Human rhabdomyosarcoma cell line (RD, ATCC® CCL-136™), African green monkey kidney cell line Vero C1008 (ATCC® CRL-1586™), Human hepatoma cell line Huh-7 (a kind gift from Dr. Tianyi Wang at University of Pittsburgh), and HEK293T expressing ACE2 (293T-ACE2, BEI Resources, NR-52511) cell lines were maintained in Dulbecco’s modified eagle’s medium (DMEM); Human fibroblast cell line MRC-5 (ATCC® CCL-171™), Human lung adenocarcinoma cell line Calu-3 (ATCC® HTB-55™), human Colorectal adenocarcinoma cell line (Caco-2, ATCC® HTB-37™) were maintained in eagle’s minimum essential medium (EMEM, ATCC® 30-2003™). Both media were supplemented with 10% fetal bovine serum (FBS) and 1% penicillin-streptomycin antibiotics. Cells were kept at cell culture incubator (humidified, 5% CO2/95% air, 37 °C). The following reagents were obtained through BEI Resources, NIAID, NIH: human coronavirus, HCoV-OC43, NR-52725; human coronavirus, HCoV-NL63, NR-470. HCoV-OC43 was propagated in RD cell line; HCoV-NL63 was initially propagated in 293T-ACE2 cell line and accommodated in Vero E6 cell line. HCoV-229E was obtained from Dr. Bart Tarbet (Utah State University) and amplified in Huh-7 or MRC-5 cell lines.
Nr 52511
Manufactured by BEI Resources
Sourced in India
The NR-52511 is a laboratory instrument designed for conducting various analytical and research-oriented tasks. It is a versatile and reliable piece of equipment suitable for a wide range of applications. The core function of the NR-52511 is to provide accurate and reproducible measurements, but the specific details of its intended use are not available.
Automatically generated - may contain errors
Lab products found in correlation
Hi fbs, by Thermo Fisher Scientific (2 mentions)
Glutamax, by Thermo Fisher Scientific (2 mentions)
Ccl 136, by American Type Culture Collection (1 mentions)
Htb 37, by American Type Culture Collection (1 mentions)
Htb 55, by American Type Culture Collection (1 mentions)
Ccl 171, by American Type Culture Collection (1 mentions)
293t ace2, by BEI Resources (1 mentions)
Hcov oc43, by BEI Resources (1 mentions)
Nr 52725, by BEI Resources (1 mentions)
Nr 470, by BEI Resources (1 mentions)
Poly l lysine (pll), by Merck Group (1 mentions)
Cellect fbs gold, by MP Biomedicals (1 mentions)
Amphotericin b, by Merck Group (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
Dulbecco s modified eagle medium, by Thermo Fisher Scientific (1 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)
7 protocols using nr 52511
1
Diverse Cell Lines for Coronavirus Propagation
2
SARS-CoV-2 Spike Pseudotype Neutralization Assay
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SARS-CoV-2 spike-pseudotyped lentivirus-neutralization assays were performed as previously described30 (link), with the following slight modifications. Infections were carried out in poly-l -lysine- (P4707, Millipore Sigma, Burlington, MA, USA) coated black-walled, clear-bottom plates (655090, Greiner Bio-One, Kremsmünster, Austria), and luciferase activity was measured in these plates without transferring to opaque-bottom plates. Sera were diluted 3-fold 7 times starting at a 1:25 dilution, and luciferase activity was measured at 52 h post infection. Target cells were HEK-293T cells transduced to express hACE230 (link) (BEI Resources, NR-52511). Samples were run in duplicate, and each plate included two no-serum controls. Fraction infectivity was calculated by normalizing the luciferase reading for each sample by the average of the two no-serum control wells in the same row. Neutralization curves were plotted using the neutcurve Python package (https://jbloomlab.github.io/neutcurve/ , 0.3.1). This package fits a three-parameter Hill curve, with the top baseline fixed to one and the bottom baseline fixed to zero. The neutralization curves are shown in Supplementary Fig. 1 .
3
SARS-CoV-2 Infection Model in ACE2-Expressing Cells
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Vero E6 and HEK 293T cells expressing human ACE2 cells (NR-52511, BEI Resources, NIAID, NIH, RRID: CVCL_A7UK) were cultured in complete media containing Dulbecco’s modified Eagle medium (#CC3004, Cell Clone, Genetix, New Delhi, India) with 10% CELLECT FBS GOLD (#2916754, MP Biomedicals, Santa Ana, CA, USA), 100 IU/mL Penicillin, 100 μg/mL Streptomycin, and 0.25 μg/mL Amphotericin-B (Antibiotic-Antimycotic Solution, #A5955, Sigma-Aldrich, St. Louis, MO, USA).
