Baculovirus Mediated Expression

Proteos provides baculovirus mediated protein expression in our fully licensed Sf9 (Spodoptera frugiperda) and Tni (Trichoplusia ni) cell lines.  Both cells lines are suitable for the production of intracellular or secreted recombinant proteins, and the resulting proteins contain a majority of the post-translational modifications found in mammalian cells.

The baculovirus expression vector systems (BEVS) that we offer utilizes site-specific transposition to produce recombinant virus encoding the protein of interest. To ensure consistent MOI (multiplicity of infection) and production reproducibility, Proteos employs a robust plate-based flow cytometry assay for baculovirus titer determination prior to infection of insect cells for protein production.  The titer of each virus stock that we produce is determined in triplicate using our proprietary plate-based flow cytometry assay.  We routinely produce high-titer virus stocks with titers ranging from 5 x 108 – 2 x 109 infection units (IU) per milliliter in a single amplification.

Learn More about Baculovirus Titer Assay Services

We utilize optimized and fully validated insect cell expression protocols for consistent reproducible high-level protein expression in serum-free shake flask culture volumes up to a 40 L batch size. Only mycoplasma-free, high viability, low passage cultures are used for expression and are monitored using a Beckman Coulter Vi-CELL XR for culture density, viability, and average cell diameter.  The Vi-CELL XR uses the trypan blue dye exclusion method and averages 50 measurements to compute the parameters for each sample.  This allows us to monitor cultures with a high degree of accuracy which is imperative for quality and reproducibility of protein production projects.  All cell culture data is included in a customized technical report delivered with each project.

Typical productions times are 8-10 weeks from construct design to purified protein.  

Services offered:

  • Full suite of molecular biology services to accompany any of our expression platforms, including construct design, fusion partner and affinity tag selection, gene synthesis with codon optimization, subcloning, endotoxin-free plasmid DNA isolation, and sequence verification.
  • Expression verification in volumes less than 100 mL; ideal for expression testing of multiple recombinant protein targets in parallel.
  • MOI (multiplicity of infection), cell line, and time of harvest optimization.
  • Pilot scale production and yield determination at culture scales ranging from 1 L to 2 L; ideal for process verification through purification prior to large-scale production.
  • Preparative scale production of recombinant proteins in Sf9 or Tni cells in shake flask culture volumes up to a 40 L batch size.
  • Generation, cryopreservation, and banking of BIIC (baculovirus infected insect cells) stocks for rapid scale-up and expression in future production runs.
  • Concentration of culture supernatant via tangential flow filtration.
  • Buffer-exchange via diafiltration for Ni-NTA purification of secreted His-tag proteins.
  • Titer determination of high-titer baculovirus stocks using Proteos’ proprietary flow cytometry assay.
  • Options for delivery include cell paste, conditioned culture supernatant, concentrated buffer-exchanged culture supernatant, high titer baculovirus stocks, or cryopreserved BIIC stocks.
  • Baculovirus mediated (insect cell) expression services can be combined with Proteos’ purification services for a fully-customized protein production process.

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