Journal: Biomedicines

Article Title: A Small Molecule That In Vitro Neutralizes Infection of SARS-CoV-2 and Its Most Infectious Variants, Delta, and Omicron

doi: 10.3390/biomedicines11030916

Figure Lengend Snippet: Lentiviral system approach and antiviral activities of the test drugs against SARS-CoV-2 and its variants Delta and Omicron in vitro. ( a ) HEK293 cells were transfected with a plasmid encoding a lentiviral backbone (genome) expressing a marker protein, a plasmid expressing spike proteins of SARS-CoV-2 and its variants Delta and Omicron, and plasmids expressing the other proteins needed for virion formation (Tat, Gag-Pol, and Rev). The transfected cells produced lentiviral particles with SARS-CoV-2 spikes on their surface. These viral particles can infect cells that overexpress the ACE2 receptor. The readout to measure the grade of viral infection is the green fluorescence of ZsGreen1 from infected cells. ( b ) The molecular structure shows the interface of the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein (white surface) with the human ACE2 receptor (pink surface). CD04872SC is predicted to bind on the face leading to the potential disruption of the complex. ( c ) Drug screening of the selected top compounds (each at a concentration of 100 µM) was evaluated for their antiviral activity against SARS-CoV-2, as defined by their percent infection inhibition. ( d ) The maximum antiviral activity against SARS-CoV-2, and its two major variants Delta and Omicron, for lead drug candidate CD04872SC. ( e ) The mean percent inhibition of viral infection and percent cell cytotoxicity of lead drug candidate CD04872SC in HEK293 cells expressing the ACE2 receptor. In brief, HEK293 cells overexpressing the ACE2 receptor were pre-treated with the lead drug candidate CD04872SC for 1 h. Next, 10 µL of the lentiviral suspensions expressing the spike protein was then added to the culture plates and incubated for 48 h. After the incubation period, the green fluorescence of ZsGreen1 protein was measured with the plate reader Synergy 2 (BioTek, Winooski, VT, USA) using the FITC channel. The mean percent inhibition and percent cytotoxicity is represented on each of the graph’s left and right y-axis, respectively. For each of the three graphs, blue curves indicate the percent of infection inhibition and red curves represent the percent of cytotoxicity. The results are expressed as mean ± s.e.m. of three experiments performed in triplicate; each triplicate was averaged before calculating the s.e.m.

Article Snippet: HEK293 cells (ATCC-1586) overexpressing the ACE2 receptor were infected with lentiviruses that express the spike protein found in SARS-CoV-2 [ ] ( a).

Techniques: In Vitro, Transfection, Plasmid Preparation, Expressing, Marker, Produced, Infection, Fluorescence, Binding Assay, Disruption, Drug discovery, Concentration Assay, Activity Assay, Inhibition, Incubation

Journal: Emerging Infectious Diseases

Article Title: Candidate Cell Substrates, Vaccine Production, and Transmissible Spongiform Encephalopathies

doi: 10.3201/eid1712.110607

Figure Lengend Snippet: Cell cultures exposed to transmissible spongiform encephalopathy agents and propagated for 30 passages*

Article Snippet: We studied the following 5 actual or candidate cell substrates used or proposed for production of biologic products: CHO-K1 (Chinese hamster ovary ATCC-CCL61); Vero C1008 (African green monkey kidney ATCC-CRL1586); WI-38 (human lung diploid fibroblasts ATCC-CCL75); MDCK (dog kidney ATCC-CCL34); and HEK-293 (human embryonic kidney ATCC-CRL-1573 transformed with defective adenovirus as a surrogate for PER.C6) (Crucell, Leiden, Netherlands) ( ).

