ArticlePloS one2021
High-level expression of the monomeric SARS-CoV-2 S protein RBD 320-537 in stably transfected CHO cells by the EEF1A1-based plasmid vector.
Article in PloS one, 2021. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 26 papers.
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Who cites it
26 citing papers in PubMed, 40 citations in OpenAlex.
- Production of a Dulaglutide Analogue by Apoptosis-Resistant Chinese Hamster Ovary Cells in a 3-Week Fed-Batch Process.Pharmaceuticals (Basel, Switzerland) · 2025Article
- Expression of Recombinant Tissue Plasminogen Activator - Plasminogen Activator Inhibitor Complex in Mammal Cells as Quality Control Materials in Immunoassay.The protein journal · 2025Article
- Leveraging Walnut Somatic Embryos as a Biomanufacturing Platform for Recombinant Proteins and Metabolites.Biotech (Basel (Switzerland)) · 2024Article
- Expression and purification of the receptor-binding domain of SARS-CoV-2 spike protein in mammalian cells for immunological assays.FEBS open bio · 2024Article
- Utilization of Receptor-Binding Domain of SARS-CoV-2 Spike Protein Expressed in Escherichia coli for the Development of Neutralizing Antibody Assay.Molecular biotechnology · 2023Article
- Comparison of vector elements and process conditions in transient and stable suspension HEK293 platforms using SARS-CoV-2 receptor binding domain as a model protein.BMC biotechnology · 2023Article
- Stable production of recombinant SARS-CoV-2 receptor-binding domain in mammalian cells with co-expression of a fluorescent reporter and its validation as antigenic target for COVID-19 serology testing.Biotechnology reports (Amsterdam, Netherlands) · 2023Article
- Spike protein receptor-binding domains from SARS-CoV-2 variants of interest bind human ACE2 more tightly than the prototype spike protein.Biochemical and biophysical research communications · 2023Article
- Development of a scalable single process for producing SARS-CoV-2 RBD monomer and dimer vaccine antigens.Frontiers in bioengineering and biotechnology · 2023Article
- Two formulations of coronavirus disease-19 recombinant subunit vaccine candidate made up of S1 fragment protein P1, S2 fragment protein P2, and nucleocapsid protein elicit strong immunogenicity in mice.Immunity, inflammation and disease · 2022Article
- The crosstalk between LINC01089 and hippo pathway inhibits osteosarcoma progression.Journal of bone and mineral metabolism · 2022Article
- Article
- Recombinant expression of SARS-CoV-2 receptor binding domain (RBD) inIranian journal of basic medical sciences · 2022Article
- Insights into potential mechanisms of asthma patients with COVID-19: A study based on the gene expression profiling of bronchoalveolar lavage fluid.Computers in biology and medicine · 2022Article
- Silent Antibodies Start Talking: Enhanced Lateral Flow Serodiagnosis with Two-Stage Incorporation of Labels into Immune Complexes.Biosensors · 2022Article
- Expression, purification and characterization of SARS-CoV-2 spike RBD in ExpiCHO cells.Protein expression and purification · 2022Article
- Designed SARS-CoV-2 receptor binding domain variants form stable monomers.Biotechnology journal · 2022Article
- Phosphate-regulated expression of the SARS-CoV-2 receptor-binding domain in the diatom Phaeodactylum tricornutum for pandemic diagnostics.Scientific reports · 2022Article
- Fast and Accurate Surrogate Virus Neutralization Test Based on Antibody-Mediated Blocking of the Interaction of ACE2 and SARS-CoV-2 Spike Protein RBD.Diagnostics (Basel, Switzerland) · 2022Article
- Glycosylation of Receptor Binding Domain of SARS-CoV-2 S-Protein Influences on Binding to Immobilized DNA Aptamers.International journal of molecular sciences · 2022Article
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Authors and funding
5 authors at 2 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
The spike (S) protein is one of the three proteins forming the coronaviruses' viral envelope. The S protein of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has a spatial structure similar to the S proteins of other mammalian coronaviruses, except for a unique receptor-binding domain (RBD), which is a significant inducer of host immune response. Recombinant SARS-CoV-2 RBD is widely used as a highly specific minimal antigen for serological tests. Correct exposure of antigenic determinants has a significant impact on the accuracy of such tests-the antigen has to be correctly folded, contain no potentially antigenic non-vertebrate glycans, and, preferably, should have a glycosylation pattern similar to the native S protein. Based on the previously developed p1.1 vector, containing the regulatory sequences of the Eukaryotic translation elongation factor 1 alpha gene (EEF1A1) from Chinese hamster, we created two expression constructs encoding SARS-CoV-2 RBD with C-terminal c-myc and polyhistidine tags. RBDv1 contained a native viral signal peptide, RBDv2 -human tPA signal peptide. We transfected a CHO DG44 cell line, selected stably transfected cells, and performed a few rounds of methotrexate-driven amplification of the genetic cassette in the genome. For the RBDv2 variant, a high-yield clonal producer cell line was obtained. We developed a simple purification scheme that consistently yielded up to 30 mg of RBD protein per liter of the simple shake flask cell culture. Purified proteins were analyzed by polyacrylamide gel electrophoresis in reducing and non-reducing conditions and gel filtration; for RBDv2 protein, the monomeric form content exceeded 90% for several series. Deglycosylation with PNGase F and mass spectrometry confirmed the presence of N-glycosylation. The antigen produced by the described technique is suitable for serological tests and subunit vaccine studies.
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