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Micro-Volume Determination of Labeling Efficiency Using Spectral Scanning Analysis in a Standard Microplate Reader

Agilent Technologies
12 Feb 2013

The use of labeled reagents in a variety of experimental methods such as immunofluorescence, TR-FRET, and ELISA has been shown to provide substantial improvements in assay sensitivity while allowing sample and reagent conservation in a high-throughput context. This application note describes the analysis of protein-antibody conjugates labeled with biotin or digoxigenin using micro-volume spectral-scanning analysis in a microplate reader.

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Agilent BioTek Gen5 Software for Detection

Agilent Technologies

Agilent BioTek Gen5 software for Agilent BioTek multimode and single-mode microplate readers is an integrated tool for endpoint, kinetic, spectral scanning, and well area scanning. It controls all the functions of the plate reader and has powerful data analysis capabilities for a broad range of applications. Gen5 microplate reader software is also used to integrate BioTek plate readers to BioTek BioStack and other automated systems.

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ELISAEnzyme-linked immunosorbent assays (ELISA), also known as enzyme immunoassays (EIA), are used for the detection and quantification of proteins, peptides and antibodies in a sample. ELISA often comes in kit format, with pre-selected antibody pairs to detect specific proteins or biomarkers of diseases such as diabetes and obesity, or cardiovascular and neurological disorders. Detection is made possible with chemiluminescent, fluorescent or colorimetric substrates and detection instruments such as microplate / ELISA readers. Other kits available include: Enzyme-linked immunospot (ELISPOT) assays for the quantification of protein-producing cells and chemiluminescent immunoassays (CLIA) kits. Find the best ELISA kits and products in our peer-reviewed product directory: compare products, check customer reviews and receive pricing direct from manufacturers.Microplate Readers / DetectorsMicroplate readers are used to automate the detection and analysis of labeled or label-free components in microplates during assays or live-cell monitoring. Microplate readers are generally distinguished by their mode of detection. Types include absorbance, luminescence, fluorescence intensity, fluorescence polarization, TRF / FRET and multimode microplate readers. Microplate readers deliver a high throughput of samples by reading multiple wells simultaneously, with the 96-well format the most commonly used. As a result, microplate readers are often used in the drug discovery, bioassays, research and pharmaceutical industries for screening applications. Microplate loading can also be automated, with robotic microplate stackers to increase throughput. Find the best microplate readers in our peer-reviewed product directory: compare products, check customer reviews and receive pricing direct from manufacturers.Cell-Based AssaysCell-based assays are used to monitor the presence, quantity and activities of a desired cellular analyte including drug molecules or biomarkers. This can reveal information on cell health (apoptosis, cytotoxicity, viability and proliferation assays), cell metabolism, cell migration and cell signaling mechanisms. Find the best cell-based assay products, kits and equipment with our peer reviewed product directory: compare products, check customer reviews and receiving pricing direct from manufacturers.ImmunofluorescenceLabelingProtein BiologyThe analysis of protein expression, identity and function is vital for many areas of life science research and drug discovery. Some of the most commonly used techniques in protein analysis include Western blotting, electrophoresis and mass spectrometry.Spectral ScanningConjugationTR-FRET

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Micro-Volume Determination of Labeling Efficiency Using Spectral Scanning Analysis in a Standard Microplate Reader