α ketoglutarate assay kit

Manufactured by Merck Group

Sourced in United States

The α-Ketoglutarate Assay Kit is a laboratory equipment product that measures the concentration of α-ketoglutarate in a sample. It provides a colorimetric detection method for quantifying α-ketoglutarate levels.

Automatically generated - may contain errors

Lab products found in correlation

Close spelling variants of this product

α-ketoglutarate (34 citations) α-Ketoglutarate Dehydrogenase Activity Colorimetric Assay Kit (3 citations)

17 protocols using α ketoglutarate assay kit

Mitochondrial Respiratory Complex Analysis

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Mitochondria were isolated from NRCMs using the Mitochondria Isolation Kit for Cultured Cells (ab110170, Abcam). The activities of complexes I and III in the mitochondrial ETC were assessed with the Complex I Enzyme Activity Assay Kit (ab109721, Abcam) and the Mitochondrial Complex III Activity Assay Kit (K520-100, Biovision) according to the manufacturer's instructions, respectively [44 (link)].
The contents of α-KG and Suc were measured using the α-Ketoglutarate Assay Kit (MAK054, Sigma) and the Succinate Colorimetric Assay Kit (MAK184, Sigma) in accordance with the manufacturer's instructions, respectively [44 (link)].

He H., Wang L., Qiao Y., Yang B., Yin D, & He M. (2021). Epigallocatechin-3-gallate pretreatment alleviates doxorubicin-induced ferroptosis and cardiotoxicity by upregulating AMPKα2 and activating adaptive autophagy. Redox Biology, 48, 102185.

+ Open protocol

+ Expand

Quantifying Glutamate and α-KG Levels

Check if the same lab product or an alternative is used in the 5 most similar protocols

The concentration of glutamate and α-KG were measured by Glutamate Assay Kit (Abcam, ab83389) and α-Ketoglutarate Assay Kit (Sigma, MAK054), respectively. All the experiments were conducted following the manufacturers’ instructions. The intracellular glutamate and α-KG abundance was normalized to the cell number.

Zhou Q., Zhan H., Lin F., Liu Y., Yang K., Gao Q., Ding M., Liu Y., Huang W, & Cai Z. (2019). LincRNA-p21 suppresses glutamine catabolism and bladder cancer cell growth through inhibiting glutaminase expression. Bioscience Reports, 39(4), BSR20182372.

+ Open protocol

+ Expand

Quantification of α-Ketoglutarate Levels

Check if the same lab product or an alternative is used in the 5 most similar protocols

α-KG was quantified using the α-Ketoglutarate Assay Kit (Sigma-Aldrich), according to the manufacturer’s instructions. In all, 2×10⁶ cells were homogenised in α-KG buffer, the samples were mixed with a coupled enzyme, and the resulting fluorometric product measured using a FLUOstar Optima plate reader (BMG Labtech, Mornington, VIC, Australia) at Ex/Em=544/590 nm. The amount of α-KG per sample in nmol (Ay) was determined from the α-KG standard curve using the equation: Ay=(corrected absorbance−(y−intercept)/slope). All experimental samples were run as replicates and all samples and standards measured in duplicate. Results are expressed as fold change in α-KG levels compared to untreated cells of each line.

Lee W.T., Sun X., Tsai T.S., Johnson J.L., Gould J.A., Garama D.J., Gough D.J., McKenzie M., Trounce I.A, & St. John J.C. (2017). Mitochondrial DNA haplotypes induce differential patterns of DNA methylation that result in differential chromosomal gene expression patterns. Cell Death Discovery, 3, 17062-.

+ Open protocol

+ Expand

Colorimetric Assay for α-Ketoglutarate

Check if the same lab product or an alternative is used in the 5 most similar protocols

The assay was performed using the α-Ketoglutarate Assay Kit according to the manufacturer’s instructions (MAK054, Sigma Aldrich, U.S.A.). In brief, intracellular α-ketoglutarate levels were determined by a coupled enzyme assay, which results in a colorimetric (570 nm) product, proportional to the levels of α-Ketoglutarate present.

Zhao J., Jiang Y., Sun X., Liu X., Liu F., Song M, & Zhang L. (2020). The mechanism and role of intracellular α-ketoglutarate reduction in hepatic stellate cell activation. Bioscience Reports, 40(3), BSR20193385.

+ Open protocol

+ Expand

Quantifying Glutamate and α-Ketoglutarate Levels

Check if the same lab product or an alternative is used in the 5 most similar protocols

Glutamate and α-ketoglutarate concentrations were measured using the Glutamate Assay Kit (Sigma-Aldrich MAK004) and α-Ketoglutarate Assay Kit (Sigma-Aldrich MAK054) according to the manufacturer’s instructions. Briefly, dissected thoraces from 15 flies (per sample) were homogenized in 100 μL of ice-cold Glutamate Assay Buffer or α-KG Assay Buffer. The samples were centrifuged at 13,000g for 10 min to remove insoluble material. The supernatant was deproteinized with 10 kDa MWCO spin filter (Life Technologies) to remove enzymes. Samples were brought to 50 μL per well with Glutamate Assay Buffer or α-KG Assay Buffer. Reaction mix was added to each well and incubated for 30 min at 37°C. Absorbance (450 nm [A450] for Glutamate Assay and 570 nm [A570] for α-Ketoglutarate Assay) was obtained using an Epoch Microplate Spectrophotometer (BioTek Instruments). Glutamate and α-Ketoglutarate levels were normalized to total protein levels using the BCA Protein Assay Kit (Pierce).

