The level of glucose uptake was determined by glucose uptake colorimetric assay kit (BioVision, Milpitas, CA, USA). Briefly, cells with a density of 1 × 104 cells were seeded, and subsequently cultured with 100 mL Krebs-Ringer-Phosphate-HEPES (KRPH) buffer containing 2% albumin from bovine serum, followed by adding 10 mM 2-deoxyglucose (2-DG). The samples were collected for the analysis of the glucose uptake level. For the determination of lactate production, cells were seeded into the 96-well plate and starved using non-serum medium for 24 h. Then, the cells were suspended in the buffer from lactate assay kit II (BioVision). The glucose uptake colorimetric assay kit and lactate assay kit II (BioVision) were also used for the determination of glucose uptake and lactate production in tumor tissues from the nude mice according to the manufacturer’s instructions.
Lactate assay kit 2
Manufactured by Abcam
Sourced in United States
The Lactate Assay Kit II is a colorimetric assay designed to quantify lactate levels in a variety of sample types. The kit uses an enzyme-based reaction to produce a colored product, which is measured spectrophotometrically. The assay is simple, sensitive, and can be used to determine lactate concentrations in biological samples.
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Lab products found in correlation
Glucose uptake colorimetric assay kit, by Abcam (19 mentions)
Atp colorimetric assay kit, by Abcam (18 mentions)
Glucose assay kit 2, by Abcam (3 mentions)
Trypsin edta, by Thermo Fisher Scientific (3 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions)
Microplate reader, by Molecular Devices (2 mentions)
Pyruvate colorimetric assay kit, by Abcam (2 mentions)
Glucose assay kit, by Eton Bioscience (2 mentions)
Hexokinase colorimetric assay kit, by Abcam (1 mentions)
Microplate reader, by Agilent Technologies (1 mentions)
Atp calorimetric assay kit, by Abcam (1 mentions)
Microplate reader, by Bio-Rad (1 mentions)
Atp assay kit, by Beyotime (1 mentions)
Cell recovery solution, by BD (1 mentions)
Matrigel, by BD (1 mentions)
Passive lysis buffer (plb), by Promega (1 mentions)
Bradford assay kit, by Bio-Rad (1 mentions)
K686 100, by Abcam (1 mentions)
Pyruvate colorimetric fluorometric assay kit, by Abcam (1 mentions)
Assay buffer, by Abcam (1 mentions)
37 protocols using lactate assay kit 2
1
Glucose Uptake and Lactate Production Assay
2
Metabolic Profile Analysis
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The levels of ATP and lactate and the consumption of glucose were analyzed using ATP Colorimetric Assay kit (Biovision, Milpitas, California, USA), Lactate Assay Kit II (Biovision), and Glucose Uptake Colorimetric Assay kit (Biovision).
3
Colorimetric Assays for Metabolic Analyses
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Hexokinase Colorimetric Assay Kit, Glucose Uptake Colorimetric Assay kit, ATP Colorimetric Assay kit and Lactate Assay Kit II were purchased from Biovision and used to detect HK activity, glucose uptake, ATP, and lactate production, respectively. These assays were detected following the manufacturer’s protocols as described previously [47 (link)].
4
Adipose Tissue Metabolic Markers
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Lactate in eWAT was determined using the Lactate Assay Kit II (Biovision, Mountain View, CA, USA) according to the manufacturer’s protocol. Mitochondrial density markers citrate synthase and aconitase activity were determined as published [24 (link)]. Paraffin embedded eWAT was sliced at 5 µm and stained with a MAC-2 antibody recognizing macrophages for detection of CLS as published [29 (link)], and using MAC-2 fluorescence staining for individual macrophage counting. HE-stained WAT sections were used for determination of adipocyte size distribution as published [24 (link)].
5
Measuring Cellular Metabolism Markers
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Glucose uptake and lactate production were measured with Glucose Assay Kit II and Lactate Assay Kit II (Biovision) according to manufacturer's instructions. Aliquots of the medium were removed from cell culture and Glucose uptake and lactate production was determined by the concentration difference between samples and blank medium control. Cellular ATP and NADH contents were measured using the ATP Colorimetric Assay Kit and NADH Quantitation Kit (Biovision) respectively as described before [9 (link)].
6
Metabolic Profiling of Tanshinone IIA
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Glucose uptake and lactate production were assessed using a Glucose Uptake Colorimetric Assay Kit (catalog no. ab136955; Abcam) and a Lactate Assay Kit II (catalog no. K267-100; BioVision, Inc.), respectively, following the manufacturers' instructions. Briefly, 5×105 cells were seeded into 96-well plates and treated with Tan IIA (5 µM) for 48 h. After washing with PBS, the cells were preincubated with 100 ml Krebs-Ringer-Phosphate-HEPES buffer containing 2% BSA for 40 min at room temperature. Next, 2-deoxy-D-glucose was added, and the cells were incubated for 20 min at 37°C. Afterward, cells were lysed with 90 ml extraction buffer and then 10 ml neutralization buffer was added. After centrifugation (1,500 × g, for 10 min at room temperature), the supernatant was used to measure glucose uptake at 412 nm and lactate production at 450 nm in a microplate reader (BioTek Instruments, Inc.).
