Pseudovirus

The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) has spread widely around the world. The full spectrum of COVID-19 ranges from mild, self-limiting respiratory diseases to severe progressive pneumonia, multiorgan failure, and death. What's more, because the SARS-CoV-2 is highly contagious and asymptomatic, its transmission is more difficult to control. As a leading company, Creative Biolabs is actively taking action to provide reliable and high-quality pseudovirus products, manufacturing services, and characterization analysis for SARS-CoV-2 research, and assists our customers in the development of vaccines or treatments for SARS-CoV-2 infection.

Introduction to SARS-CoV-2

SARS-CoV-2 is an enveloped, non-segmented, positive-sense RNA virus that is widely distributed in humans and other mammals and belongs to the sarbecovirus subfamily. It is about 65-125 nm in diameter, contains single-stranded RNA, and has a coronal protrusion on its outer surface. Among them, the glycosylated spike (S) protein is the major surface protein of SARS-CoV-2, responsible for receptor binding and fusion of viral membrane and cell membrane. It is one of the key components that bind receptors and mediate membrane fusion and virus entry. Therefore, S protein is the main goal of neutralizing antibody and vaccine design.

Figure 1. Schematic structure of SARS-CoV-2.¹

Our Products

At present, we offer the following pseudovirus products for SARS-CoV-2 S protein:

In addition, if there is no product you need in our product list, we can customize the pseudovirus according to your needs.

Why Need Pseudovirus?

Due to its high infectivity and pathogenicity, SARS-CoV-2 research should be handled in the Biosafety Level (BSL)-3 laboratory, which limits the development of antiviral measures. Pseudoviruses are considered to be safe and effective virological tools, especially for emerging and re-emerging viruses, because they have the basic components of cell entry and virus infection, but lack of nucleic acid and lose the ability of self-replication. Therefore, using the SARS-CoV-2 S protein as a prototype and designing pseudoviruses to mimic the SARS-CoV-2 virus for further research is the choice advocated by many researchers. With extensive experience in pseudoviruses, Creative Biolabs provides pseudovirus services based on lentivirus (LV) and vesicular stomatitis virus (VSV) systems for SARS-CoV-2 S protein.

LV vector packaging systems

Because of their high efficiency, LV vectors are often the first choice for packaging systems to produce encapsulated pseudoviruses. These vectors are mainly from human immunodeficiency virus (HIV-1), simian immunodeficiency virus (SIV) or feline immunodeficiency virus (FIV).

VSV vector packaging systems

VSV vector packaging system is a general tool for producing pseudotype virus. The advantage of this system is that it has no strict selectivity for envelope protein, and the resulting virus can be operated in the BSL-2 laboratory.

Types of Pseudoviruses

The practicality of a pseudovirus system fundamentally depends on the chosen core viral backbone, which in turn determines factors such as host cell extent, integration capability, and expression kinetics.

A. Lentiviral Pseudoviruses (LPs)

• Backbone: Most commonly derived from human immunodeficiency virus type 1 (HIV-1) or feline immunodeficiency virus (FIV). These systems are genetically engineered to lack all the accessory genes required for viral replication.
• Key Characteristics: LPs possess the unique ability to infect both dividing and non-dividing cells and can mediate the stable integration of reporter gene constructs into the host cell genome.
• Applications: Ideal for long-term expression studies, and due to their high biosafety and stable transduction efficiency in various cell lines expressing the target receptor, they are currently the most widely used method for constructing SARS-CoV-2 spike protein pseudoviruses.

B. Rhabdovirus Pseudoviruses (based on VSV-G)

• Backbone: Typically based on vesicular stomatitis virus (VSV). These systems are usually produced in the form of single-cycle infectious particles (SCIPs).
• Key characteristics: These particles can express reporter genes very rapidly and at high levels, typically yielding results within 24 hours post-infection. They do not integrate their own genetic material and do not self-replicate.
• Applications: Ideal for ultra-rapid, high-throughput screening where extremely fast result readout is required, such as screening large libraries of small molecules or compounds to find viral entry inhibitors.

