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Open accessJournal ArticleDOI: 10.1021/ACS.ANALCHEM.0C04974

Microdroplet Ultrafast Reactions Speed Antibody Characterization.

02 Mar 2021-Analytical Chemistry (American Chemical Society (ACS))-Vol. 93, Iss: 8, pp 3997-4005
Abstract: Recently, microdroplet reactions have aroused much interest because the microdroplet provides a unique medium where organic reactions could be accelerated by a factor of 103 or more. However, microdroplet reactions of proteins have been rarely studied. We report the occurrence of multiple-step reactions of a large protein, specifically, the digestion, reduction, and deglycosylation of an intact antibody, which can take place in microseconds with high reaction yields in aqueous microdroplets at room temperature. As a result, fast structural characterization of a monoclonal antibody, essential for assessing its quality as a therapeutic drug, can be enabled. We found that the IgG1 antibody can be digested completely by the IdeS protease in aqueous microdroplets in 250 microseconds, a 7.5 million-fold improvement in speed in comparison to traditional digestion in bulk solution (>30 min). Strikingly, inclusion of the reductant tris(2-carboxyethyl)phosphine in the spray solution caused simultaneous antibody digestion and disulfide bond reduction. Digested and reduced antibody fragments were either collected or analyzed online by mass spectrometry. Further addition of PNGase F glycosylase into the spray solution led to antibody deglycosylation, thereby producing reduced and deglycosylated fragments of analytical importance. In addition, glycated fragments of IgG1 derived from glucose modification were identified rapidly with this ultrafast digestion/reduction technique. We suggest that microdroplets can serve as powerful microreactors for both exploring large-molecule reactions and speeding their structural analyses.

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10 results found

Open accessPosted Content
Abstract: We investigate the solvation of methane and methane decorated with charges in spherically confined water droplets. Free energy profiles for a single methane molecule in droplets, ranging in diameter D, from 1 to 4 nm, show that the droplet surfaces are strongly favorable as compared to the interior. From the temperature dependence of the free energy in D=3 nm, we show that this effect is entropically driven. The potentials of mean force (PMFs) between two methane molecules show that the solvent separated minimum in the bulk is completely absent in confined water, independent of the droplet size since the solute particles are primarily associated with the droplet surface. The tendency of methanes with charges (Mq+ and Mq- with q+ = q- = 0.4e, where e is the electronic charge) to be pinned at the surface depends dramatically on the size of the water droplet. When D=4 nm, the ions prefer the interior whereas for D<4 nm the ions are localized at the surface, but with much less tendency than for methanes. Increasing the ion charge to e makes the surface strongly unfavorable. Reflecting the charge asymmetry of the water molecule, negative ions have a stronger preference for the surface compared to positive ions of the same charge magnitude. With increasing droplet size, the PMFs between Mq+ and Mq- show decreasing influence of the boundary due to the reduced tendency for surface solvation. We also show that as the solute charge density decreases the surface becomes less unfavorable. The implications of our results for the folding of proteins in confined spaces are outlined.

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Topics: Solvation (54%), Charge density (53%), Ion (51%)

48 Citations

Journal ArticleDOI: 10.1063/5.0057353
16 Jul 2021-Physics of Fluids
Abstract: Using micro-particle image velocimetry (μPIV), the convective flow inside a silicone oil droplet was investigated in detail during its formation in coaxial capillaries under co-flow in a water/glycerol mixture continuous phase. The analysis of μPIV measured flow field revealed that two characteristic flow areas exist in the droplet in formation: an inflow zone and a circulation zone. The intensity of vortex flow in these zones was estimated by calculating the average angular velocity of these vortices under the condition of no shear for different dispersed phase and continuous phase flow rates and for different viscosity ratios between the two phases. The evolution of the vortex flow pattern inside the droplet was investigated thoroughly all the way from the step of their formation to the step of the free-moving droplet. The results of this study are important for understanding the mixing processes inside the droplet at different stages of its formation.

