Characterization of murine anti-human Fab antibodies for use in an immunoassay for generic quantification of human Fab fragments in non-human serum samples including cynomolgus monkey samples

Stubenrauch, Kay; Wessels, Uwe; Essig, Ulrich; Kowalewsky, Frank; Vogel, Rudolf; Heinrich, Joachim

doi:10.1016/j.jpba.2012.08.023

Cited by 21 publications

(5 citation statements)

References 11 publications

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“…Serial aqueous humor samples were collected at 72, 168, and 336 h post‐dose. Serum, aqueous, and vitreous humor samples were analyzed for RG7716 concentration with an ELISA that uses anti‐idiotypic antibodies (Stubenrauch et al , ).…”

Section: Methodsmentioning

confidence: 99%

Targeting key angiogenic pathways with a bispecific Cross MA b optimized for neovascular eye diseases

Leithner²,

et al. 2016

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Anti‐angiogenic therapies using biological molecules that neutralize vascular endothelial growth factor‐A (VEGF‐A) have revolutionized treatment of retinal vascular diseases including age‐related macular degeneration (AMD). This study reports preclinical assessment of a strategy to enhance anti‐VEGF‐A monotherapy efficacy by targeting both VEGF‐A and angiopoietin‐2 (ANG‐2), a factor strongly upregulated in vitreous fluids of patients with retinal vascular disease and exerting some of its activities in concert with VEGF‐A. Simultaneous VEGF‐A and ANG‐2 inhibition was found to reduce vessel lesion number, permeability, retinal edema, and neuron loss more effectively than either agent alone in a spontaneous choroidal neovascularization (CNV) model. We describe the generation of a bispecific domain‐exchanged (crossed) monoclonal antibody (CrossMAb; RG7716) capable of binding, neutralizing, and depleting VEGF‐A and ANG‐2. RG7716 showed greater efficacy than anti‐VEGF‐A alone in a non‐human primate laser‐induced CNV model after intravitreal delivery. Modification of RG7716's FcRn and FcγR binding sites disabled the antibodies' Fc‐mediated effector functions. This resulted in increased systemic, but not ocular, clearance. These properties make RG7716 a potential next‐generation therapy for neovascular indications of the eye.

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Section: Methodsmentioning

confidence: 99%

Targeting key angiogenic pathways with a bispecific Cross MA b optimized for neovascular eye diseases

Leithner²,

et al. 2016

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“…Affinity (expressed in K D values) of interactions involving Atg8/LC3/GABARAPs is not so high in comparison with affinities of the strongest biointeractions (K D ~10 −6 nM for streptavidin:biotin binding [ 97 ], up to 10 −3 nM for antigen:antibody [ 98 ]). The K D values for interactions between the canonical p62/SQSTM1 LIR motif and all six human Atg8 proteins were measured by the isothermal titration calorimetry (ITC) experiments and are around 1 µM without any selectivity [ 92 ].…”

Section: Interactions Between Atg8/lc3/gabarap Proteins and Their mentioning

confidence: 99%

Atg8-Family Proteins—Structural Features and Molecular Interactions in Autophagy and Beyond

Wesch

Kirkin

Rogov

2020

Cells

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Autophagy is a common name for a number of catabolic processes, which keep the cellular homeostasis by removing damaged and dysfunctional intracellular components. Impairment or misbalance of autophagy can lead to various diseases, such as neurodegeneration, infection diseases, and cancer. A central axis of autophagy is formed along the interactions of autophagy modifiers (Atg8-family proteins) with a variety of their cellular counter partners. Besides autophagy, Atg8-proteins participate in many other pathways, among which membrane trafficking and neuronal signaling are the most known. Despite the fact that autophagy modifiers are well-studied, as the small globular proteins show similarity to ubiquitin on a structural level, the mechanism of their interactions are still not completely understood. A thorough analysis and classification of all known mechanisms of Atg8-protein interactions could shed light on their functioning and connect the pathways involving Atg8-proteins. In this review, we present our views of the key features of the Atg8-proteins and describe the basic principles of their recognition and binding by interaction partners. We discuss affinity and selectivity of their interactions as well as provide perspectives for discovery of new Atg8-interacting proteins and therapeutic approaches to tackle major human diseases.

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“…By comparison, antigen binding fragments (Fab) typically possess K d values in the pM–nM range ,. Monoclonal anti‐biotin antibodies have a K d value of 900 nM .…”

Section: Resultsmentioning

confidence: 99%

A Novel Assay Format as an Alternative to ELISA: MINA Test for Biotin

et al. 2018

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A novel abiotic assay based on biotin-specific fluorescent molecularly imprinted polymer nanoparticles (nanoMIPs) which acted as both reporter probes and binding agents, was developed. This is a first report of an assay which, unlike ELISA, required no washing steps or addition of enzyme substrates, making it more user-friendly. The components of the molecularly imprinted polymer nanoparticles assay (MINA) were assembled in microtiter plates fitted with magnetic inserts. The fluorescent nanoMIPs were bound to biotin-conjugated magnetic particles, which were attracted to the inserts. The addition of free biotin caused a displacement of the fluorescent nanoMIPs into solution, generating a signal proportional to the concentration of biotin. The nanoMIPs had a dissociation constant (Kd) of 14 nM, allowing the assay to detect biotin at nano-molar concentrations. The preassembled assay only required the addition of the sample and measurement of the fluorescence, and it functioned well after six weeks of storage without refrigeration. The assay did not show the susceptibility to several compounds which are known to interfere with avidin and streptavidin-based assays, such as mercaptoethanol and sugars. The protocols optimized in this work could be used to develop the abiotic assays for any other compound of interest.[a] S.

show abstract

Characterization of murine anti-human Fab antibodies for use in an immunoassay for generic quantification of human Fab fragments in non-human serum samples including cynomolgus monkey samples

Cited by 21 publications

References 11 publications

Targeting key angiogenic pathways with a bispecific Cross MA b optimized for neovascular eye diseases

Targeting key angiogenic pathways with a bispecific Cross MA b optimized for neovascular eye diseases

Atg8-Family Proteins—Structural Features and Molecular Interactions in Autophagy and Beyond

A Novel Assay Format as an Alternative to ELISA: MINA Test for Biotin

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