Gentamicin

Manufactured by BioLegend

Gentamicin is an antibiotic that inhibits bacterial protein synthesis, effectively killing a wide range of gram-negative and some gram-positive bacteria. It is commonly used in cell culture applications to prevent bacterial contamination.

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Lab products found in correlation

Brefeldin a, by BioLegend (1 mentions) Gentamicin, by Lonza (1 mentions) M csf, by Thermo Fisher Scientific (1 mentions) 100 mm cell culture dishes, by BD (1 mentions) Iscove modified dulbecco media (imdm), by Lonza (1 mentions) Recombinant mouse m csf, by BioLegend (1 mentions) Recombinant mouse il 10, by Thermo Fisher Scientific (1 mentions) Recombinant mouse il 4, by Thermo Fisher Scientific (1 mentions) Ip 10, by R&D Systems (1 mentions) Il 12p70, by BioLegend (1 mentions) Il 23p19, by R&D Systems (1 mentions) Rantes, by R&D Systems (1 mentions) Gentamicin, by Merck Group (1 mentions) Prism software, by GraphPad (1 mentions) Il 1β, by BioLegend (1 mentions) Interleukin 6 (il 6), by Immunotools (1 mentions)

3 protocols using gentamicin

MAIT Cell Activation by Salmonella

Cited 1 time

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PBMCs were seeded in 96-well round-bottom plates (5 to 8 × 10⁵ cells per well) and infected with the different Salmonella strains from frozen midlog phase stocks, at the indicated MOI. Upon 80-min incubation at 37 °C, 100 μg/mL gentamicin (Lonza) was added to kill extracellular bacteria. A total of 200 μg/mL of gentamicin was required for the experiments carried out in Malawi using ST313 strains from lineages 1 and 2.
At 180 min postinfection, 5 ng/mL brefeldin A (BioLegend) solution was added to every well in order to achieve accumulation of intracellular cytokines. Samples were incubated overnight at 37 °C for no more than 15 h.
For MR1 blocking experiments, infection was performed in the presence of 30 μg/mL MR1 blocking antibody 26.5 (42 (link)) or mouse IgG2a isotype control (ATCC). The MAIT agonists 5-A-RU was synthesized as described in ref. 71 (link) and 1 μg/mL was combined to 50 μM MG (Sigma).

Preciado-Llanes L., Aulicino A., Canals R., Moynihan P.J., Zhu X., Jambo N., Nyirenda T.S., Kadwala I., Sousa Gerós A., Owen S.V., Jambo K.C., Kumwenda B., Veerapen N., Besra G.S., Gordon M.A., Hinton J.C., Napolitani G., Salio M, & Simmons A. (2020). Evasion of MAIT cell recognition by the African Salmonella Typhimurium ST313 pathovar that causes invasive disease. Proceedings of the National Academy of Sciences of the United States of America, 117(34), 20717-20728.

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Murine Macrophage Differentiation Protocol

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Bone marrow was collected from 12- to 16-week-old female wild-type C57BL/6 mice (experiments approved by the animal experiments committee of the Academic Medical Center, Amsterdam) and cultured in 100 mm cell culture dishes (BD Falcon) in IMDM (Lonza) supplemented with 10% FBS, 86 μg ml⁻¹ gentamicin and 40 ng ml⁻¹ recombinant mouse M-CSF (Biolegend). At day 3, fresh medium containing M-CSF was added. At day 6, M-CSF and either 10 ng ml⁻¹ recombinant mouse IL-10 or 20 ng ml⁻¹ recombinant mouse IL-4 (both from eBioscience) were added. At day 7, cells were harvested and stimulated as described for human macrophages, using 2 μg ml⁻¹ mouse IgG (Equitech-Bio) coated in Maxisorp plates. Cells were analysed by quantitative RT–PCR as described; primer pairs are listed in Supplementary Table 2.

Vogelpoel L.T., Hansen I.S., Rispens T., Muller F.J., van Capel T.M., Turina M.C., Vos J.B., Baeten D.L., Kapsenberg M.L., de Jong E.C, & den Dunnen J. (2014). Fc gamma receptor-TLR cross-talk elicits pro-inflammatory cytokine production by human M2 macrophages. Nature Communications, 5, 5444.

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Cytokine Production by AIEC-infected MoDCs

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MoDC were obtained from CD14⁺ human monocytes purified from blood of healthy subjects as previously described.^{30 (link)} MoDCs were seeded into round-bottom, 96-well culture plates at a density of 10⁵ cells/well in complete RPMI medium without penicillin/streptomycin and infected with 10⁷ CFU/mL of AIEC-LF82 or E. coli Nissle 1917 strain alone [MOI 10:1 bacteria per phagocytic cell]. Following 1 h of phagocytosis, gentamicin [20 ug/mL, Sigma-Aldrich, Darmstadt, Germany] was added to kill extracellular bacteria. Subsequently, infected MoDC were washed and maintained in RPMI supplemented with gentamicin [2 ug/mL] for additional 22 h. The levels of IL-1β [BioLegend], IL-12p70 [BioLegend], IL-23p19 [R&D Systems], IL-6 [ImmunoTools], IP-10 [R&D Systems], and RANTES [R&D Systems] in supernatants of infected MoDC were analysed after 24 h by ELISA. The ELISA plates were read on a microplate reader [SAFAS MP96], and data were analysed with Prism software [version 7; GraphPad Software, La Jolla, CA].

Paroni M., Leccese G., Ranzani V., Moschetti G., Chiara M., Perillo F., Ferri S., Clemente F., Noviello D., Conforti F.S., Ferrero S., Karnani B., Bosotti R., Vasco C., Curti S., Crosti M.C., Gruarin P., Rossetti G., Conte M.P., Vecchi M., Pagani M., Landini P., Facciotti F., Abrignani S., Caprioli F, & Geginat J. (2023). An Intestinal Th17 Subset is Associated with Inflammation in Crohn’s Disease and Activated by Adherent-invasive Escherichia coli. Journal of Crohn's & Colitis, 17(12), 1988-2001.

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