The future of
single-domain antibodies



Nanobody Selection & Validation for Cell Surface Antigens

Antibody Phage Display Selection & IF/FACS Validation Process for Cell Surface Antigens

Do you need an affine antibody recognizing your cell surface receptor? Let us select a nanobody for you.

Applications

  • Detection and quantification of cell surface receptors
  • Identification of nanobodies active on receptors/transporters (e.g. agonists, antagonists…)
  • Structural studies (co-crystallization)

Key benefits

  • Nanobodies binding native receptors or cell surface antigens
  • Cost-effective production of nanobodies
  • No variability from batch to batch
  • Versatile tool to use in a vast array of experiments thanks to the possible fusion with Fc fragments from human, mouse or rabbit (see Minibody Cloning & Production)

How does it work?

Hybrigenics can select nanobodies by phage display directly on living cells when in vitro selection is not possible e.g for receptors with transmembrane domains like GPCR's. Our hs2dAb library coding for 3.109 VHH nanobodies is expressed at the phage surface and incubated with the cell expressing the antigen of interest. After a washing step with cells that do not express the antigen, the elution of the specific nanobodies is performed and their enrichment is measured. 3 to 4 rounds of selection are performed.

 Selection & IF-validation of cell surface antigen

Selected nanobodies are validated by immunofluorescence (IF). Each nanobody is tagged and expressed as an E.coli supernatant. Its colocalization with the antigen is detected with a secondary labeled anti-tag antibody by fluorescence microscopy.

IF validation for cell surface antigens

Selected VHHs can also be validated by Phage FACS. After selection by phage display selection, cells are incubated with an anti-M13 antibody labelled with a fluorescent marker and sorted in a FACS. Fluorescence signal is compared between cells expressing the antigen of interest and cells which do not.

Phage FACS validation for cell surface antigensMaterial required

  • 5 µg of the cDNA of your antigen
  • A pellet of the cell line expressing your antigen of interest (stable inducible cell line)
  • If you have other material, please contact us.

Deliverables

  • Full report containing the selection process description and the immunofluorescence data (and phage FACS data if performed)
  • Sequences and cDNA clones of up to 10 IF/FACS validated Hybribodies in the pHEN2 vector (myc fusion, expression in E.coli supernatant)

 

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