U-Protein Express BV is specialized in the production of research and preclinical grade recombinant proteins and recombinant antibodies in mammalian expression systems. Fully post-translational modified mammalian recombinant proteins and antibodies are produced via our proprietary rPEx and transient production platforms. The platform consist of serum free suspension growing HEK and CHO cells which are transiently transfected. Dedicated purification protocols yield highly pure protein preparations with extreme low endotoxin levels, in general below 0.05 EU/mg.

Recombinant protein production

Fully post-translational modified mammalian proteins are produced via the rPEx transient protein production platform. The rPEx production platform consists of serum free suspension growing HEK and CHO cells which are transiently transfected with expression vectors containing the gene of interest. For each recombinant protein to be expressed, several proprietary expression vectors (pUPE expression vectors) are available. Due to the modular architecture, the expression vectors can be easily adapted to meet specific requirements. This allows, for example, choice of nature and position of the purification tag.

Large amount of recombinant protein can be produced in a very short time span (generally less than 8 weeks). The pipeline from gene to purified protein comprises the following steps:

  • Generation of coding cDNA via synthetic gene design and codon optimization.
  • Ligation of the coding sequence in several expression vectors.
  • Small scale transient protein production to suspension growing HEK293 cells. Three to 6 days post transfection recombinant protein production is assayed and the most optimal expression vector is selected.
  • Large scale transient protein production in suspension growing HEK293 or CHO cells.
  • Purification and characterization of the recombinant protein.

The rPEx platform allows for several transfection and delivery options to match your needs. For more information on the transfection and delivery options, please send us an email.

Recombinant antibody production

Fully post-translational modified mammalian recombinant antibodies (rAb’s) are produced via the rPEX expression platform. Antibody variable domains can be seamlessly ligated in antibody expression vectors to generate each antibody class.

Gram amounts of purified antibodies with low levels of endotoxin (< 0.05 EU/mg) can be provided in 4 to 6 weeks. The recombinant antibody production comprises the following steps:

  • Generation of coding sequences of the antibody variable domains via synthetic gene design and codon optimization.
  • Expression vector generation by ligation of antibody variable domain cDAN fragments in antibody expression vectors.
  • Transient production of antibody expression vectors to HEK293 or CHO-Potelligent cells via the rPEx technology.
  • Purification of the recombinant antibody via affinity chromatography (proteinA) and gel filtration chromatography.
  • Assaying the function of the purified antibody.

The platform has a modular architecture, thereby allowing several transfection and delivery options. For more information on the transfection and delivery options, please send us an email.

Mouse Rapid Prime monoclonal antibody development

Rapid Prime is ImmunoPrecise’s proprietary custom monoclonal antibody development service. This rapid immunization strategy saves weeks in development time. When coupled with the Single-Step cloning procedure, positive hybridoma cell lines are identified in 65 days. This rapid immunization procedure does not compromise on affinity. It consistently generates large numbers of IgG clones specific for antigen.

PROJECT PHASESTIMEFRAME
I.Immunization of 4 female BALB/c mice 19 Days
II.Fusion and Selection of Hybridomas 16 Days
III.Screening of Hybridoma Clones 32 Days
IV.Subcloning and Cryopreservation 35 Days
V.Purification (Optional) 4-7 Weeks

IPA’s proprietary Rapid Prime Method in mice identifies positive monoclonal antibodies in as little as 32 days. While this method also uses Single-Step cloning to accelerate the process, it is especially good for generating anti-idiotypic antibodies and has also been extremely useful for generating monoclonal antibodies against conformational epitopes. Other sources of immunogens include conserved or smaller proteins, peptides, and whole cell bacteria.

More information about the Rapid Prime method can be found here.

Standard method monoclonal antibody development

ImmunoPrecise’s Standard Method monoclonal antibody development service in mice and rats is fast and efficient. IPA’s single-step cloning method shaves weeks off of the traditional time line for hybridoma production. IPA custom develops monoclonal antibodies to an array of antigens including peptides, modified peptides, recombinant proteins, DNA and small molecules.

PROJECT PHASESTIMEFRAME
I.Immunization of 4 female BALB/c mice or 2 female F244 (SAS FISCH) rats 63 Days
II.Fusion and HAT Selection of Hybridomas 16 Days
III.Screening and Expansion of Hybridoma Clones 32 Days
IV.Subcloning and Cryopreservation 35 Days
V.Purification Antibody Concentration and Purification (Optional) 4-7 Weeks

More information about the Standard Method monoclonal antibody development can be found here.

Rabbit monoclonal antibody development

ImmunoPrecise Antibodies offers custom Rabbit Monoclonal Antibody development service using RMAT (Recombinant Monoclonal Antibody Technology). This proprietary technology screens the repertoire of the rabbit creating the power to select the desired antibody directly from B cells.

The RMAT Advantage

  • - High affinity antibodies without the traditional cell fusion step; avoids stability issues commonly associated with hybridomas
  • - Recombinant antibody ensures preservation of genetic information
  • - Both a monoclonal and polyclonal end product
  • - Utilization of rabbit immune system to generate antibodies against different epitopes on the same antigen
PROJECT PHASESTIMEFRAME
I.Immunization of 2x female 78 Days
II.Screening and Selection 11 Days
III.Cloning and Expression
  • - Clone heavy and light chains into separate expression vectors
  • - Cotransfect heavy and light chain plasmids in mammalian cell line for antibody expression
  • - Identification of antigen specific antibodies by indirect ELISA
30 Days

More information about the Rabbit monoclonal antibody development can be found here.

Polyclonal development

ImmunoPrecise is a leading producer of high-titre custom mouse, rat, rabbit, goat, and sheep polyclonal antibodies. We offer a comprehensive range of services to get you the polyclonal product you need:

  • - Epitope prediction
  • - Custom peptide synthesis
  • - Conjugation
  • - Immunization
  • - Purification
  • - Assay design
PROJECT PHASESTIMEFRAME
I.Pre-immune bleeds are taken. Primary immunization Day 0
II.1st boost Day 28
III.2nd boost Day 47
IV.Test bleeds are taken. Sera may be shipped to client upon request for testing in specific assays Day 59
V.3rd boost Day 66
VI.Final Bleed Day 78

More information about the Polyclonal development can be found here.

Single domain antibodies (VHHs) development

The technology behind the products and services provided by QVQ is based on the smallest naturally occurring antibody-based molecules, called VHH or Nanobdy (~15 kDa). Next to the small size, VHH are highly soluble, show the ability to refold to their functional structure after denaturation and are able to target alternative epitopes, which are not targeted by conventional monoclonal antibodies and their minimized versions. VHHs can be obtained from the immunization of llamas.

PROJECT PHASESTIMEFRAME
I.Immunization and library construction
  • - QVQ Research outsources immunizations of llamas to a company specialized in the immunizations and production of polyclonal antibodies.
-- Days
II.II. Phage display and VHH selection -- Days
III.VHH sequence analysis
  • - QVQ’s database of VHH sequences is used for the analysis of VHH production, stability and protease resistance.
-- Days
IV.VHH production
  • - Production of recombinant VHH from bacterial and yeast hosts for samples up to 100 mg. Ook vermelden dat dit door UPE kan worden gedaan (gram amounts en in CHO cellen oid?)
-- Days
V.VHH labeling -- Days