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Topics: Vortex (56%)

4 Citations

Journal ArticleDOI: 10.1021/ACS.ANALCHEM.1C01011
Zhengzhou Li1, Yuze Li1, Lingpeng Zhan, Lingwei Meng1  +4 moreInstitutions (1)
Abstract: Volatile organic compounds (VOCs) from exhaled breath (EB) are considered to be promising biomarkers for lung diseases. A convenient and sensitive point-of-care (POC) testing method for EB VOCs is essential. Here, we developed a POC test paper for the analysis of EB aldehydes, which are potential biomarkers for lung cancer. A probe molecule, 4-aminothiophenol (4-ATP), was anchored on a paper substrate to specifically capture gas-phase aldehydes through the Schiff base reaction. Meanwhile, thin-film reaction acceleration was utilized to increase capture efficiency. By directly coupling the test paper to a mass spectrometer through paper spray, high sensitivity (0.1 ppt) and a wide quantification linear range (from 10 ppt to 1 ppm) were obtained. Analysis of EB from lung cancer patients with the test paper showed a significant increase in several reported aldehyde markers compared to EB from healthy volunteers, indicating the potential of this method for sensitive, low-cost, and convenient lung cancer screening and diagnosis.

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2 Citations

Journal ArticleDOI: 10.1016/J.IJMS.2021.116639
Xin Yan1Institutions (1)
Abstract: Recent studies have shown that remarkable reaction acceleration can be achieved at the air/solution interface of microdroplets. The unique environment of microdroplets can drive reactions that are thermodynamically unfavorable in bulk. These features allow microdroplet chemistry to impact many fields including chemical derivatization, reaction mechanistic studies, high-throughput reaction screening, and rapid small-scale, green and sustainable synthesis. This minireview describes the unique reactivities in microdroplets and acceleration mechanisms with examples of experimental and theoretical studies. Various methods for scale-up of microdroplet reactions and the accelerated derivatization for in-situ molecular analysis are also discussed.

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2 Citations

Journal ArticleDOI: 10.1002/CPLU.202100373
Lingqi Qiu1, Zhenwei Wei2, Zhenwei Wei1, Honggang Nie3  +2 moreInstitutions (3)
01 Oct 2021-ChemPlusChem
Abstract: The kinetics of organic reactions of different types in microvolumes (droplets, thin films, and sealed tubes) show effects of gas/solution interfacial area, reaction molecularity and solvent polarity. Partial solvation at the gas/solution interface is a major contributor to the 104 -fold reaction acceleration seen in bimolecular but not unimolecular reactions in microdroplets. Reaction acceleration can be used to manipulate selectivity by solvent choice.

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Topics: Molecularity (59%), Solvation (58%), Organic reaction (53%)


71 results found

Journal ArticleDOI: 10.1038/NRD2804
Roy Jefferis1Institutions (1)
Abstract: To date, more than 20 recombinant immunoglobulin G (IgG) antibody therapeutics are licensed for the treatment of various diseases The mechanism of action of recombinant monoclonal antibodies (rMAbs) has been extensively investigated and several distinct pathways have been defined; selective activation of specific pathways may optimize clinical outcomes for different diseases, such as cancer and chronic inflammation Human IgG is a glycoprotein with oligosaccharides attached at a single site These are essential to the mode of action of rMAbs, and the antibody efficacy can vary depending on the particular oligosaccharide that is attached Methods are now becoming available that allow the production of rMAbs bearing pre-selected oligosaccharides - glycoforms - to provide maximum efficacy for a given disease indication This Review summarizes current knowledge of these methods and avenues for their exploitation in the clinic

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Topics: Immunoglobulin G (52%), Monoclonal antibody (51%)

733 Citations

Journal ArticleDOI: 10.1039/B602614A
Abstract: On-line droplet–droplet extraction occurs when a sample spray intersects a reagent electrospray; this allows continuous analysis of trace amounts of compounds directly in complex matrices including undiluted urine, milk and polluted water over extended periods of time.

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409 Citations

Journal ArticleDOI: 10.1038/NRD.2017.227
Paul Carter1, Greg A. Lazar1Institutions (1)
Abstract: Antibody therapeutics are now established as a major drug class. Here, Carter and Lazar comprehensively discuss current and emerging platforms and technologies for antibody therapeutics, with an emphasis on approaches that could extend their therapeutic applications, including antibody–drug conjugates, bispecific antibodies and antibody engineering to enable more effective delivery.

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375 Citations

Journal ArticleDOI: 10.1146/ANNUREV.MED.59.090506.155819
Janet Woodcock1, Raymond WoosleyInstitutions (1)
Abstract: Societal expectations about drug safety and efficacy are rising while productivity in the pharmaceutical industry is falling. In 2004, the US Food and Drug Administration introduced the Critical Path Initiative with the intent of modernizing drug development by incorporating recent scientific advances, such as genomics and advanced imaging technologies, into the process. An important part of the initiative is the use of public-private partnerships and consortia to accomplish the needed research. This article explicates the reasoning behind the Critical Path Initiative and discusses examples of successful consortia.

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Topics: Drug development (51%)

366 Citations

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