Tumor cell lines were plated at a concentration of 100,000 cells/ml in the corresponding media. The following day tumor cells were replaced with fresh serum free medium with ACLYi, AKTi (GSK690693), AKTi-1/2 and C38 Mab and then stimulated with α2M* and acetate. Medium samples were collected for the assay. Glucose and lactate were measured using colorimetric kits according to the manufacturer's instructions. High sensitivity Glucose assay kit and Lactose assay kit were obtained from Sigma.
Lactose assay kit
Manufactured by Merck Group
Sourced in Canada
The Lactose Assay Kit is a laboratory tool designed to quantitatively measure the concentration of lactose in various samples. It provides a simple and reliable method for the enzymatic determination of lactose levels.
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Lab products found in correlation
High sensitivity glucose assay kit, by Merck Group (2 mentions)
Alanine assay kit, by Merck Group (1 mentions)
Labassay nefa kit, by Fujifilm (1 mentions)
Tag reagent, by Merck Group (1 mentions)
Ketone body assay, by Merck Group (1 mentions)
Free glycerol reagent, by Merck Group (1 mentions)
Atp assay kit, by Beyotime (1 mentions)
Acetate solution, by Merck Group (1 mentions)
P foxo1, by Cell Signaling Technology (1 mentions)
Gsk690693, by Selleck Chemicals (1 mentions)
Irdye 800cw, by Rockland Immunochemicals (1 mentions)
P irs1, by Cell Signaling Technology (1 mentions)
P gsk3β, by Cell Signaling Technology (1 mentions)
P erk1 2, by Cell Signaling Technology (1 mentions)
Akti 1 2, by Selleck Chemicals (1 mentions)
Gapdh antibody, by GenScript (1 mentions)
Alexa fluor 680, by Thermo Fisher Scientific (1 mentions)
P acly, by Cell Signaling Technology (1 mentions)
Acecs1, by Cell Signaling Technology (1 mentions)
Alexa fluor 647, by Thermo Fisher Scientific (1 mentions)
4 protocols using lactose assay kit
1
Glucose and Lactate Metabolic Assay
2
Comprehensive Metabolic Analysis in Mice
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Blood glucose concentrations were measured through tail veil bleeding with glucose analyzer (GM9, Analox Instruments Ltd, UK). The concentrations of serum triacylglycerides (TAGs) and nonesterified fatty acids (NEFAs) were measured using TAG reagent (Sigma-Aldrich, T2449) and free glycerol reagent (Sigma-Aldrich, F6248), and a Lab Assay NEFA kit (Wako Pure Chemical Industries, Japan, 294-63601), respectively. Serum ketone body concentrations and serum alanine were determined by Ketone Body Assay (Sigma-Aldrich, MAK134) and Alanine assay kit (Sigma-Aldrich, MAK001), respectively. Milk lactose was measured by Lactose Assay Kit (Sigma-Aldrich, MAK017). Pyruvate carboxylase activity was determined by Pyruvate Carboxylase Activity Assay Kit (Boxbio, AKSU070M). Brain ATP levels were determined by ATP Assay Kit (Beyotime, S0026).
3
Profiling Acetylation and Phosphorylation
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Antibodies recognizing Acetyl-Histone H2A, H2B, H3 and H4, Hisotne H2A, H2B, H3 and H4, P-ACLY (Ser455), ACLY, ACeCS1, P-Akt (Ser473), P-Akt (Thr308), AKT, P-IRS1(Ser612), IRS1, P-FoxO1(Thr24)/FoxO3a(Thr32), P-ERK1/2(Thr202/Tyr204), ERK1/2, P-PRAS40(T246), PRAS40, P-GSK3β(S21), GSK3β, Acetyl-Stat3(Lys685), Acetyl-β-catenin (Lys49), Acetyl-α-Tubulin (Lys40), were purchased from Cell Signaling Technologies. GAPDH antibody was purchased from Genscript. Secondary antibodies conjugated with Alexa fluor 680, Alexa fluor 790, and Alexa fluor 647 were purchased from Invitrogen. IR dye 800 CW was purchased from Rockland. Acetate solution, High sensitivity Glucose assay kit and Lactose assay kit were purchased from Sigma-Aldrich. SB-204990, GSK690693 and AKTi-1/2 were purchased from Selleckchem. α2M* was prepared as described previously [59 (link)]. GRP78 murine monoclonal antibody (C38) was produced in our laboratory [35 (link)].
4
Biochemical Markers of Metabolism Assays
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Serum urea concentration was determined using the QuantiChrom urea assay kit (BioAssay Systems, Hayward, CA) according to the manufacturer's instructions. The intra-and interassay CV were 3.2 and 2.8%, respectively. Plasma glucose concentration was determined as previously described (Loiselle et al., 2009) using a glucose assay kit (Sekisui Diagnostics P.E.I. Inc., Charlottetown, PE, Canada) with a modification of the glucose oxidase/peroxidase method described by Trinder (1969) . The intra-and inter-assay CV were 1.9 and 1.3%, respectively. Plasma NEFA and BHB concentrations were determined as described in Ollier et al. (2014) using NEFA-HR(2) reagents (Wako Diagnostics, Richmond, VA) and a BHB reagent set (Pointe Scientific Inc., Canton, MI), respectively. The intraand inter-assay CV were 3.8 and 4.0% for NEFA and 2.3% and 2.5% for BHB, respectively. Plasma lactose concentration was determined using a lactose assay kit (Sigma-Aldrich, Oakville, ON, Canada) according to the manufacturer's instructions. The intra-and interassay CV were 2.4 and 12.3%, respectively.
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