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Hybridoma binding (6HIS antigen) kit HTRF®

The hybridoma binding (6HIS antigen) kit is designed to screen mouse hybridoma supernatants against 6HIS-tagged targets.

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  • No-wash No-wash
  • All inclusive kit All inclusive kit
  • Low sample consumption Low sample consumption

The hybridoma binding (6HIS antigen) kit is designed to screen mouse hybridoma supernatants against 6HIS-tagged targets.

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Overview

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.

Benefits

  • Mouse hybridoma screening
  • Selection of all biotherapeutic binders
  • 6HIS-tagged antigen compatible

Assay Principle

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.

Principle of the HTRF HIS Hybridoma assay

Assay Protocol

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.

Protocol of the HTRF GST Hybridoma assay

HIS tagged TNFa binding assay

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.

Validation of the HTRF HIS hybridoma kit on HIS-TNF alpha protein

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Plate Reader Requirement

Choosing the right microplate reader ensures you’ll get an optimal readout. Discover our high performance reader, or verify if your lab equipment is going to be compatible with this detection technology.

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