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VHH generation

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Phase I – Immunization 
You have the choice between dromedary and lama. The dromedary provides more sequence variability and better stability.

Standard immunization (each two weeks) or Rapid immunization (each week).

This step is done in collaboration with our partner Dromelys ( for more information www.dromelys.com)

Antigens : protein, peptide, Toxins, chemical molecules, cells, cell membrane.

Protocol of immunization and recovery :

After seven injections, peripheral blood lymphocytes (PBLs) of the immunized dromedary or Llama were isolated and used to construct the library.

Phase II – Lymphocyte isolation, cDNA preparation, Amplification and phage library
cDNA is generated via RNA isolation, amplified via PCR, and a diverse phage library is generated for screening against the antigen.

Total RNA was isolated from PBLs . To avoid contamination with VH genes, the variable regions of heavy-chain immunoglobulins (VHH) were amplified by nested PCR.  The VHH was extracted from the gel. The amplified second PCR products were digested with Pst I and Not I restriction enzymes, then inserted into the phagemid. Ligation products were transformed into Escherichia coli TG1 cells by electroporation. Many clones were selected randomly and used to estimate the percentage of clones with a proper insert size within library.

A fraction of the VHH library was cultured and infected with helper phages to express the VHH at the tip of phage particles for screening against the antigen. This process represented one round of panning. The generated phage particles were used in the next round of panning. During three to four rounds of panning, Proteins-specific phages were enriched gradually.

Phase III – Antibody Screening
The phage library will be screened against the desired antigen and ideally 6 potential clones will be identified and evaluated.

Phase IV – Production
We express the final antibody in a protein expression system. The desired quantity of antibody is produced and purified.

 

  • In the case of unit non-response, we will re-inject the antigen (no additional charges).
  • You are the only owner of intellectual property