Hybridoma binding (GST antigen) kit
HTRF Hybridoma binding (GST antigen) kit
The hybridoma binding (6HIS antigen) kit is designed to screen mouse hybridoma supernatants against 6HIS-tagged targets.
The hybridoma technology is still the most common technique used to generate antibodies for therapeutic applications. The generation and screening of hybridomas from an immunized animal involve a time-consuming process and sample only a fraction of the antibodies generated during the adaptive immune response. Therefore, screening hybridoma supernatants for their ability to efficiciently bind the target of interest has become crucial. The hybridoma binding (6HIS antigen) kit has been designed to reliably screen mouse hybridoma supernatants against specific 6HIS-tagged antigens.
The mouse hybridoma supernatant binding to a 6HIS-tagged antigen is measured using a sandwich immunoassay, involving an anti-6HIS antibody labeled with an HTRF donor and an anti-mouse antibody labeled with an acceptor dye. The binding of murine antibodies produced by the hybridoma clone to their 6HIS-tagged target generates an HTRF Signal.
The HTRF hybridoma binding kit can be run in a 96- or 384-well low volume white plate (20 µL final). As described here, samples are dispensed directly into the assay plate. The 6HIS antigen is then added, followed by the dispensing of the HTRF reagents: the Anti 6HIS antibody labeled with europium cryptate and the anti mouse antibody labeled with d2. The protocol can be further miniaturized or upscaled by simply resizing each addition volume proportionally.
Binders of TNF-alpha fused to the HIS tag were detected using an anti HIS europium cryptate antibody and an anti mouse d2 antibody.
The hybridoma binding kit protocol was performed using 5 nM HIS tagged TNF-alpha protein for 1H at room temperature.
The graph shows the signal obtained with a positive sample (red) and a negative sample (orange).
The Hybridoma (HIS antigen) control was also used to validate the experiment.
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