Assume use of GE CM7 chip to maximize RU for small analyte and large ligand. Assume NHS/EDC and ethanolamine provided by GE. For optimal signal (~100 RU), determine the first appropriate amount of ligand to attach to SPR chip assuming one binding site on ligand Sec18 monomer (Sec18mon) 84,056 Da for one molecule analyte di-C8 phosphatidic acid (PA) MW 446.453 Da. Using formula: [Responsemax = (ResponseLigand × MassAnalyte)/MassLigand] you would need 18,827.5 RU of Sec18 cross-linked to dextran surface to yield Responsemax 100 assuming 1:1 binding of PA to Sec18mon.
Cm7 chip
Manufactured by GE Healthcare
The CM7 chip is a core component of GE Healthcare's medical imaging equipment. It is designed to process and analyze data from various imaging modalities, including computed tomography (CT), magnetic resonance imaging (MRI), and positron emission tomography (PET). The CM7 chip's primary function is to perform complex mathematical calculations and signal processing tasks necessary for the reconstruction and enhancement of medical images, enabling clinicians to make accurate diagnoses and treatment decisions.
Automatically generated - may contain errors
Lab products found in correlation
Vitronectin, by R&D Systems (4 mentions)
Fibronectin, by Roche (4 mentions)
Recombinant human integrin αvβ3, by R&D Systems (4 mentions)
Acetate buffer ph5, by GE Healthcare (3 mentions)
Biacore x100 system, by GE Healthcare (2 mentions)
Acetate buffer, by GE Healthcare (1 mentions)
Biacore t200 system, by GE Healthcare (1 mentions)
8 protocols using cm7 chip
1
Optimizing SPR Assay for PA-Sec18 Interaction
2
Surface Plasmon Resonance Analysis of MEST
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The experiment was performed as previously described.26 (link) SPR analysis was performed using the Biacore X100 system (GE Healthcare Life Sciences, Marlborough, MA, USA). The MEST protein was immobilized by amine coupling onto flow cell 2 of a CM7 chip (GE Healthcare Life Sciences). Following immobilization, the chip was washed for 30 min with PBS buffer. Small molecules in PBS buffer were passed over the chip at 30 μl/min for 90 s at 25 °C.
3
Integrin αVβ3 Binding Assay
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Example 13
Recombinant human integrin αVβ3 (R&D Systems, Minneapolis Minn.) was diluted to 40 ug/mL in Acetate buffer pH5.0 (GE Healthcare, Piscataway N.J.), and. then immobilized on a CM7 chip (GE Healthcare) using standard amine coupling techniques. 500 nM fibronectin (Roche Diagnostics, Indianapolis, Ind.) and vitronectin (R&D Systems) and 5 μM of either non-binding control 10Fn3 molecule (consisting of SEQ ID NO: 6 with an additional MG at the N-terminus and with a single amino acid substitution that changes RGD to RGE) or targeted 10Fn3 molecules (having a mutated FG loop that does not contain an RGD motif) were flowed over the top of the immobilized integrin. Binding RU was collected at the end of the sample injection. The results indicate that the lack of RGD in the FG loop results in abolishing binding of 10Fn3 molecules to fibronectin and vitronectin.
4
Integrin αVβ3 Binding Inhibition
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Example 13
Recombinant human integrin αVβ3 (R&D Systems, Minneapolis Minn.) was diluted to 40 ug/mL in Acetate buffer pH5.0 (GE Healthcare, Piscataway N.J.), and. then immobilized on a CM7 chip (GE Healthcare) using standard amine coupling techniques. 500 nM fibronectin (Roche Diagnostics, Indianapolis, Ind.) and vitronectin (R&D Systems) and 5 μM of either non-binding control 10Fn3 molecule (consisting of SEQ ID NO: 6 with an additional MG at the N-terminus and with a single amino acid substitution that changes RGD to RGE) or targeted 10Fn3 molecules (having a mutated FG loop that does not contain an RGD motif) were flowed over the top of the immobilized integrin. Binding RU was collected at the end of the sample injection. The results indicate that the lack of RGD in the FG loop results in abolishing binding of 10Fn3 molecules to fibronectin and vitronectin.
5
Integrin αVβ3 Binding Characterization
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Example 13
Recombinant human integrin αVβ3 (R&D Systems, Minneapolis Minn.) was diluted to 40 ug/mL in Acetate buffer pH5.0 (GE Healthcare, Piscataway N.J.), and. then immobilized on a CM7 chip (GE Healthcare) using standard amine coupling techniques. 500 nM fibronectin (Roche Diagnostics, Indianapolis, Ind.) and vitronectin (R&D Systems) and 5 μM of either non-binding control 10Fn3 molecule (consisting of SEQ ID NO: 6 with an additional MG at the N-terminus and with a single amino acid substitution that changes RGD to RGE) or targeted 10Fn3 molecules (having a mutated FG loop that does not contain an RGD motif) were flowed over the top of the immobilized integrin. Binding RU was collected at the end of the sample injection. The results indicate that the lack of RGD in the FG loop results in abolishing binding of 10Fn3 molecules to fibronectin and vitronectin.
6
Binding of 10Fn3 Molecules to Integrin
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Example 13
Recombinant human integrin αVβ3 (R&D Systems, Minneapolis Minn.) was diluted to 40 ug/mL in Acetate buffer pH 5.0 (GE Healthcare, Piscataway N.J.), and. then immobilized on a CM7 chip (GE Healthcare) using standard amine coupling techniques. 500 nM fibronectin (Roche Diagnostics, Indianapolis, Ind.) and vitronectin (R&D Systems) and 5 μM of either non-binding control 10Fn3 molecule (consisting of SEQ ID NO: 6 with an additional MG at the N-terminus and with a single amino acid substitution that changes RGD to RGE) or targeted 10Fn3 molecules (having a mutated FG loop that does not contain an RGD motif) were flowed over the top of the immobilized integrin. Binding RU was collected at the end of the sample injection. The results indicate that the lack of RGD in the FG loop results in abolishing binding of 10Fn3 molecules to fibronectin and vitronectin.
7
Investigating IGPD-Azithromycin Binding Kinetics
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Surface plasmon resonance method was used to further investigate the relationship between IGPD and azithromycin. The sensor surfaces were set up using the Biacore T200 system (GE Healthcare). EDC-NHS (70 μL) was used to activate the surface (CM7 chip; GE Healthcare) at a flow rate of 0.5 mL/min, IGPD was diluted to 30 μg/mL in 10 mM sodium acetate and immobilized in a flow cell on a sensor chip. In addition, 1 M ethanolamine was used to block the surface-activated groups after immobilizing IGPD. The running buffer used for immobilization contained 900 mL of 1.1× PBS and 100 mL of 100% methanol (pH 7.4). Azithromycin (320 nM) was injected (100 μL) at a flow rate of 2 mL/min. In addition, positive control (with ceftiofur) and control (with the mobile phase) were included. All experiments were performed at 25°C.
8
Probing Azelastine-ARF1 Interactions
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The interaction between azelastine and ARF1 was analyzed with a Biacore X100 system (GE Healthcare Life Sciences). In brief, the purified protein solution was adjusted to an appropriate pH value with acetic acid. ARF1 was coupled with CM7 chip (GE Healthcare Life Sciences), which was pre-balanced with PBS containing 0.4% P20 according to the manufacturer's instructions. Different concentrations of azelastine (150, 75, 37.5, 18.75, and 9.375 μM) were diluted with running buffer, and the samples were loaded to detect the response values. Biacore analysis software was used to fit the curve, and the Kd value of the small molecule and the protein was obtained.
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