Vero 76

Manufactured by American Type Culture Collection

Sourced in United States

The Vero 76 is a cell line derived from the kidney of an African green monkey. It is a commonly used model system for virus propagation and cytotoxicity assays.

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Vero 76 cells (69 citations)

33 protocols using vero 76

Cell Culture and Transfection Protocol

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HEK293T and Vero 76 cells (both from ATCC) were cultured in Dulbecco's modified Eagle's medium (DMEM) with GlutaMAX (Gibco) supplemented with 10% fetal bovine serum (FBS) (Sigma-Aldrich). Cell lines were tested for mycoplasma contamination after receipt, before expansion and cryopreservation. For studies including nonhuman primate samples, Vero 76 and Vero CCL81 cells (both from ATCC) were cultured in DMEM (Gibco) containing 2% HyClone fetal bovine and 100 U ml -1 penicillium-streptomycin (Gibco). Expi293F cells were grown in Expi293 medium and transiently transfected using Expi-Fectamine293 (all from Thermo Fisher Scientific).

Vogel A.B., Kanevsky I., Che Y., Swanson K.A., Muik A., Vormehr M., Kranz L.M., Walzer K.C., Hein S., Güler A., Loschko J., Maddur M.S., Ota-Setlik A., Tompkins K., Cole J., Lui B.G., Ziegenhals T., Plaschke A., Eisel D., Dany S.C., Fesser S., Erbar S., Bates F., Schneider D., Jesionek B., Sänger B., Wallisch A.K., Feuchter Y., Junginger H., Krumm S.A., Heinen A.P., Adams-Quack P., Schlereth J., Schille S., Kröner C., de la Caridad Güimil Garcia R., Hiller T., Fischer L., Sellers R.S., Choudhary S., Gonzalez O., Vascotto F., Gutman M.R., Fontenot J.A., Hall-Ursone S., Brasky K., Griffor M.C., Han S., Su A.A.H., Lees J.A., Nedoma N.L., Mashalidis E.H., Sahasrabudhe P.V., Tan C.Y., Pavliakova D., Singh G., Fontes-Garfias C., Pride M., Scully I.L., Ciolino T., Obregon J., Gazi M., Carrion R J.r., Alfson K.J., Kalina W.V., Kaushal D., Shi P.Y., Klamp T., Rosenbaum C., Kuhn A.N., Türeci Ö., Dormitzer P.R., Jansen K.U, & Sahin U. (2021). BNT162b vaccines protect rhesus macaques from SARS-CoV-2. Nature, 592(7853).

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Cell Culture and Virus Propagation

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HEK293T (human) (CRL-11268; American Type Culture Collection [ATCC]), NIH3T3 (mouse) (CRL-1658; ATCC), MDTF (mouse), CRFK (cat) (CCL-94; ATCC), FEA (cat) (feline embryonic fibroblasts), Cf2Th (dog) (CRL-1430; ATCC), A72 (dog) (CRL-1542; ATCC), Mpf (ferret) (CRL-1656; ATCC), PK15 (pig) (CCL-33; ATCC), Vero 76 (CRL-1587; ATCC), and BHKT7/9 (a hamster kidney–derived BHK cell clone stably expressing T7 RNA polymerase) (31 (link)) cells were cultured in Dulbecco’s modified Eagle's medium (Sigma–Aldrich), supplemented with 10% heat-inactivated fetal calf serum and antibiotics (Thermo Fisher Scientific). The SFTSV YG1 strain, a field isolate from an SFTS patient in Japan, was kindly provided by Dr Ken Maeda, NIID. Virus stocks were prepared from the culture supernatants of Vero 76 cells.

Yoshikawa R., Kawakami M, & Yasuda J. (2023). The NSs protein of severe fever with thrombocytopenia syndrome virus differentially inhibits the type 1 interferon response among animal species. The Journal of Biological Chemistry, 299(6), 104819.

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Comparative Study of Diverse Cell Lines

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Human neuroblastoma cell line SK-N-AS (ATCC # CRL-2137™), African green monkey kidney epithelial cell line Vero 76 (ATCC # CRL-1587™), embryonic fibroblasts from the NIH/Swiss mouse 3T3 (ATCC # CRL-1658™) and brain neuroblastoma line from strain A albino mouse Neuro2A (ATCC # CCL-131™) were obtained from American Type Culture Collection (USA). SK-N-AS cells were cultured in DMEM with 10% FBS and 1% of penicillin/streptomycin. Vero76 cells were cultured in DMEM with 5% FBS and 1% of penicillin/streptomycin. 3T3 cells were cultured in Dulbecco's Modified Eagle's Medium with 10% FBS and 1% of penicillin/streptomycin. Neuro-2A cells were cultured in Eagle's Minimum Essential Medium with 10% FBS and 1% of penicillin/streptomycin. Experiments were performed within the first 15 passages after receipt form ATCC.

