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Minimum essential medium eagle

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Minimum Essential Medium Eagle is a cell culture medium developed by Harry Eagle. It provides a balanced salt solution and essential nutrients required for the in vitro maintenance of a variety of cell types.

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Lab products found in correlation

Hsd icr cd 1 female mice, by Inotiv (2 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions) Dual glo luciferase assay, by Promega (1 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions) Prl sv40, by Promega (1 mentions) Microvette 500 k3 edta tubes, by Sarstedt (1 mentions) Trypsin edta, by Corning (1 mentions) Hanks balanced salt solution (hbss), by Corning (1 mentions) Streptomycin, by Corning (1 mentions) Penicillin g, by Corning (1 mentions) Dmem f12 media, by Corning (1 mentions)

5 protocols using minimum essential medium eagle

1

Dual-Glo Luciferase Assay for OR Activation

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The Dual-Glo Luciferase Assay (E2940, Promega) was used to determine the activities of Firefly and Renilla luciferase in Hana3A cells (34 (link)). Firefly luciferase, driven by a cAMP response element promoter (CRE-Luc; Stratagene), was used to determine OR activation levels. Renilla luciferase, driven by a constitutively active SV40 promoter (pRLSV40; Promega), functioned as an internal control for transfection efficiency and cell viability. Hana3A cells were plated on 96-well plates (Nalge Nunc) and incubated overnight in minimum essential medium eagle (Corning) with 10% FBS (ThermoFisher) at 37 °C and 5% CO2. The following day, cells were transfected using Lipofectamine 2000 (Invitrogen). For each 96-well plate, 2.4 μg of pRL-SV40, 2.4 μg of CRE-Luc, 2.4 μg of mouse RTP1S and 12 μg of receptor plasmid DNA were transfected. After transfection (24 h), medium was replaced with 100 μL of odorant solution diluted in Optimal MEM medium (ThermoFisher), and cells were further incubated for 4 h at 37 °C and 5% CO2. The manufacturer’s protocols were followed to measure firefly luciferase and renilla luciferase activities by using MD SPECTRAMAX L plate reader.
Xu R., Cong X., Zheng Q., Xu L., Ni M.J., de March C.A., Matsunami H., Golebiowski J., Ma M, & Yu Y. (2022). Interactions among key residues regulate mammalian odorant receptor trafficking. FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 36(7), e22384.
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2

West Nile Virus Infection in Mice

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Experimental infections in mice were performed in biosafety level 3 (BSL-3) facilities at Centro de Investigación en Sanidad Animal, Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (CISA, INIA-CSIC). Animals were handled in strict accordance with the guidelines of the European Community 86/609/CEE. A total of 59 six-week-old Hsd:ICR(CD-1) female mice (Envigo; Inotiv) were used (29 uninfected and 30 WNV-infected). Mice were infected intraperitoneally (i.p.) with 104 plaque forming units (PFU) of WNV NY99 (GenBank: KC407666.1) in 200 µL of Minimum Essential Medium Eagle (Corning). Mock-infected animals were inoculated i.p. with the same volume of culture medium. Animals were kept with ad libitum access to food and water, and daily monitored for weight and clinical signs. At different days post-infection (dpi), a representative number of animals (8-10 mice per group) was anesthetized under isoflurane, and humanly sacrificed. Blood from submandibular vein, and left hemispheres of brain and cerebellum from euthanized mice, were collected at 3- (n = 9 uninfected and n = 10 infected), 7- (n = 10 uninfected and n = 10 infected), and 10-days post infection (dpi) (n = 10 uninfected and n = 8 infected). Plasma was separated using Microvette 500 K3 EDTA tubes (Sarstedt) by centrifugation (1123 × g, 15 min at 4°C). All samples were frozen at −80°C until analyses.
Mingo-Casas P., Sanchez-Céspedes J., Blázquez A.B., Casas J., Balsera-Manzanero M., Herrero L., Vázquez A., Pachón J., Aguilar-Guisado M., Cisneros J.M., Saiz J.C, & Martín-Acebes M.A. (2023). Lipid signatures of West Nile virus infection unveil alterations of sphingolipid metabolism providing novel biomarkers. Emerging Microbes & Infections, 12(2), 2231556.
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3