Techniques: Infection

Journal: Vaccines

Article Title: Development of an Inactivated Vaccine against SARS CoV-2

doi: 10.3390/vaccines9111266

Figure Lengend Snippet: Preparation of ERUCoV-VAC. ( a ) The viral kinetics of the hCoV-19/Turkey/ERAGEM-001/2020 strain at different multiplicities of infection (MOI). ( b ) Flowchart of ERUCoV-VAC preparation. ( c ) Protein profiles of the hCoV-19/Turkey/ERAGEM-001/2020 virus strain before and after purification. Total proteins were separated by Nu-page 10% bis-tris SDS-PAGE gel. Lane 1, protein molecular weight marker (M). Lane A, uninfected Vero E-6 cells as a negative control. Lane B, Vero E-6 cells infected with the hCoV-19/Turkey/ERAGEM-001/2020 virus strain. Lane C, purified hCoV-19/Turkey/ERAGEM-001/2020 virus strain. Western blot analysis of purified hCoV-19/Turkey/ERAGEM-001/2020 virus strain probed with a human antibody to the SARS-CoV-2 nucleocapsid protein (1:2500) (GenScript; HC2003) (Lane E) or a rabbit polyclonal to SARS-CoV-2 spike glycoprotein (lane G). Uninfected cell lysates probed with the SARS-CoV-2 nucleocapsid protein (Lane D) and a rabbit polyclonal to SARS-CoV-2 spike glycoprotein (F) were used as a negative control. The arrows indicate that the bands at approximately 180 kDa and 48 kDa represent spike glycoprotein and nucleocapsid protein, respectively.

Article Snippet: Vero E6 cells (African green monkey kidney) obtained from ATCC (CRL 1586) were maintained in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% heat-inactivated fetal bovine serum (FBS) and 100 mM L-glutamine (Sigma–Aldrich, Darmstadt, Germany).

Techniques: Infection, Virus, Purification, SDS Page, Molecular Weight, Marker, Negative Control, Western Blot

Journal: Virology

Article Title: Replication and budding of simian immunodeficiency virus in polarized epithelial cells.

doi: 10.1006/viro.1999.9637

Figure Lengend Snippet: FIG. 1. Virus released after SIVmac239 proviral DNA transfection of Vero C1008 cells that were then cultured on Millicell filters. Culture medium was collected daily and dilutions were used to infect CEMx174 cells. Virus titers (TCID50) were determined from IFA-positive staining of infected CEMx174 cells with anti-gp130 MAb.

Article Snippet: MATERIALS AND METHODS ell culture Two African green monkey cell lines, Vero C1008 ATCC CRL 1587) and CV-1 (ATCC CCL 70), were obained from the American Type Culture Collection.

Techniques: Virus, Transfection, Cell Culture, Staining, Infection

Journal: Virology

Article Title: Replication and budding of simian immunodeficiency virus in polarized epithelial cells.

doi: 10.1006/viro.1999.9637

Figure Lengend Snippet: FIG. 2. Polarized release of SIVmac239. (A, C) Vero C1008 cells were transfected with SIVmac239 proviral DNA in tissue culture plates. One day after transfection, cells were trypsinized and seeded on Millicell-HA filters. (B, D) CD41 Vero C1008 cells were infected with SIVmac239 in tissue culture plates. After 24 h of adsorption, virus inoculum was removed and the cells were washed 6 times with MEM. Cells were incubated in fresh medium at 37°C for an additional 24 h, after which the cells were trypsinized and transferred to Millicell-HA filters. Culture medium was collected from both apical and basolateral domains and fresh medium was added daily. Viral p27 antigen was quantified by ELISA. (A, B) Quantity of p27 antigen released from each cellular domain. (C, D) Percentage of total virus released from each domain.

Article Snippet: MATERIALS AND METHODS ell culture Two African green monkey cell lines, Vero C1008 ATCC CRL 1587) and CV-1 (ATCC CCL 70), were obained from the American Type Culture Collection.

Techniques: Transfection, Infection, Adsorption, Virus, Incubation, Enzyme-linked Immunosorbent Assay

Journal: Virology

Article Title: Replication and budding of simian immunodeficiency virus in polarized epithelial cells.

doi: 10.1006/viro.1999.9637

Figure Lengend Snippet: FIG. 3. SIV maturation and budding at the basolateral surface of a polarized epithelial cell. SIVmac239 proviral DNA transfected Vero C1008 cells were grown to confluence on Millicell-HA filters and prepared for electron microscopy. Virus budding and release of progeny virions were observed only at the basolateral surface (arrows) (bar indicates 2 mm). The insert shows a higher magnification of basolateral budding (bar indicates 0.1 mm). The apical surface shows microvilli and an underlying tight junction between adjacent cells. M indicates filter membrane.

Article Snippet: MATERIALS AND METHODS ell culture Two African green monkey cell lines, Vero C1008 ATCC CRL 1587) and CV-1 (ATCC CCL 70), were obained from the American Type Culture Collection.

