Heat inactivated fbs

Manufactured by Corning

Heat-inactivated FBS is a serum product derived from fetal bovine blood. It has been processed through heat inactivation to denature proteins and inactivate potential contaminants. This product is commonly used as a supplement in cell culture media to support the growth and maintenance of various cell types.

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Lab products found in correlation

6 protocols using heat inactivated fbs

Culturing B3Z and DC2.4 Cell Lines

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The hybridoma CD8⁺ T cell line
B3Z was generously provided by Dr. J. Woodward (University of Kentucky
Medical Center, KY, USA). B3Z cells were cultured in DMEM (Corning)
supplemented with 10% heat-inactivated FBS (Corning), 100 unit/mL
penicillin, 100 μg/mL streptomycin, 0.292 mg/mL l-glutamine
(Gibco), 0.1 mM non-essential amino acids (Gibco), and 1 mM sodium
pyruvate (Gibco). The immature dendritic cell line DC2.4 was purchased
from Millipore Sigma. DC2.4 cells were cultured in RPMI 1640 medium
(Gibco) supplemented with 10% FBS (Corning), 2 mM l-glutamine
(Gibco), 0.1 mM non-essential amino acids (Gibco), 10 mM HEPES buffer
(Gibco), and 0.5 mM β-mercaptoethanol (Fisher Scientific)
for a maximum of 10 passages. Cells were cultured at 37 °C with
5% CO₂.

Harvey B.T., Fu X., Li L., Neupane K.R., Anand N., Kolesar J.M, & Richards C.I. (2022). Dendritic Cell Membrane-Derived Nanovesicles for Targeted T Cell Activation. ACS Omega, 7(50), 46222-46233.

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Characterization of SARS-CoV-2 Infection in Cell Lines

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African green monkey (Cercopithecus aethiops) kidney epithelial cells (Vero E6 cells; ATCC CRL-1586) were maintained in complete DMEM containing 1× DMEM supplemented with 25 mM HEPES, 2 mM l-glutamine,1 mM sodium pyruvate, 1× non-essential amino acids, 1× antibiotic/antimycotic solution (all from Corning Life Sciences) and 10% heat-inactivated FBS (Life Technologies), unless indicated otherwise. Human lung adenocarcinoma epithelial cells (Calu-3 cells; ATCC HTB-55) were maintained in complete MEM (cMEM) containing 1× MEM (Corning Life Sciences) supplemented with 1× antibiotic/antimycotic solution and 10% heat-inactivated FBS unless indicated otherwise. Primary leukocytes from the airways of severe COVID-19 patients were collected bedside via endotracheal aspiration, and whole blood was collected by standard venipuncture, then processed as previously described (D.J. Eddins et al., manuscript posted on bioRxiv, DOI: 10.1101/2021.06.02.446468). SARS-CoV-2 USA-WA1/2020 (hereafter SCV2-WA1) was provided by BEI Resources (Manassas, VA, USA). Virus was propagated in Vero E6 cells as previously described (15 , 31 ), and titer was determined by 50% tissue culture infective dose (TCID₅₀/ml) or plaque assays (PFU/ml). Low-passage (P1 or P2) virus stocks were used throughout this study.

Eddins D.J., Bassit L.C., Chandler J.D., Haddad N.S., Musall K.L., Yang J., Kosters A., Dobosh B.S., Hernández M.R., Ramonell R.P., Tirouvanziam R.M., Lee F.E., Zandi K., Schinazi R.F, & Ghosn E.E. (2022). Inactivation of SARS-CoV-2 and COVID-19 Patient Samples for Contemporary Immunology and Metabolomics Studies. ImmunoHorizons, 6(2), 144-155.

