Enzymatic assay

Manufactured by Merck Group

Sourced in United States

An enzymatic assay is a quantitative analytical procedure used to measure the activity or concentration of a specific enzyme in a sample. It provides a means to determine the amount of a particular enzyme present in a biological sample, such as tissue, cells, or fluids. The core function of an enzymatic assay is to detect and quantify the enzymatic activity in a given sample.

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Lab products found in correlation

Oxiselect tbars assay kit, by Cell Biolabs (1 mentions) Cellytic m cell lysis reagent, by Merck Group (1 mentions) Enzymatic kit, by Merck Group (1 mentions) Atplite 1step, by PerkinElmer (1 mentions)

Close spelling variants of this product

Enzymatic Assay of Cathepsin B Colorimetric enzymatic assay Enzymatic Assay of Pyruvate Kinase kit Enzymatic assay kit

8 protocols using enzymatic assay

Hepatic Index and Histological Evaluation

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After sacrifice, liver tissues were removed from the animals and weighed. The
hepatic index was obtained by dividing the wet liver weight by the rabbit weight
and multiplying by 100. Macroscopic and microscopic analyses of specimens were
performed in a blinded fashion. Liver tissues were fixed in 10% neutral buffered
formalin and then embedded in paraffin. After deparaffinization and dehydration,
5 µm thick sections were stained with hematoxylin and eosin (H&E) using
standard protocols. The tissues were histologically graded according to the
NAFLD activity score system by two pathologists blinded to the treatment groups
(14 (link)). Hepatic triglycerides were
extracted from frozen tissue and measured by enzymatic assays (Sigma, USA). TG
values were normalized to protein concentration.

Li S., Wang X., Zhang J., Li J., Liu X., Ma Y., Han C., Zhang L, & Zheng L. (2018). Exenatide ameliorates hepatic steatosis and attenuates fat mass and FTO gene expression through PI3K signaling pathway in nonalcoholic fatty liver disease. Brazilian Journal of Medical and Biological Research, 51(8), e7299.

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Triglyceride and Free Fatty Acid Quantification

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Triglyceride (TG) contents were measured with enzymatic assays (Sigma). Free fatty acids (FFA) concentrations in cell extracts were measured with a commercial kit from Wako (Wako Pure Chemical Industries, Osaka, Japan). All values of TG and FFA in cell extracts were normalized to the protein concentration.

Yang S.J., Choi J.M., Chang E., Park S.W, & Park C.Y. (2014). Sirt1 and Sirt6 Mediate Beneficial Effects of Rosiglitazone on Hepatic Lipid Accumulation. PLoS ONE, 9(8), e105456.

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Quantifying Liver Biomarkers and Oxidative Stress

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Serum glucose, alanine aminotransferase (ALT), and aspartate aminotransferase (AST) were analyzed by enzymatic enzyme methods (Sigma-Aldrich, St. Louis, MO, USA). For hepatic triglyceride (TG) quantification, lipid was extracted as described in a previous study [28 (link)]. Briefly, liver tissue was homogenized in 1 mL solution containing 5% Nonidet P-40 (NP-40) substitute (Amresco, Solon, OH, USA). The homogenates were slowly heated to 80–100°C in a water bath for 2–5 min until the NP-40 became cloudy, then cooled down to room temperature. Samples were then centrifuged for 2 min to remove any insoluble materials. Hepatic TG levels were measured by enzymatic assay (Sigma-Aldrich) and normalized to their respective protein concentrations. Lipid peroxidation (MDA) levels in the liver tissue were measured by a OxiSelect^TM TBARS assay kit (Cell Biolabs, San Diego, CA, USA) according to the manufacturer’s protocol, normalized to their respective protein concentrations, and expressed as fold change compared to LETO control group.

Chang E., Kim D.H., Yang H., Lee D.H., Bae S.H, & Park C.Y. (2018). CB1 receptor blockade ameliorates hepatic fat infiltration and inflammation and increases Nrf2-AMPK pathway in a rat model of severely uncontrolled diabetes. PLoS ONE, 13(10), e0206152.

