Interaction of albumins and heparinoids investigated by affinity capillary electrophoresis and free flow electrophoresis

Mozafari, Mona; Deeb, Sami El; Krull, Friederike; Wildgruber, Robert; Weber, Gerhard; Reiter, Christian; Wätzig, Hermann

doi:10.1002/elps.201700202

Cited by 14 publications

(8 citation statements)

References 40 publications

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“…Mozafari et al. analyzed the interaction between human serum albumin (HSA) as well as bovine serum albumin (BSA) and PPS at 23 and 37°C, respectively . The raw data of the systems # 1.1–1.4 was provided by Dr. Mona Mozafari from Prof. Wätzig's group at TU Braunschweig.…”

Section: Methodsmentioning

confidence: 99%

Experimental design and measurement uncertainty in ligand binding studies by affinity capillary electrophoresis

et al. 2019

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Experimental design and measurement uncertainty in ligand binding studies by affinity capillary electrophoresisIn all life sciences ligand binding assays (LBAs) play a crucial role. Unfortunately these are very error prone. One part of this uncertainty results from the unavoidable random measurement uncertainty, another part can be attributed to the experimental design. To investigate the latter, uncertainty propagation was evaluated as a function of the given experimental design. A design space including the normalized maximum response range (nMRR), the data point position (DPP), the data point range (DPR) and the number of data points (NoDP) was defined. Based on ten measured ms ACE source data sets 20 specific parameter sets were selected by Design of Experiments. Monte Carlo simulations using 100 000 repeats for every parameter set were employed. The resulting measurement uncertainty propagation factors (measurement uncertainty multiplier: MUM) were used to describe the whole design space by polynomial regression. The resulting 5-dimensional response surface was investigated to evaluate the design parameter's influence and to find the minimal uncertainty propagation. It could be shown, that the nMRR is of highest importance, followed by DPP and DPR. Interestingly, the NoDP is less relevant. However, the interactions of the four parameters need to be carefully considered during design optimization. Using at least five data points which cover over 40% of the upper part of the binding hyperbola (DPP Ͼ 0.57) the MUM will be minimized (MUM approximately 1.5) when the nMRR is appropriate. It is possible to reduce the measurement uncertainty propagation more than one order of magnitude.Abbreviations: , electrophoretic mobility; c KD , theoretical mobility at the ligand concentration equals the K D -value; c , mobility of the complex; f , mobility of the free analyte; AIQ, analytical instrument qualification; aMRR, absolute maximum response range; BGE, background electrolyte; BSA, bovine serum albumin; c, concentration; c 0 , in absence of ligand; c Ý , in presence of a theoretically infinite ligand concentration; CD, sulfobutyl ether betacyclodextrine derivates; CI mean , confidence interval of the mean; const, constant parameter; conti, continuous parameters; DPP, data point position; DPR, data point range; DS, design space; E-systems, example analyte/ligand systems; G, Gradient; HC, hydrocorti-sone; HPAC, high performance affinity chromatography; HSA, human serum albumin; K A , association constant; K D , dissociation constant; LBA, ligand binding assay; LBAs, ligand binding assays; MRR, maximum response range; ms ACE, mobility shift affinity capillary electrophoresis; MUM, measurement uncertainty multiplier; MUP, measurement uncertainty propagation; nDS, normalized design space; nMRR, normalized maximum response range; NoDP, number of data points; PMS, parameter sets; PPS, pentosan polysulfate; R", complexed fraction; RSD, relative standard deviation; SOP, standard operating procedure; SPR, surface plasmon resona...

