 1.9, and 181.3 ± 20.9 protein molecules per nanoparticle, respectively.…”

Section: ■ Results and Discussionmentioning

confidence: 99%

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Controlling Conjugated Antibodies at the Molecular Level for Active Targeting Nanoparticles toward HER2-Positive Cancer Cells

Dong

Cheng

et al. 2021

Mol. Pharmaceutics

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For active targeting nanodrug delivery systems conjugated with antibodies, both lack of control of the antibody at the molecular level and protein corona formation remarkably decreases targeting efficacy. Herein, we designed a series of silica nanoparticles toward HER2-positive breast cancer cells, with an anti-HER2 Fab-6His density ranging from 50 to 180 molecules per nanoparticle. Through the site-directed immobilization method we developed, the antigen-binding domain of anti-HER2 Fab was mostly accessible to the HER2 receptor. Both polyethylene glycol (PEG) chains and a high density of Fab were shown to suppress protein corona formation and macrophage uptake. The dependency of targeting efficacy and cytotoxicity on Fab density was shown using a series of breast cancer cell lines with different levels of the HER2 expression. The high density of Fab stimulates quick responses from HER2-positive cells. Combined with PEG chains, conjugated antibodies with a well-controlled orientation and density significantly improves delivery performance and sheds light on the design and preparation of an improved active targeting nanodrug delivery system.

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Section: ■ Results and Discussionsupporting

confidence: 91%

Section: ■ Results and Discussionmentioning

confidence: 99%

Controlling Conjugated Antibodies at the Molecular Level for Active Targeting Nanoparticles toward HER2-Positive Cancer Cells

Dong

Cheng

et al. 2021

Mol. Pharmaceutics

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“…Oligo(ethylene glycol) on the self-assembly monolayer surfaces provides the terminal hydroxyl group for conjugation while effectively reduces nonspecific protein adsorption . PPh 3 was selected as the catalyst, which could result in a high surface density of VS groups …”

Section: Resultsmentioning

confidence: 99%

“…28 PPh 3 was selected as the catalyst, which could result in a high surface density of VS groups. 29 The preparation of EG 6 -OH SAMs on the cleaned Aucoated substrates and the surface functionalization with DVS were characterized by XPS. The self-assembly and DVS functionalization are illustrated in Figure 1a.…”

Section: ■ Experimental Sectionmentioning

confidence: 99%

“…According to the HER2 + antigen-binding assay, this new immobilization method provides a much stronger binding affinity with lower LOD. Combined with the ease in the fabrication of VS-bearing surfaces on Au, 25 nanoparticles, 30 and chromatographic resins, 29 our new VS immobilization method paved the way for improving the sensitivity of biosensors and performance of biocatalysts.…”

Section: ■ Conclusionmentioning

confidence: 99%

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Site-Specific and Covalent Immobilization of His-Tagged Proteins via Surface Vinyl Sulfone–Imidazole Coupling

Cheng

Li³

et al. 2019

Langmuir

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Site-specific immobilization of proteins on a surface has been a long-lasting challenge in the fields of biosensing and biotechnology because of the need for improving the biological activity of immobilized protein via the maximal exposure of its bioactive domain. Herein, we reported a new site-specific immobilization method for His-tagged proteins onto a vinyl sulfone (VS)-bearing surface in a covalent manner. X-ray photoelectron spectroscopy characterization indicated the specificity of the addition reaction of the imidazole group in histidine on the VS-bearing surface at pH 7.0. Solution-based experiments were carried out to verify the reaction priority of the imidazole residue of histidine with the VS group at neutral conditions. The real-time immobilization process of two His-tagged proteins (HaloTag-6His and anti-HER2 Fab-6His) on surfaces presenting VS, preactivated carboxyl, and NTA groups were studied by quartz crystal microbalance. Compared to the existing methods utilizing covalent (NHS/EDC activated carboxyl) and coordinate (Ni2+-NTA) linking, our method offers two significant advantages for protein immobilization: high density and high specificity. The orientation of the two His-tagged proteins on the VS-bearing surface was confirmed by an enzyme-linked assay and an HER2+ liposome binding experiment. Our method of site-specific immobilization of His-tagged proteins is efficient and straightforward, which would be helpful to expand the applications of recombinant proteins in enzyme immobilization, biosensor and array fabrication, and drug delivery system.

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Divinyl Sulfone

Lucchi¹,

Fabbri²,

Santoyo‐González³

et al. 2021

Encyclopedia of Reagents for Organic Synthesis

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Activation of resin with controllable ligand density via catalytic oxa-Michael addition and application in antibody purification

Cited by 15 publications

References 29 publications

Controlling Conjugated Antibodies at the Molecular Level for Active Targeting Nanoparticles toward HER2-Positive Cancer Cells

Controlling Conjugated Antibodies at the Molecular Level for Active Targeting Nanoparticles toward HER2-Positive Cancer Cells

Site-Specific and Covalent Immobilization of His-Tagged Proteins via Surface Vinyl Sulfone–Imidazole Coupling

Divinyl Sulfone

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