Hsiao J., Yuan T.Y., Tsai M.S., Lu C.Y., Lin Y.C., Lee M.L., Lin S.W., Chang F.C., Liu Pimentel H., Olive C., Coito C., Shen G., Young M., Thorne T., Lawrence M., Magistri M., Faghihi M.A., Khorkova O, & Wahlestedt C. (2016). Upregulation of Haploinsufficient Gene Expression in the Brain by Targeting a Long Non-coding RNA Improves Seizure Phenotype in a Model of Dravet Syndrome. EBioMedicine, 9, 257-277.

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Infection Protocols for VEEV and CHIKV

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Baby hamster kidney (BHK) clone 21 cells (ATCC CCL-10) and Vero 76 (ATCC® CRL-1587™) were maintained in Modified Eagle’s Medium with Earle’s Balanced Salt Solution and L-glutamine (MEM-E) supplemented with 10% fetal bovine serum (FBS) (Corning CellGro). Cells were maintained at 37 °C in humidified incubators with 5% CO₂. VEEV strain TC-83 (gift of Dr. Connie Schmaljohn, USAMRIID) was used for this study. The strain V3526 was generated from a plasmid as described previously (Chung et al., 2014 (link)). Infections were carried out using a virus infection medium (Modified Eagle’s Medium with Earle’s Balanced Salt Solution supplemented with 1x GlutaMAX, 25 mM HEPES, and 10% FBS. For Chikungunya virus (CHIKV) experiments, CHIKV strain 181/25 (BEI Resources NR-13222) was used. V3526 experiments were carried out in an infection media that contained L-glutamine instead of GlutaMAX (Modified Eagle’s Medium with Earle’s Balanced Salt Solution with L-glutamine, 25 mM HEPES, 10% FBS)

Skidmore A.M., Adcock R.S., Jonsson C.B., Golden J.E, & Chung D.H. (2019). Benzamidine ML336 inhibits plus and minus strand RNA synthesis of Venezuelan equine encephalitis virus without affecting host RNA production.. Antiviral research, 174, 104674.

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Culturing Coronavirus Cell Lines for Research

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Cell lines were purchased from the American Type Culture Collection (ATCC). The absence of mycoplasma contamination was checked periodically by the Hoechst staining method. Cell lines supporting the multiplication of Coronaviruses were the following: Monkey kidney (Vero-76) [ATCC CRL 1587 Cercopithecus Aethiops], Monkey kidney (Vero C1008, clone E6) [ATCC CRL 1586 Cercopithecus Aethiops].
Human coronaviruses were: (i) Coronaviridae: Betacoronavirus strain OC43 (ATCC VR-1558), Alphacoronavirus strain 229E (ATCC VR-740), Betacoronavirus SARS-CoV-2 (strain VR PV10734) clinical isolate, kindly provided by Lazzaro Spallanzani Hospital, Rome, Italy. All experimental steps involving the SARS-CoV-2 virus were performed in a biosafety level 3 (BSL3) containment laboratory.

Manca V., Chianese A., Palmas V., Etzi F., Zannella C., Moi D., Secci F., Serreli G., Sarais G., Morone M.V., Galdiero M., Onnis V., Manzin A, & Sanna G. (2024). Exploring the Antiviral Potential of Esters of Cinnamic Acids with Quercetin. Viruses, 16(5), 665.

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Antiviral Activity Evaluation in Cell Lines

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Antiviral activity was evaluated in Caco-2 (human colon carcinoma; American Type Culture Collection (ATCC) HTB-37), Vero-76 (African green monkey (Chlorocebus aethiops) kidney; ATCC CRL-1587), and Calu-3 (human lung adenocarcinoma) cells.

Taylor R., Bowen R., Demarest J.F., DeSpirito M., Hartwig A., Bielefeldt-Ohmann H., Walling D.M., Mathis A, & Babu Y.S. (2021). Activity of Galidesivir in a Hamster Model of SARS-CoV-2. Viruses, 14(1), 8.