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Section: Methodsmentioning

confidence: 99%

Experimental design and measurement uncertainty in ligand binding studies by affinity capillary electrophoresis

et al. 2019

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“…Another study employed ms ACE to investigate the interactions of polysulfate sodium with HSA and BSA [106].…”

Section: Mobility Shift Acementioning

confidence: 99%

“…Anti-T 4 antibody [163] Insulin and glucagon Anti-insulin antibodies and anti-glucagon antibodies [169] Fluoroquinolones HSA [158] Mobility shift assay Tryptophan, warfarin, and quercetin HSA, BSA, and β-lactoglobulin [6] Catechins HSA [105] Heparinoids HSA, BSA [106] Enantioseparation and equilibrium constant measurements using proteins as buffer additives…”

Section: Direct Separation Anionic Carbohydrates and Oligonucleotidesmentioning

confidence: 99%

Analysis of drug interactions with serum proteins and related binding agents by affinity capillary electrophoresis: A review

et al. 2022

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Biomolecules such as serum proteins can interact with drugs in the body and influence their pharmaceutical effects. Specific and precise methods that analyze these interactions are critical for drug development or monitoring and for diagnostic purposes. Affinity capillary electrophoresis (ACE) is one technique that can be used to examine the binding between drugs and serum proteins, or other agents found in serum or blood. This article will review the basic principles of ACE, along with related affinity-based capillary electrophoresis (CE) methods, and examine recent developments that have occurred in this field as related to the characterization of drug-protein interactions. An overview will be given of the various formats that can be used in ACE and CE for such work, including the relative advantages or weaknesses of each approach. Various applications of ACE and affinity-based CE methods for the analysis of drug interactions with serum proteins and other binding agents will also be presented. Applications of ACE and related techniques that will be discussed include drug interaction studies with serum agents, chiral drug separations employing serum proteins, and the use of CE in hybrid methods to characterize drug binding with serum proteins.

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“…The interaction between chaperone HSP70 and co-chaperone BAG3, in the presence and absence of drug candidates, was observed through MCE fractionation of free and complex-bound HSP70, allowing high-throughput identification of hit compounds (Rauch et al 2013). MCE has recently been employed for the quantification of PPIs between serum albumins and heparinoids (Mozafari et al 2018), and for the characterisation of the binding interaction of an anti-lyzosyme antibody and dimerisation of HSP70 through a cross-linking approach (Ouimet et al 2016). MCE has also been used in the analysis of the PPI between actin-scavenger protein Gc-globulin and free actin (Pedersen et al 2008) (Fig.…”

Section: Analysis Of Ppis By Microchip Electrophoresismentioning

confidence: 99%

Microfluidic approaches for the analysis of protein–protein interactions in solution

et al. 2020

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Exploration and characterisation of the human proteome is a key objective enabling a heightened understanding of biological function, malfunction and pharmaceutical design. Since proteins typically exhibit their behaviour by binding to other proteins, the challenge of probing protein-protein interactions has been the focus of new and improved experimental approaches. Here, we review recently developed microfluidic techniques for the study and quantification of protein–protein interactions. We focus on methodologies that utilise the inherent strength of microfluidics for the control of mass transport on the micron scale, to facilitate surface and membrane-free interrogation and quantification of interacting proteins. Thus, the microfluidic tools described here provide the capability to yield insights on protein–protein interactions under physiological conditions. We first discuss the defining principles of microfluidics, and methods for the analysis of protein–protein interactions that utilise the diffusion-controlled mixing characteristic of fluids at the microscale. We then describe techniques that employ electrophoretic forces to manipulate and fractionate interacting protein systems for their biophysical characterisation, before discussing strategies that use microdroplet compartmentalisation for the analysis of protein interactions. We conclude by highlighting future directions for the field, such as the integration of microfluidic experiments into high-throughput workflows for the investigation of protein interaction networks.

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Interaction of albumins and heparinoids investigated by affinity capillary electrophoresis and free flow electrophoresis

Cited by 14 publications

References 40 publications

Experimental design and measurement uncertainty in ligand binding studies by affinity capillary electrophoresis

Experimental design and measurement uncertainty in ligand binding studies by affinity capillary electrophoresis

Analysis of drug interactions with serum proteins and related binding agents by affinity capillary electrophoresis: A review

Microfluidic approaches for the analysis of protein–protein interactions in solution

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