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Citrate synthase activity assay

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Citrate synthase activity was determined by commercially available enzymatic assay (Sigma). Briefly, cells were washed twice in ice-cold PBS and lysed (CelLytic M Cell Lysis Reagent; Sigma) in the presence of a protease inhibitor cocktail. Lysed cells were centrifuged at 20,000g for 15 min, and the supernatant was transferred to a fresh tube for determination of total protein content. Protein lysate, 8 μg, was combined with 1× assay buffer, 30 mM acetyl CoA solution, and 10 mM DTNB and plated in triplicate in a 96-well plate. Basal absorbance was measured on a plate reader set on a kinetic program (1.5 min duration, 10 s interval) at 412 nm. 10 mM oxaloacetate was then added to each well, the plate was briefly shaken, and the absorbance was read again at 412 nm on the same kinetic program.

King W.T., Axelrod C.L., Zunica E.R., Noland R.C., Davuluri G., Fujioka H., Tandler B., Pergola K., Hermann G.E., Rogers R.C., López-Domènech S., Dantas W.S., Stadler K., Hoppel C.L, & Kirwan J.P. (2021). Dynamin-related protein 1 regulates substrate oxidation in skeletal muscle by stabilizing cellular and mitochondrial calcium dynamics. The Journal of Biological Chemistry, 297(4), 101196.

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Serum Lactate Measurement in ARSB Mice

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Serum lactate was measured in the ARSB-deficient and control mice under non-stress conditions. L(+)-Lactate concentration was determined by an enzymatic assay (Sigma-Aldrich) in which a lactase enzyme produced a colorimetric (570 nm) product that was proportional to the lactate present in the serum samples.

Bhattacharyya S., Feferman L, & Tobacman J.K. (2016). Restriction of Aerobic Metabolism by Acquired or Innate Arylsulfatase B Deficiency: A New Approach to the Warburg Effect. Scientific Reports, 6, 32885.

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Enzymatic Plasma Amylase Assay

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Plasma amylase levels were determined at 37°C by means of an enzymatic assay (Sigma, St. Louis, MO, USA) with a spectrophotometer according to the manufacturer's instructions. All plasma samples were assayed in duplicate, and the results were averaged at the end of each experiment.

Zhao J., Zhong C., He Z., Chen G, & Tang W. (2015). Effect of Da-Cheng-Qi Decoction on Pancreatitis-Associated Intestinal Dysmotility in Patients and in Rat Models. Evidence-based Complementary and Alternative Medicine : eCAM, 2015, 895717.

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Measuring Cardiac LDH Activity

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Aliquots of the perfusate were collected at 1-min intervals, and LDH activity was measured by an enzymatic kit (Sigma-Aldrich) sensitive to detect changes in the µU/mL range, and expressed as U/L. Similar determinations were carried out in isolated cardiac myocytes and H9C2 cells. In both cell models, LDH activity was assessed as a percentage of the total amount of the enzyme released from digitonin-permeabilized cells.
Protein content was determined by Lowry assay (Lowry, Rosebrough, Farr, & Randall, 1951 (link)), using bovine serum albumin as a standard.
After excision of the heart, blood samples were collected by heparinized capillaries. The tubes were centrifuged at 500 × g for 20 min, and plasma alcohol content was determined by enzymatic assay (Sigma-Aldrich). Exposure to Lieber-DeCarli diet for 3 weeks resulted in a plasma ethanol concentration of 42 ± 3 mmol/L (n = 8), which is consistent with other published reports (Fisher et al., 2001).

, & Romani A.M. (2015). Effect of Acute and Prolonged Alcohol Administration on Mg2+ Homeostasis in Cardiac Cells. Alcohol (Fayetteville, N.Y.), 49(3), 265-273.

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Cellular ATP and Lactate Quantification

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Cellular ATP changes were determined by a luminescence assay (ATPLite-1step, PerkinElmer). At the end of the experiments, media were collected and centrifuged to eliminate cell debris. Lactate in the media was determined spectrophotometrically by an enzymatic assay (Sigma-Aldrich) according to manufacturer's instructions. Lactate concentration was calculated as μmol/mg protein and expressed as percent vs control condition.

Fumarola C., Cretella D., La Monica S., Bonelli M.A., Alfieri R., Caffarra C., Quaini F., Madeddu D., Falco A., Cavazzoni A., Digiacomo G., Mazzaschi G., Vivo V., Barocelli E., Tiseo M., Petronini P.G, & Ardizzoni A. (2017). Enhancement of the anti-tumor activity of FGFR1 inhibition in squamous cell lung cancer by targeting downstream signaling involved in glucose metabolism. Oncotarget, 8(54), 91841-91859.

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