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Characterization of SARS-CoV-2 Infection in Cell Lines

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T Cell Differentiation Protocols

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Naïve CD4 T cells were isolated from mouse spleens and lymph nodes by magnetic bead separation (Miltenyi Biotec) following the manufacturers’ protocols. Cells were cultured in glucose-free RPMI (Gibco) supplemented with 10% heat-inactivated FBS (Corning), 1% penicillin/streptomycin (Gibco), 50μM 2-Mercaptoethanol (Gibco), and either 10mM glucose (Sigma) or 10mM galactose (Sigma). Naïve CD4 T cells were activated with plate-bound anti-CD3e (5μg/ml) and soluble anti-CD28 (2μg/ml) in the presence of mIL-1β (10 ng/ml; R&D Systems), mIL-23 (10ng/ml; R&D Systems), mIL-6 (50ng/ml; R&D Systems), and h TGF-β (5ng/ml; Peprotech) for T_H17 cell polarization; mIL-12 (10ng/ml; R&D Systems) for T_H1 cell polarization, or TGF-β (10ng/ml; Peprotech) and mIL-2 (10ng/ml; R&D Systems) for T_reg polarization. All cells were cultured at 37°C and 5% CO₂. T_H17 differentiation was verified on day 3 or 4, and cytokine expression on day 5, at which point cells were used for subsequent assays.

Hong H.S., Mbah N.E., Shan M., Loesel K., Lin L., Sajjakulnukit P., Correa L.O., Andren A., Lin J., Hayashi A., Magnuson B., Chen J., Li Z., Xie Y., Zhang L., Goldstein D.R., Carty S.A., Lei Y.L., Opipari A.W., Argüello R.J., Kryczek I., Kamada N., Zou W., Franchi L, & Lyssiotis C.A. (2022). OXPHOS Promotes Apoptotic Resistance and Cellular Persistence in TH17 cells in the Periphery and Tumor Microenvironment. Science immunology, 7(77), eabm8182.

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PBMC Isolation and Cryopreservation Protocol

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Approximately 50 mL of blood was drawn by venipuncture into heparinized tubes (Vacutainer, BD Biosciences). PBMCs were isolated by density gradient centrifugation with Ficoll-Hypaque medium (Amersham) and resuspended in a volume of RPMI 1640 medium (Corning) supplemented with 10% heat-inactivated FBS (NOTACOR). Cells were then cryopreserved with an equal volume of freezing media containing 20% DMSO and 80% FBS and stored in liquid nitrogen until use. For serum separation, blood was allowed to coagulate at room temperature and centrifuged at 2000 rpm for 10 min. Then, serum was aliquoted and stored at -70°C until use. Cell viability was evaluated by trypan blue staining (80–95% viable cells/sample) prior to use. Due to sample availability, the assays were not run for all samples.

Natale M.A., César G.A., Alvarez M.G., Castro Eiro M.D., Lococo B., Bertocchi G., Albareda M.C, & Laucella S.A. (2018). Trypanosoma cruzi-specific IFN-γ-producing cells in chronic Chagas disease associate with a functional IL-7/IL-7R axis. PLoS Neglected Tropical Diseases, 12(12), e0006998.

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Engineered Chimeric PD-1 Receptor

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A Chimeric PD-1 receptor, P3Z, was constructed by fusing the extracellular and transmembrane domains of human PD-1 to the cytoplasmic domain of human CD3ζ chain, which was stably transduced into HuT78 cells (ATCC) to generate HuT78/P3Z cells. The HEK293/PD-L1 and HEK293/FcγRI cell lines were generated by stable transfection. HuT78/P3Z cells were cultured in complete RPMI1640 (Hyclone) supplemented with 10% heat-inactivated FBS (Corning), and incubated at 37 °C with 5% CO₂. HEK293-based cell lines were cultured in DMEM (Invitrogen) supplemented with 10% FBS (Corning).

Zhang T., Song X., Xu L., Ma J., Zhang Y., Gong W., Zhang Y., Zhou X., Wang Z., Wang Y., Shi Y., Bai H., Liu N., Yang X., Cui X., Cao Y., Liu Q., Song J., Li Y., Tang Z., Guo M., Wang L, & Li K. (2018). The binding of an anti-PD-1 antibody to FcγRΙ has a profound impact on its biological functions. Cancer Immunology, Immunotherapy, 67(7), 1079-1090.

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