Spike (B.1.618 Variant) Pseudotyped Lentivirus (Luc Reporter)

• Description:

  The pandemic coronavirus disease 2019 (COVID-19) is caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) . As the first step of the viral replication, the virus attaches to the host cell surface before entering the cell. The viral Spike protein recognizes and attaches to the Angiotensin-Converting Enzyme 2 (ACE2) receptor found on the surface of type I and II pneumocytes, endothelial cells, and ciliated bronchial epithelial cells. Drugs targeting the interaction between the Spike protein of SARS-CoV-2 and ACE2 may offer protection against the viral infection. A variant called B.1.618 was identified in India in the spring of 2021. This variant has a number of mutations that allow the virus to spread more easily and quickly than other variants.The Spike (B.1.618 Variant) (SARS-CoV-2) Pseudotyped Lentiviruses were produced with SARS-CoV-2 B.1.618 Variant Spike (Genbank Accession #QHD43416.1 with B.1.618 mutations; see below for details) as the envelope glycoproteins instead of the commonly used VSV-G. These pseudovirions contain the firefly luciferase gene driven by a CMV promoter (Figure 1), therefore, the spike-mediated cell entry can be measured via luciferase activity. The Spike (B.1.618 Variant) (SARS-CoV-2) pseudotyped lentivirus can be used to measure the activity of neutralizing antibody against SARS-CoV-2 B.1.618 variant in a Biosafety Level 2 facility.Spike interacts with host cells by binding to membrane receptor ACE2 (angiotensin converting enzyme 2) . Based on experiments performed by scientists at BPS Bioscience, we know that the wild-type SARS-CoV-2 spike pseudotyped lentiviruses transduce the following cells with great efficiency:  ACE2-HEK293 cells (BPS Bioscience #79951), ACE2-CHO cells (BPS Bioscience #79959), ACE2-HeLa cells (BPS Bioscience #79958) . They also efficiently transduce TMPRSS2-Vero E6 cells (BPS Bioscience #78081), which express high endogenous levels of ACE2 and were stably transfected with human serine protease TMPRSS2  required for the priming of Spike and fusion of the virion with the plasma membrane. By contrast, it has been shown by others that SARS-CoV-2 spike pseudotyped lentiviruses do not transduce parental Calu3 and Vero E6 cells very well [Neerukondaet al. 2021, PlosOne PMID:33690649; Tandonet al. 2020, Scientific Reports PMID:33154514; Condor Capchaet al.2021, Front. Cardiovasc. Med. PMID:33521067; Pisilet al. 2021, Pathogens PMID:33540924].SARS-CoV-2 variant pseudoviruses have been validated using ACE2-HEK293 cells but have not been tested in other cells.As recommended in our protocol, 5 µl of virus/well in a 96-well plate provides a sufficient signal-to-noise ratio to perform inhibition studies. The amount of virus added to the cells can also be scaled down according to the user’s need.Spike Mutations in B.1.618 Variant:Y145delH146delE484KD614GFigure 1. Schematic of the Luciferase Reporter in SARS-CoV-2 Spike Pseudovirion

• Applications:

  1. Study the mechanism of viral transduction.2. Screening for neutralizing antibodies for SARS-CoV-2 Spike and ACE2.

• Shipping Conditions:

  -80°C

• Storage Conditions:

  Lentiviruses are shipped with dry ice. For long-term storage, it is recommended to store the lentiviruses at -80°C for up to 12 months from date of receipt. Avoid repeated freeze/thaw cycles. Titers can drop significantly with each freeze/thaw cycle.

• Notes:

  Troubleshooting GuideVisitbpsbioscience.com/lentivirus-faqfor detailed troubleshooting instructions. For all further questions, please email[email protected].

• Formulation:

  The lentiviruses were produced from HEK293T cells. Supplied in medium containing 90% DMEM + 10% FBS.Media FormulationThaw Medium 1 (BPS Bioscience #60187) :MEM medium supplemented with 10% FBS, 1% non-essential amino acids, 1 mM Na pyruvate, 1% Penicillin/Streptomycin.