C. Alphavirus and Retrovirus Systems

• Backbone: Older-generation retroviruses or self-replicating alphavirus vectors (which can be modified to be replication-deficient).
• Key characteristics: While these systems may be less common in highly specific neutralization assays compared to lentivirus systems, they provide alternatives for specific cell lines or applications, particularly suitable for applications requiring non-integrative transient expression in dividing cells.
• Applications: These systems are particularly important in cancer immunotherapy research, where they can be used to rapidly and transiently express immunomodulatory genes or tumour-associated antigens in dividing tumour cells.

Why Choose Our Services?

At Creative Biolabs, we offer comprehensive and fully customizable pseudovirus services to accelerate your R&D process. Our capabilities are designed to provide end-to-end solutions from molecular design to final data delivery.

Custom Pseudovirus Packaging

We specialize in producing high-titer, highly infectious pseudoviruses, customizable to your needs. You can provide the gene sequence of the envelope glycoprotein you are interested in. Our team will handle molecular cloning, vector construction, and virus production using a VSV or lentiviral backbone.

Neutralization Assay Development and Testing

• Vaccine Evaluation: Evaluating neutralizing antibody responses induced by candidate vaccines.
• Therapeutic Antibody Screening: Identifying and characterizing monoclonal antibodies that block viral invasion.
• Convalescent Serum Analysis: Detecting the levels and persistence of neutralizing antibodies in patient samples.
• Cytotoxicity and Invasion Mechanism Studies: We can help you uncover the complex invasion pathways of viral pathogens.

Our Collaborative Process

Creative Biolabs employs a rigorous, phased approach to ensure complete client satisfaction and scientific excellence in every pseudovirus project. Our workflow is clearly designed, flexible, and maximizes efficiency.

1. Phase 1: Scientific Consultation and Design

   • Identifying the target viral envelope protein and its required mutations.
   • Selecting the optimal pseudovirus backbone (e.g., lentivirus or VSV-based pseudovirus) based on the target cell line and required detection kinetics.
   • Finalizing the reporter system and production scale.

2. Phase 2: Molecular Biology and Plasmid Construction

   • Sequence Optimization: Optimizing the EGP gene sequence for high-level transient expression in selected host cells.
   • Vector Assembly: Cloning the optimized gene along with the reporter gene cassette into a transfer vector. All packaging and envelope expression plasmids are prepared and rigorously quality-controlled via sequencing.

3. Phase 3: Virus Particle Production and Purification

   • Transient Mammalian Cell Transfection: Efficiently transfecting production cell lines using a complete set of essential plasmids (packaging plasmid, envelope plasmid, and transfer plasmid).
   • Collection and Concentration: The supernatant containing pseudoviruses is collected, filtered to remove cell debris, and concentrated using advanced methods such as ultracentrifugation or tangential flow filtration to ensure high purity and high titer.

4. Phase 4: Rigorous Quality Assessment and Validation

   • Physical Titer Assay: The number of viral particles is quantified using techniques such as p24 content assay (targeting the lentiviral core).
   • Functional Titer Assay: Infective Units (IU) or Transduction Units (TU) per milliliter of virus are measured by reporter gene expression assay in susceptible target cell lines.

Frequently Asked Questions

Connect with Us Anytime!

At Creative Biolabs, our deep understanding of viral biology, coupled with our commitment to manufacturing excellence and rigorous intellectual property compliance, ensures that the resources we provide not only meet but exceed the needs of the global scientific community. With our custom synthesis and high-volume production capabilities, researchers are able to respond rapidly to emerging pathogens and accelerate the discovery of life-saving interventions. For more extensive information, please contact us and request a price. We will contact you within 24 hours.

Reference

1. Santos I A, Grosche V R, Bergamini F R G, et al. Antivirals against coronaviruses: candidate drugs for SARS-CoV-2 treatment? Frontiers in microbiology, 2020, 11: 1818. https://doi.org/10.3389/fmicb.2020.01818
   (Distributed under Open Access license CC BY 4.0, without modification.)