SARS-CoV-2 isolates (Hong Kong/VM20001061/2020, NR-52282; hCoV-19/USA/MD-HP05285/2021, NR-55671 (Delta Variant)) were obtained from BEI Resources, the NIAID, and the NIH and were propagated and tittered by using standard plaque assay in Vero E6 cells. The Omicron variant was isolated from the patient sample from the South Karnataka region, India, and was confirmed using S gene target failure PCR. All the experiments pertaining to live SARS-CoV-2 were performed in a viral biosafety level 3 laboratory at CIDR, IISc, Bangalore.
SARS-CoV-2 isolates (Hong Kong/VM20001061/2020, NR-52282; hCoV-19/USA/MD-HP05285/2021, NR-55671 (Delta Variant)) were obtained from BEI Resources, the NIAID, and the NIH and were propagated and tittered by using standard plaque assay in Vero E6 cells. The Omicron variant was isolated from the patient sample from the South Karnataka region, India, and was confirmed using S gene target failure PCR. All the experiments pertaining to live SARS-CoV-2 were performed in a viral biosafety level 3 laboratory at CIDR, IISc, Bangalore.
4
Maintenance of HEK293T and HEK293T-ACE2 Cells
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HEK293T cells (CRL-11268, CVCL_1926, ATCC, Manassas, VA) and HEK293T-ACE2 cells (NR-52511, BEI Resources, ATCC, Manassas, VA) were maintained in Dulbeco’s Modified Eagles Medium (Gibco, 11965–092, ThermoFisher Scientific, Waltham, MA) supplemented with 10% (v/v) fetal bovine serum (F1051, Sigma-Aldrich, St. Louis, MO) and 1% (v/v) penicillin/streptomycin (SV30010, HyClone Laboratories Inc., Logan, UT). Cells were maintained in at 37°C and 5% CO2.
5
Cell Culture of Human Derived Cell Lines
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All cell lines were incubated at 37°C in the presence of 5% CO2. 293-6E (human female, RRID:CVCL_HF20) and 293T cells (human female, RRID:CVCL_0063) cells were maintained in Freestyle 293 media with gentle shaking. HEK-293T-hACE2 (human female, BEI Resources Cat# NR-52511) were maintained in DMEM containing 10% FBS, 2 mM L-glutamine, 100 U/ml penicillin, and 100 μg/ml streptomycin (cDMEM).
6
SARS-CoV-2 Propagation and Titration
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Authenticated (relevant documentation regarding authentication by suppliers is available in Supplemental Data) HEK 293T cells expressing human ACE2 (NR-52511, BEI Resources, NIAID, NIH, RRID: CVCL_A7UK) and VeroE6 cells (CRL-1586, ATCC, RRID:CVCL_0574) were cultured in complete media containing Dulbecco's modified Eagle medium (12100-038, Gibco) with 10% HI-FBS (16140-071, Gibco), 100 IU/ml Penicillin, 100 μg/ml Streptomycin and 0.25μg/ml Amphotericin-B (Penicillin-Streptomycin-Amphotericin B, ICN1674049, MP Biomedicals) supplemented with GlutaMAX™ (35050-061, Gibco). SARS-CoV2 (Isolate Hong Kong/VM20001061/2020, NR-52282, BEI Resources, NIAID, NIH) was propagated and titered by plaque assay in Vero E6 cells as described before [19] (link).
7
SARS-CoV-2 Entry Receptor Expression
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The following cell lines were used in this study, namely, HEK293 (ATCC), HEK293T-ACE2 (HEK293T cells stably expressing human angiotensin-converting enzyme 2) (BEI Resources NR-52511, NIAID, NIH. RRID: CVCL_A7UK ) and Vero-E6 cells (CRL-1586, ATCC, RRID: CVCL_0574 ). The authenticity of HEK293T-ACE2 and Vero-E6 cell lines was confirmed by Certificate of Analysis from their respective sources. HEK293T-ACE2 are human embryonic kidney 293T cells that express the human ACE2 receptor, which is required for SARS-CoV-2 entry. HEK293T-ACE2 and Vero E6 cells are of human and primate origin respectively, and express ACE2 receptor. All cell lines tested negative for mycoplasma contamination. Cells were cultured in complete media prepared using Dulbecco’s modified Eagle medium (12100–038, Gibco) supplemented with 10% HI-FBS (16140–071, Gibco), 100 U/mL Penicillin-Streptomycin (15140122, Gibco) and GlutaMAX (35050–061, Gibco).
SARS-CoV2 (Isolate Hong Kong/VM20001061/2020, NR-52282, BEI Resources, NIAID, NIH) was propagated and quantified by plaque assay in Vero-E6 cells as described before (Case et al., 2020 (link)).
SARS-CoV2 (Isolate Hong Kong/VM20001061/2020, NR-52282, BEI Resources, NIAID, NIH) was propagated and quantified by plaque assay in Vero-E6 cells as described before (Case et al., 2020 (link)).
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