Net328250uc

Manufactured by PerkinElmer

Sourced in United States

The NET328250UC is a laboratory instrument designed for sample processing. It features a compact and modular design to accommodate various sample types and volumes. The core function of this product is to facilitate efficient sample preparation and handling in a controlled laboratory environment.

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Lab products found in correlation

Net328a250uc, by PerkinElmer (2 mentions) Cacl2, by Merck Group (2 mentions) Hgcl2, by Merck Group (2 mentions) Sodium chloride (nacl), by Merck Group (1 mentions) Potassium chloride (kcl), by Merck Group (1 mentions) α tubulin clone b 5 1 2, by Merck Group (1 mentions) 3h 2 deoxyglucose, by PerkinElmer (1 mentions) Phospho p70 s6 kinase thr389, by Cell Signaling Technology (1 mentions) Goat anti rabbit 111 035 003, by Jackson ImmunoResearch (1 mentions) Oil red o o0625, by Merck Group (1 mentions) Mgcl2, by Merck Group (1 mentions)

4 protocols using net328250uc

Hippocampal Glucose Uptake Assay

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Hippocampal slices were washed with buffer (15 mM HEPES (cat: H3375), 135 mM NaCl (cat: s3014), 5 mM KCl (cat: P5405), 1.8 mM CaCl₂ (cat: C1016), and 0.8 mM MgCl₂ (cat: 208337) supplemented with 0.5 mM glucose (Cisternas et al., 2014a (link)). Then, the slices were incubated for 0–90 min with 1–1.2 μCi 2-[1,2-³H(N)]-deoxy-D-glucose (cat: NET328250UC, PerkinElmer, United States) or ³H-2-deoxyglucose (cat: NET328A250UC, PerkinElmer, United States) at a final specific activity of 1–3 disintegrations/min/pmol (∼1 mCi/mmol). Glucose uptake was arrested by washing the cells with ice-cold PBS supplemented with 1 mM HgCl₂ (cat: 203777, Sigma-Aldrich, United States). The incorporated radioactivity was quantified by liquid scintillation counting.

Cisternas P., Oliva C.A., Torres V.I., Barrera D.P, & Inestrosa N.C. (2019). Presymptomatic Treatment With Andrographolide Improves Brain Metabolic Markers and Cognitive Behavior in a Model of Early-Onset Alzheimer’s Disease. Frontiers in Cellular Neuroscience, 13, 295.

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Hippocampal Slice Preparation for Glucose Uptake Assay

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Hippocampal slices were prepared according to standard procedures. Briefly, we sectioned transverse slices (350 μm) from the dorsal hippocampus in cold artificial cerebrospinal fluid (ACSF; 124 mM NaCl, 26 mM NaHCO₃, 3 mM D- glucose, 2.69 mM KCl, 1.25 mM KH₂PO₄, 2.5 mM CaCL₂, 1.3 mM MgSO₄, and 2.60 mM NaHPO₄) using a vibratome (LeicaVT 1000s). Slices were incubated in ACSF for 1 h at room temperature, then were washed with washing buffer (15 mM HEPES [#H3375, Sigma Millipore], 135 mM NaCl [#s3014, Sigma-Aldrich], 5 mM KCl [#P5405 Sigma-Aldrich], 1.8 mM CaCl₂ (#C1016 Sigma-Aldrich), and 0.8 mM MgCl₂ [#208337, Sigma-Aldrich]) supplemented with 0.5 mM glucose. Then, the slices were incubated for 30 min with 1-1.2 μCi 2-[1,2-³H(N)]-deoxy-D-glucose (#NET328250UC, PerkinElmer) at a final specific activity of 1-3 disintegrations/min/pmol (~1 mCi/mmol). Glucose uptake was arrested by washing the cells with ice-cold PBS supplemented with 1 mM HgCl₂ (#203777, Sigma-Aldrich). The incorporated radioactivity was quantified by liquid scintillation counting (45 (link)).

Cisternas P., Gherardelli C., Gutierrez J., Salazar P., Mendez-Orellana C., Wong G.W, & Inestrosa N.C. (2023). Adiponectin and resistin modulate the progression of Alzheimer´s disease in a metabolic syndrome model. Frontiers in Endocrinology, 14, 1237796.

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Biochemical Reagents for Cell Signaling

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Biochemical reagents, such as Cytochalasin B (C-6762), 2-deoxyglucose (D-8375), Dexamethasone (D2915), isobutymethylxanthine (I5879), Insulin (I0516), Rosiglitazone (R2408), BCS (Bovine Calf serum, 12133C), BSA (Bovine Serum Albumin, A9647) and Oil Red O (O0625), were all purchased from Sigma-Aldrich (St. Louis, MO, USA). 2-[3H] deoxyglucose was purchased from Perkin Elmer (NET328250UC). All lentiviral plasmids harboring shRNA sequences (mission shRNA) were obtained from Sigma. Antibodies against β-actin (Sigma-Aldrich, St Louis, MO, USA; a5441), α-tubulin (clone B-5-1-2) and total PKN1 were purchased from Sigma-Aldrich. Phospho-IRS-1 (Tyr895), Phospho-PRK1 (Thr774)/PRK2 (Thr816), Phospho-Akt (Ser473) and Phospho-p70 S6 kinase (Thr 389) were purchased from cell signaling. Peroxidase conjugated goat anti-mouse (115-035-003) and goat anti-rabbit (111-035-003) antibodies were obtained from Jackson Immunoresearch (West Grove, PA, USA).

Herrerías-González F., Yeramian A., Baena-Fustegueras J.A., Bueno M., Fleitas C., de la Fuente M., Serrano J.C., Granado-Serrano A., Santamaría M., Yeramian N., Zorzano-Martínez M., Mora C, & Lecube A. (2023). PKN1 Kinase: A Key Player in Adipocyte Differentiation and Glucose Metabolism. Nutrients, 15(10), 2414.

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Quantification of Glucose Uptake in Cells

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After activator/inhibitor treatments, cells were washed with incubation buffer (15 mM HEPES (cat: H3375), 135 mM NaCl (cat: s3014), 5 mM KCl (cat: P5405), 1.8 mM CaCl₂ (cat: C1016), and 0.8 mM MgCl₂ (cat: 208337), all the reagents were purchased from Sigma-Aldrich, USA) supplemented with 0.5 mM glucose (Cisternas et al. 2014a). Cells were then incubated for 15 s with 1–1.2 μCi 2-deoxy-D-[1,2-(N)3H] (cat: NET328250UC, PerkinElmer, USA) or glucose ([2-³H]-DG (cat: NET328A250UC, PerkinElmer, USA) at a final specific activity of 1–3 disintegrations/min/pmol (~1 mCi/mmol). Glucose uptake was arrested by washing the cells with ice-cold PBS supplemented with 1 mM HgCl₂ (cat: 203777, Sigma-Aldrich, USA). The incorporated radioactivity was quantified by liquid scintillation counting. In separate experiments, brain slices were incubated with different drugs for 1 h, then treated with [2-³H]-DG for 15 min and processed as described above. The kinetic parameters were determined using a single rectangular hyperbola of the form 𝑉max ∗ [glc]/(𝐾𝑚 + [glc]), adjusted to the data by nonlinear regression using SigmaPlot 12 (Barros et al. 2009 (link)).

Cisternas P., Zolezzi J.M., Martinez M., Torres V.I., Wong G.W, & Inestrosa N.C. (2018). Wnt-induced activation of glucose metabolism mediates the in vivo neuroprotective roles of Wnt signaling in Alzheimer disease. Journal of neurochemistry, 149(1), 54-72.

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