Zhao X, & Karpac J. (2021). Glutamate Metabolism Directs Energetic Trade-offs to Shape Host-Pathogen Susceptibility in Drosophila. Cell metabolism, 33(12), 2428-2444.e8.

+ Open protocol

+ Expand

Metabolite Quantification in Ly6G+ Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

Measurement of glucose, pyruvate, and α-ketoglutarate levels was performed using the glucose assay kit (Sigma, catalog no GAHK20), pyruvate assay kit (Sigma, catalog no MAK071), and α-ketoglutarate assay kit (Sigma, catalog no MAK054), respectively. Briefly, samples were prepared from freshly isolated Ly6G⁺ cells according to the assay kits’ instructions. Lysates were then incubated with reaction mix for a designated time and ready for the absorbance measuring.

Zhao T., Liu S., Hanna N.H., Jalal S., Ding X., Wan J., Yan C, & Du H. (2023). LAL deficiency induced myeloid-derived suppressor cells as targets and biomarkers for lung cancer. Journal for Immunotherapy of Cancer, 11(3), e006272.

+ Open protocol

+ Expand

Molecular Cloning and Western Blotting

Check if the same lab product or an alternative is used in the 5 most similar protocols

The kits used for molecular cloning were from TransGen (China) or Takara (Japan). Taq DNA polymerase and restriction enzymes were obtained from Fermentas (Canada) or Takara (Japan). The antibodies and chemicals used for western blotting were from Sangon Biotech (China). Pure ethylene gas standard was purchased from Heli Gas Co., Ltd (China). The α-ketoglutarate assay kit was ordered from Sigma-Aldrich (USA). Unless otherwise specified, all other chemical reagents were purchased from Sigma-Aldrich (USA).

Mo H., Xie X., Zhu T, & Lu X. (2017). Effects of global transcription factor NtcA on photosynthetic production of ethylene in recombinant Synechocystis sp. PCC 6803. Biotechnology for Biofuels, 10, 145.

+ Open protocol

+ Expand

α-Ketoglutarate Quantification in Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

Cells treated with TPA and/or WA were collected and resuspended with α-KG Buffer. The supernatant was deproteinized by passing through a 10 kD MWCO spin filter (Thermo Scientific Pierce). Two microliters of the whole-cell lysate filtrate was diluted to 50 µL with α-KG Buffer. Alpha-ketoglutarate concentrations were measured using α-Ketoglutarate Assay Kit (Sigma) according to the manufacturer’s instructions. For each sample, the concentration of α-KG was recorded by spectrophotometry at 570 nm, and calculated by the equation provided by the manufacturer (Gene Company Limited).

Xu K., Zhang C., Li Y., Xi X., Zheng L., Meng M., Liu T., Zhao Y, & Li W. (2019). Withaferin A suppresses skin tumor promotion by inhibiting proteasome-dependent isocitrate dehydrogenase 1 degradation. Translational Cancer Research, 8(6), 2449-2460.

+ Open protocol

+ Expand

Intracellular Metabolite Profiling of ADSCs

Check if the same lab product or an alternative is used in the 5 most similar protocols

ADSCs (2 × 10⁶ in quantity) transfected with shNC or shBcat1#1 were used for intracellular metabolite concentration measurement. Each metabolite level was determined using the following assay kits: BCAA detection kit (BioVision, K564‐100, α‐ketoglutarate assay kit (Sigma‐Aldrich, MAK054), glutamate assay kit (Sigma‐Aldrich, MAK004), citrate assay kit (Sigma‐Aldrich, MAK057), fumarate assay kit (Sigma‐Aldrich, MAK060), succinate assay kit (Sigma‐Aldrich, MAK184), and malate assay kit (Sigma‐Aldrich, MAK196). BCKA concentration was measured by high‐performance liquid chromatography (HPLC) as previously described.^[⁴²^] Intracellular metabolite levels were normalized to the ADSCs transfected with shNC.

Hu G., Cui Z., Chen X., Sun F., Li T., Li C., Zhang L., Guo X., Zhao H., Xia Y., Yan W., Yi W., Fan M., Yang R., Wang S., Tao L, & Zhang F. (2023). Suppressing Mesenchymal Stromal Cell Ferroptosis Via Targeting a Metabolism‐Epigenetics Axis Corrects their Poor Retention and Insufficient Healing Benefits in the Injured Liver Milieu. Advanced Science, 10(13), 2206439.

+ Open protocol

+ Expand

Metabolic Profiling in Cell Cultures

Check if the same lab product or an alternative is used in the 5 most similar protocols

Cellular lactate levels, α-KG levels, IDH1/IDH2 and IDH3 activities, and NADH/NAD⁺ ratios were measured using the Lactate Assay Kit (Sigma, MAK064), α-Ketoglutarate Assay Kit (Sigma, MAK054), Isocitrate Dehydrogenase Activity Assay Kit (Sigma, MAK062), and NAD/NADH Quantification Kit (Sigma, MAK037), respectively, according to the manufacturer's instructions. The measurements were performed and recorded by a Synergy 2 multidetection microplate reader (BioTek, USA).

La T., Chen S., Guo T., Zhao X.H., Teng L., Li D., Carnell M., Zhang Y.Y., Feng Y.C., Cole N., Brown A.C., Zhang D., Dong Q., Wang J.Y., Cao H., Liu T., Thorne R.F., Shao F.M., Zhang X.D, & Jin L. (2021). Visualization of endogenous p27 and Ki67 reveals the importance of a c-Myc-driven metabolic switch in promoting survival of quiescent cancer cells. Theranostics, 11(19), 9605-9622.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!