The future of
single-domain antibodies



CDR Library Construction

 

Looking for VHH nanobodies with enhanced affinity? We build CDR libraries for further antibody optimization.

Applications

  • Antibody optimization after Phage Display and/or Yeast Two-Hybrid selection.

Key benefits

  • Generation of a very high number of CDR variants (diversity of a million CDRs)
  • Selection of quality sequences only without codon bias (deletions/insertions removed)
  • Library ready for further antibody optimization

How does it work?

Using Agilent-based DNA chips and primer extension, Hybrigenics is able to generate highly complex CDR variant libraries. For example, we generate 1 million variants of the 13 amino-acid CDR3, representing all single, double and triple variants in aminoacid sequence to optimize the efficiency of an antibody.

 CDR Library Construction by NGS

The actual quality of the DNA chips oligos coding for CDR is limited by the presence of deletions or insertions. To overcome this technical challenge, we developed and efficient yeast-based selection that suppresses deleted or inserted sequences based on natural gap repair.
After primer extension and linker ligation on DNA chips, a denaturation step and the addition of long PCR oligos of 45 nucleotides allow efficient recombination with a yeast plasmid.
An engineered yeast strain selects CDR sequences with no stop codon using a protocol based on the Yeast Two-Hybrid  method: yeast are able to grow on a selective medium lacking histidine only if 2 proteins can interact and reconstitute a full transcription factor that will enable the expression of HIS3 gene. One of these proteins is coded by the vector that recombined with the CDR sequence. Positive yeast clones express only correct CDR sequences.

 

CDR Library validation by Yeast Two-Hybrid

Material required

  • 5 µg of the DNA of your antibody

Deliverables

  • DNA library of 1 million CDRs.

Associated services

Hybrigenics Services can use high diversity CDR library to select the most affine antibodies by Yeast Two-Hybrid for intrabodies (Super-Binders) or by Yeast Display. Contact us for specific enquiries about antibody optimization.

 

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