For ATP level analysis, cells were lysed in the same manner as for the glucose uptake and lactate production assays. The cells were collected and extracted in 100 ml of the ATP Assay Buffer from the ATP calorimetric assay kit (BioVision, Inc.). After centrifugation (1,500 × g, for 10 min at room temperature), the supernatants were incubated for 30 min at room temperature, and then absorbance at 570 nm was measured using a microplate reader (BioTek Instruments, Inc.).
For ATP level analysis, cells were lysed in the same manner as for the glucose uptake and lactate production assays. The cells were collected and extracted in 100 ml of the ATP Assay Buffer from the ATP calorimetric assay kit (BioVision, Inc.). After centrifugation (1,500 × g, for 10 min at room temperature), the supernatants were incubated for 30 min at room temperature, and then absorbance at 570 nm was measured using a microplate reader (BioTek Instruments, Inc.).
7
Lactate Production Quantification
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Lactate Assay Kit II was used to assess the lactate according to the manufacturer's protocols (BioVision). Cells were plated into 96-well plates at 1000 cells per well and then grown in DMEM containing 10% FBS for 10 h. The media was removed, and the cells were incubated in DMEM without FBS for 1 h. The supernatant was collected for assessment of lactate production.
8
Lactate and Glucose Quantification
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Lactate and glucose in culture medium were measured with the respective lactate assay kit II and glucose assay kit II according to the manufacturer's instructions (BioVision, Mountain View, CA). Briefly, after centrifugation (3,500 rpm, 15 min, 4 °C), cell culture medium supernatants were frozen at −20 °C. Samples were later thawed, diluted in assay buffer, and mixed with lactate or glucose reaction mixture for 30 min. The optical density of the mixture in each well was read at 450 nm on a microplate reader (Molecular Devices). The lactate concentration was calculated from a standard curve and normalized to cell numbers and culture time. Glucose consumption was calculated from a standard curve, subtracting background from cell-free medium, and normalizing to cell numbers and time.
9
Metabolic Profiling of Transfected Cells
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The levels of lactate production, glucose uptake, and ATP production were detected by Lactate Assay Kit II, Glucose Uptake Colorimetric Assay Kit, and ATP Colorimetric Assay Kit (BioVision, Milpitas, CA, USA) according to the manufacturer’s instructions, respectively. For the examination of glucose uptake, the transfected cells with the density of 1 × 104 cells per well were seeded into the 96-well plate, followed by culturing in 100 mL Krebs-Ringer-Phosphate-HEPES (KRPH) buffer supplemented with 2% albumin from bovine serum for 40 min. Then, 10 mM 2-deoxyglucose (2-DG) was added and the cells were cultured for 20 min. For the measurement of lactate and ATP production, 1 × 104 cells were suspended in the buffer from the corresponding assay kit. Then, the samples were centrifuged at 4°C for analyzing the soluble fraction.
10
Quantification of Lactate and ATP in Cells
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For lactate level assay, 1 × 105 cells were seeded into a 12-well plate and grew overnight. The medium was replaced by FBS-free DMEM/F12 followed by incubation for 2 h. Then, the supernatant was collected for specific reactions according to the protocol of Lactate Assay Kit II (Biovision, USA). The reaction mixture was protected from light at room temperature for 30 min. Lactate level was measured at 450 nm by a microplate reader (Bio-Rad, USA). Additionally, we also detected the lactate level within tumor tissue.
ATP Colorimetric Assay Kit (Biovision, USA) was used to analyze ATP production. According to the manufacturer’s protocol, a total of 5 × 105 cells were collected and extracted with ATP Assay Buffer. After centrifugation at 1.2 × 104 rpm for 5 min, the supernatants were collected for further reaction for 30 min in the dark at room temperature. Optical density (OD) values of reaction mixture were measured at 570 nm using a microplate reader. All above measurements were normalized to protein or tumor tissue weight.
ATP Colorimetric Assay Kit (Biovision, USA) was used to analyze ATP production. According to the manufacturer’s protocol, a total of 5 × 105 cells were collected and extracted with ATP Assay Buffer. After centrifugation at 1.2 × 104 rpm for 5 min, the supernatants were collected for further reaction for 30 min in the dark at room temperature. Optical density (OD) values of reaction mixture were measured at 570 nm using a microplate reader. All above measurements were normalized to protein or tumor tissue weight.
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