Experimental West Nile Virus Infections in Mice

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Experimental infections in mice were performed in the biosafety level 3 (BSL-3) facilities at Centro de Investigación en Sanidad Animal, Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (CISA, INIA-CSIC). Six-week-old Hsd:ICR(CD-1) female mice (Inotiv) were used. Mice were infected intraperitoneally (i.p.) with 104 PFU of WNV New York 99 strain [14 (link)] (GenBank: KC407666.1) in 200 µL of Minimum Essential Medium Eagle (Corning); mock-infected animals were also inoculated i.p. with 200 µL of culture media. In the case of drug treatments, drugs or vehicle (saline buffer, 0.9% NaCl) were intraperitoneally administered at a dose of 200 mg/kg for DCA or 500 mg/kg in the case of 2-DG. Mice were daily injected (QD) from the first day of infection up to day 6 post-infection. Animals were kept with ad libitum access to food and water and were anesthetized under isoflurane and humanely killed at 7 days post-infection.
Mingo-Casas P., Blázquez A.B., Gómez de Cedrón M., San-Félix A., Molina S., Escribano-Romero E., Calvo-Pinilla E., Jiménez de Oya N., Ramírez de Molina A., Saiz J.C., Pérez-Pérez M.J, & Martín-Acebes M.A. (2023). Glycolytic shift during West Nile virus infection provides new therapeutic opportunities. Journal of Neuroinflammation, 20, 217.
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4

Isolation and Expansion of Amniotic Membrane Cells

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AMCs were isolated as described in our prior reports [14 , 40 (link)]. Briefly, the amnion (~10 g) was peeled from the chorion layer and rinsed 3 times in sterile Hanks’ Balanced Salt Solution (HBSS; Cat# 21–021-CV, Corning) to remove blood debris. The sample was then incubated with 0.05% trypsin/EDTA (Cat# 25–053-CI, Corning) for 1 hour at 37°C (water bath) to disperse the cells and remove the epithelial cell layer. The membrane pieces were then washed 3 times using cold HBSS to inactivate the enzyme. The washed membrane was then transferred into a second digestion buffer containing Minimum Essential Medium Eagle (Cat# 10–010-CV, Corning), 1-mg/mL collagenase type IV, and 25-μg/mL DNase I and incubated in a rotator at 37°C for 1 hour. The digested membrane solution was neutralized using Dulbecco’s Modified Eagle’s Medium (DMEM)/F12 media (Cat# 16–405-CV, Corning), filtered using a 70-μm cell strainer, and centrifuged at 3000 rpm for 10 minutes. The cell pellet was resuspended in complete DMEM/F12 media supplemented with 5% heat-inactivated FBS (Cat# 35–010-CV, Corning), 100-U/mL penicillin G, and 100-mg/mL streptomycin (Cat# 30–001-CI, Corning); plated at 3–5 million cells per T75; and incubated at 37°C with 5% CO 2 until they were 80%–90% confluent. AMC purity was confirmed with morphological analysis before passaging to P1s.
Richardson L.S., Menon P.R, & Menon R. (2019). The effects of extracellular matrix rigidity on 3-dimensional cultures of amnion membrane cells. Placenta, 90, 82-89.
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5

Experimental West Nile Virus Infections in Mice

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Experimental infections in mice were performed in the biosafety level 3 (BSL-3) facilities at Centro de Investigación en Sanidad Animal, Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (CISA, INIA-CSIC). Six-week-old Hsd:ICR(CD-1) female mice (Inotiv) were used. Mice were infected intraperitoneally (i.p.) with 104 PFU of WNV New York 99 strain [14 (link)] (GenBank: KC407666.1) in 200 µL of Minimum Essential Medium Eagle (Corning); mock-infected animals were also inoculated i.p. with 200 µL of culture media. In the case of drug treatments, drugs or vehicle (saline buffer, 0.9% NaCl) were intraperitoneally administered at a dose of 200 mg/kg for DCA or 500 mg/kg in the case of 2-DG. Mice were daily injected (QD) from the first day of infection up to day 6 post-infection. Animals were kept with ad libitum access to food and water and were anesthetized under isoflurane and humanely killed at 7 days post-infection.
Mingo-Casas P., Blázquez A.B., Gómez de Cedrón M., San-Félix A., Molina S., Escribano-Romero E., Calvo-Pinilla E., Jiménez de Oya N., Ramírez de Molina A., Saiz J.C., Pérez-Pérez M.J, & Martín-Acebes M.A. (2023). Glycolytic shift during West Nile virus infection provides new therapeutic opportunities. Journal of Neuroinflammation, 20, 217.
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