Ammonia production and glutamine consumption levels were determined using Ammonia Assay Kit (Abcam) and Glutamine/Glutamate-Glo assay (Promega), respectively. Intracellular ROS levels were quantified using CM-H2DCFDA (Invitrogen). Intracellular ATP concentrations were measured by ATP bioluminescent somatic cell assay kit (Sigma-Aldrich). Intracellular α-KG levels were determined using alpha-KG Assay Kit (Abcam). Briefly, 2 × 106 cells were collected for each group and cell volume was estimated by comparing the size of the cell pellet with the size of the known volume of PBS. Cell pellets were homogenized and centrifuged. The supernatant was further deproteinized using Amicon 10k centrifugal filter (Millipore) and applied to alpha-KG measurement.
Alpha kg assay kit
Manufactured by Abcam
The Alpha-KG Assay Kit is a quantitative colorimetric assay designed to measure alpha-ketoglutarate (α-KG) levels in a variety of sample types, including cell lysates, tissue homogenates, and biological fluids. The kit utilizes an enzymatic reaction that produces a colored product, the intensity of which is proportional to the α-KG concentration in the sample.
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Lab products found in correlation
3 protocols using alpha kg assay kit
1
Metabolic Profiling of Cell Lines
2
Quantifying α-Ketoglutarate in Hypoxic Cells
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WT and BBS Mut cells of MCF7 and HCC1806 cells subjected to 24 h of hypoxia were processed for α-ketoglutarate estimation. Briefly, equal number of cells were lysed using assay buffer provided in the alpha KG assay kit (Abcam, ab83431, lot no. GR3410863-1). The lysates were then deproteinized using perchloric acid and neutralized using 2 M KOH as per manufacturer's instructions. Finally, an equal amount of sample was taken, and the reaction was incubated at 37°C for 30 min. The absorbance was measured using a microplate reader set at an optical density of 450 nm.
3
Metabolic Profiling of Cell Lines
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Ammonia production and glutamine consumption levels were determined using Ammonia Assay Kit (Abcam) and Glutamine/Glutamate-Glo assay (Promega), respectively. Intracellular ROS levels were quantified using CM-H2DCFDA (Invitrogen). Intracellular ATP concentrations were measured by ATP bioluminescent somatic cell assay kit (Sigma-Aldrich). Intracellular α-KG levels were determined using alpha-KG Assay Kit (Abcam). Briefly, 2 × 106 cells were collected for each group and cell volume was estimated by comparing the size of the cell pellet with the size of the known volume of PBS. Cell pellets were homogenized and centrifuged. The supernatant was further deproteinized using Amicon 10k centrifugal filter (Millipore) and applied to alpha-KG measurement.
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