Techniques: Transfection, Electron Microscopy, Virus, Membrane

Journal: Virology

Article Title: Replication and budding of simian immunodeficiency virus in polarized epithelial cells.

doi: 10.1006/viro.1999.9637

Figure Lengend Snippet: FIG. 4. Western blot analysis of SIVmac envelope proteins expressed in proviral DNA transfected Vero C1008 cells, virus infected Vero C1008 cells, and virus infected HUT.78 cells. Proteins from lysates of SIV- mac239 virus infected HUT.78 cells (lane 1), SIVmac239 virus infected CD41 Vero C1008 cells (lane 2), SIVmac239 proviral DNA transfected Vero C1008 cells (lane 3), SIVmac251 virus (cloned virus with a prema- ture stop codon in the TM portion of the envelope gene) infected CD41

Article Snippet: MATERIALS AND METHODS ell culture Two African green monkey cell lines, Vero C1008 ATCC CRL 1587) and CV-1 (ATCC CCL 70), were obained from the American Type Culture Collection.

Techniques: Western Blot, Transfection, Virus, Infection, Clone Assay

Journal: Virology

Article Title: Replication and budding of simian immunodeficiency virus in polarized epithelial cells.

doi: 10.1006/viro.1999.9637

Figure Lengend Snippet: FIG. 5. Polarized release of SIVmac1A11 and SIVmac251 in polarized Vero cells and symmetrical release of SIVmac239 in nonpolarized CV-1 cells. CD41 Vero C1008 cells were infected with SIVmac251 virus (A, D), Vero C1008 cells were transfected with SIVmac1A11 proviral DNA (B, E), or CV-1 cells were transfected with SIVmac239 proviral DNA (C, F) in tissue culture plates. Cells were subsequently transferred to Millicell-HA filters and culture medium was collected for analysis as described in the legend to Fig. 2. (A, B, C) Quantity of p27 antigen released from each domain. (D, E, F) Percentage of total virus released from each domain.

Article Snippet: MATERIALS AND METHODS ell culture Two African green monkey cell lines, Vero C1008 ATCC CRL 1587) and CV-1 (ATCC CCL 70), were obained from the American Type Culture Collection.

Techniques: Infection, Virus, Transfection

Journal: Virology

Article Title: Replication and budding of simian immunodeficiency virus in polarized epithelial cells.

doi: 10.1006/viro.1999.9637

Figure Lengend Snippet: FIG. 6. Polarized release of intact HIV and HIV with truncated gp41 in polarized Vero cells. Vero C1008 cells were transfected in tissue culture plates with HIV proviral DNA and with full-length gp41 (clone ATG-(1)gp160) (A, D) or truncated gp41, clone ATG-(1)gp41D726-856 (B, E) or ATG- (1)gp41D754-856 (C, F). Cells were subsequently transferred to Millicell-HA filters and culture medium was collected for analysis as described in the legend to Fig. 2. (A, B, C) Quantity of p24 antigen released from each domain. (D, E, F) Percentage of total virus released from each domain.

Article Snippet: MATERIALS AND METHODS ell culture Two African green monkey cell lines, Vero C1008 ATCC CRL 1587) and CV-1 (ATCC CCL 70), were obained from the American Type Culture Collection.

Techniques: Transfection, Virus

Journal: Journal of Chemical Information and Modeling

Article Title: AI-Driven Discovery of SARS-CoV-2 Main Protease Fragment-like Inhibitors with Antiviral Activity In Vitro

doi: 10.1021/acs.jcim.3c00409

Figure Lengend Snippet: Anti-SARS-CoV-2 activity (A) and cytotoxicity (B) of compounds 818, 737, and 183 in Vero E6 (left) and Calu-3 (right) cells.

Article Snippet: Vero E6 (African green monkey kidney, ATCC CRL-1586) and Calu-3 (sub-mucosal gland cell line, generated from a bronchial adenocarcinoma, kindly donated by Farmanguinhos platform RPT11M) cells were infected with SARS-CoV-2 B.1 lineage isolate (GenBank MT710714, SisGen AC58AE2) in MOI 0.01.

Techniques: Activity Assay

Journal: Journal of Chemical Information and Modeling

Article Title: AI-Driven Discovery of SARS-CoV-2 Main Protease Fragment-like Inhibitors with Antiviral Activity In Vitro

doi: 10.1021/acs.jcim.3c00409

Figure Lengend Snippet: Pharmacological Parameters of SARS-CoV-2-Infected Cells in the Presence of Compounds 818, 737, and 183 a

Article Snippet: Vero E6 (African green monkey kidney, ATCC CRL-1586) and Calu-3 (sub-mucosal gland cell line, generated from a bronchial adenocarcinoma, kindly donated by Farmanguinhos platform RPT11M) cells were infected with SARS-CoV-2 B.1 lineage isolate (GenBank MT710714, SisGen AC58AE2) in MOI 0.01.

Techniques: