Determination of Kinetic Data Using Surface Plasmon Resonance Biosensors

Hahnefeld, Claudia; Drewianka, Stephan; Herberg, Friedrich W.

doi:10.1385/1-59259-679-7:299

Cited by 36 publications

(56 citation statements)

References 40 publications

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“…RGS-(His) 4 antibody (Qiagen AG, Germany) was used to capture RGS-(His) 6 -tagged proteins on a CM5 sensor chip for subsequent binding analysis. The RGS-(His) 4 antibody was dialyzed for 1 h at 47C against 10 mM sodium acetate buffer, pH 5.0 and immobilized on three flow cells using EDC/NHS chemistry [7]. The antibody (30 mg/mL) was injected for 7 min at a flow rate of 5 mL/min over the NHS/ EDC-activated surface to generate surface densities of 6500-8500 RU.…”

Section: Conventional Setupmentioning

confidence: 99%

“…Nonspecific binding was subtracted using an activated and deactivated flow cell [7]. Double referencing, i.e., subtracting a blank run with running buffer after subtracting a reference cell, was applied to all curves.…”

Section: Conventional Setupmentioning

confidence: 99%

“…lized employing a specific capture approach [7], and analyzed with respect to their detailed binding kinetics using the complementary interaction partner in a microflow system.…”

Section: Generalmentioning

confidence: 99%

“…Repetitive immobilization and regeneration of anti-RGS-(His) 4 antibodies may reduce the biological activity. Therefore, control experiments had to be performed as shown before [7].…”

Section: Immobilization Strategiesmentioning

confidence: 99%

“…Recent technological developments like protein microarrays or biosensor analyses using surface plasmon resonance (SPR) allow protein-protein interactions to be characterized with high accuracy and with reasonable throughput [7,8]. With the Biacore technology binding partners can be detected in real time and separate kinetic parameters for the association and dissociation phases can be derived.…”

Section: Introductionmentioning

confidence: 99%

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Differential binding studies applying functional protein microarrays and surface plasmon resonance

et al. 2006

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A variety of different in vivo and in vitro technologies provide comprehensive insights in proteinprotein interaction networks. Here we demonstrate a novel approach to analyze, verify and quantify putative interactions between two members of the S100 protein family and 80 recombinant proteins derived from a proteome-wide protein expression library. Surface plasmon resonance (SPR) using Biacore technology and functional protein microarrays were used as two independent methods to study protein-protein interactions. With this combined approach we were able to detect nine calcium-dependent interactions between Arg-Gly-Ser-(RGS)-His 6 tagged proteins derived from the library and GST-tagged S100B and S100A6, respectively. For the protein microarray affinity-purified proteins from the expression library were spotted onto modified glass slides and probed with the S100 proteins. SPR experiments were performed in the same setup and in a vice-versa approach reversing analytes and ligands to determine distinct association and dissociation patterns of each positive interaction. Besides already known interaction partners, several novel binders were found independently with both detection methods, albeit analogous immobilization strategies had to be applied in both assays.

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Section: Conventional Setupmentioning

confidence: 99%

Section: Conventional Setupmentioning

confidence: 99%

“…lized employing a specific capture approach [7], and analyzed with respect to their detailed binding kinetics using the complementary interaction partner in a microflow system.…”

Section: Generalmentioning

confidence: 99%

“…Repetitive immobilization and regeneration of anti-RGS-(His) 4 antibodies may reduce the biological activity. Therefore, control experiments had to be performed as shown before [7].…”

Section: Immobilization Strategiesmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Differential binding studies applying functional protein microarrays and surface plasmon resonance

et al. 2006

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Applications of Lipid Nanodiscs for the Study of Membrane Proteins by Surface Plasmon Resonance

Trahey

Kwon

et al. 2015

CP Protein Science

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Methods for the initial steps of surface plasmon resonance analysis of membrane proteins incorporated in lipid nanodiscs are described. Several types of BiacoreTM sensor chips are available and require distinct strategies to immobilize proteonanodiscs on the chip surface. The procedures for immobilization on three of these chips (NTA, antibody coupled CM5, and L1) are described and results are demonstrated for a model system with cytochrome P4503A4 (CYP3A4) in nanodiscs binding to a polyclonal anti-CYP3A4 antibody. Advantages and disadvantages of each chip type are considered.

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Construction of an antimyoglobin single‐chain variable fragment with rapid reaction kinetics

Jang

Kim

Paek

et al. 2015

Biotech and App Biochem

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Antibodies with rapid reaction kinetics (high association and dissociation rates), named reversible antibodies, are used to perform continuous monitoring of sensitive disease biomarkers. In cases of acute myocardial infarction (AMI), continuous monitoring and early diagnosis are important. Human myoglobin (Myo) is a useful biomarker for AMI during the early stage after the onset of symptoms. In this study, a single-chain variable fragment (scFv) specific to Myo was derived from an IgG antibody that has rapid reaction kinetics. Enzyme-linked immunosorbent assay revealed that recombinant scFv exhibited 3.8-fold reduced affinity compared with the parent IgG antibody based on the antibody concentration necessary for 50% of the maximum signal. The scFv retained the rapid reaction kinetic mode with average kon and koff of 2.63 × 10(5) M(-1) Sec(-1) and 3.25 × 10(-3) Sec(-1) , respectively, which were reduced to 10- and 2.3-fold compared with those of the parent antibody. The equilibrium constant for the association of the scFv (KA = 8.09 × 10(7) M(-1) ) was 4.6-fold lower than that of its parent IgG antibody. This scFv may be a starting point for further mutagenesis/kinetic and structural analyses providing valuable insight into the mechanism of reversible antibodies.

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Determination of Kinetic Data Using Surface Plasmon Resonance Biosensors

Cited by 36 publications

References 40 publications

Differential binding studies applying functional protein microarrays and surface plasmon resonance

Differential binding studies applying functional protein microarrays and surface plasmon resonance

Applications of Lipid Nanodiscs for the Study of Membrane Proteins by Surface Plasmon Resonance

Construction of an antimyoglobin single‐chain variable fragment with rapid reaction kinetics

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