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Comprehensive Virus Cell Line Characterization

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All cell lines were obtained from American Type Culture Collection (ATCC): CD4+ human T-cells (MT-4), Madin-Darby bovine kidney (MDBK) (ATCC CCL 22 (NBL- 1) Bos taurus), baby hamster kidney (BHK-21) (ATCC CCL 10 (C-13) Mesocricetus auratus), and monkey kidney (Vero-76) (ATCC CRL 1587 Cercopithecus aethiops).
Human immunodeficiency virus type-1 (HIV-1) IIIB laboratory strain was derived from supernatant of H9/IIIB cells (NIH 1983). Viruses representative of ssRNA+, of ssRNA-, and of DNA virus were, respectively, (i) yellow fever virus (YFV) (strain 17-D vaccine (Stamaril Pasteur J07B01)), bovine viral diarrhea virus (BVDV) (strain NADL (ATCC VR-534)), west Nile virus (WNV) (clinical isolate), dengue virus (DENV-2) (clinical isolate), coxsackie type B5 (CV-B5), strain Faulkner (ATCC VR-185), and poliovirus type-1 (Sb-1), Sabin strain Chat (ATCC VR-1562); (ii) human respiratory syncytial virus (hRSV) (strain A2 (ATCC VR-1540)) and vesicular stomatitis virus (VSV) (lab strain Indiana (ATCC VR 1540)); and (iii) vaccinia virus (VV) (vaccine strain Elstree-Lister (ATCC VR-1549)) and human herpes 1 (HSV-1) (strain KOS (ATCC VR- 1493)).

Madeddu S., Marongiu A., Sanna G., Zannella C., Falconieri D., Porcedda S., Manzin A, & Piras A. (2021). Bovine Viral Diarrhea Virus (BVDV): A Preliminary Study on Antiviral Properties of Some Aromatic and Medicinal Plants. Pathogens, 10(4), 403.

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Cell Lines for Protein Production

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Spodoptera frugiperda Sf9 cells, Drosophila S2 cells, Expi-CHO cells and HEK-293F cells were purchased from Thermo Fisher Scientific. HEK-293T (CRL-3216) were acquired from the American Type Culture Collection (ATCC). Vero (CCL-81) and Vero 76 (CRL-1587) cells were also obtained from ATCC. HEK-293T cells stably expressing like-acetylglucosaminyltransferase (LARGE, UniProt: O95461) were generated using puromycin selection.

Eis-Hübinger A.M., Däumer M., Matz B, & Schneweis K.E. (1999). Evaluation of Three Glycoprotein G2-Based Enzyme Immunoassays for Detection of Antibodies to Herpes Simplex Virus Type 2 in Human Sera. Journal of Clinical Microbiology, 37(5), 1242-1246.

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Plaque Assay for SARS-CoV-2 Quantification

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Plaque titrations for SARS-CoV-2 were conducted as previously described (26 (link)). Briefly, 10-fold dilutions of clarified tissue homogenate, or stock virus, starting at a 1:10 dilution were adsorbed for 1 h on Vero 76 (ATCC CRL-1587) cell monolayers in 6-well tissue culture plates and then overlaid with 0.6% agarose. After 2 days (day 2) in a 37°C, 5% CO₂ incubator, an overlay containing 5% neutral red was added. On day 3, plaques were counted and the number of PFU per milliliter was calculated. The number of PFU per gram of lung was calculated as the PFU per milliliter of lung homogenate divided by the mass of lung tissue per volume of homogenate (in grams per milliliter).

Golden J.W., Li R., Cline C.R., Zeng X., Mucker E.M., Fuentes-Lao A.J., Spik K.W., Williams J.A., Twenhafel N., Davis N., Moore J.L., Stevens S., Blue E., Garrison A.R., Larson D.D., Stewart R., Kunzler M., Liu Y., Wang Z, & Hooper J.W. (2022). Hamsters Expressing Human Angiotensin-Converting Enzyme 2 Develop Severe Disease following Exposure to SARS-CoV-2. mBio, 13(1), e02906-21.

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RSV Infection in Primate and Human Cells

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Cell lines and RSV were purchased from American Type Culture Collection (ATCC). The absence of mycoplasma contamination was checked periodically by the Hoechst staining method. Cell line supporting the multiplication of RSV was the Monkey kidney (Vero 76) [ATCC CRL 1587 Cercopithecus Aethiops], while CD4⁺human T cells containing an integrated HTLV-1 genome (MT-4) was used as human cellular model. Human respiratory syncytial virus (RSV) [strain A2 (ATCC VR-1302)].

Cichero E., Tonelli M., Novelli F., Tasso B., Delogu I., Loddo R., Bruno O, & Fossa P. (2017). Benzimidazole-based derivatives as privileged scaffold developed for the treatment of the RSV infection: a computational study exploring the potency and cytotoxicity profiles. Journal of Enzyme Inhibition and Medicinal Chemistry, 32(1), 375-402.

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