Pyruvate, which is produced in the final step of glycolysis, participates in anabolic metabolism, catabolic metabolism, and signal transduction. Invitrogen Molecular Probes Amplex UltraRed Reagent 5 x 1mg Electrophoresis, Western Blotting and ELISA:Western Blotting Reagents, Membranes. Invitrogen Molecular Probes Amplex Red/UltraRed Stop Reagent 1 set Life Sciences:Cell Analysis:Cell Based Assays:Cell Growth and Differentiation Reagents.

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Free Radic Biol Med. We next evaluated the selectivity of the method by performing the same assay with phosphoenolpyruvate PEPlactate, acetoacetate, ketoglutarate, 2-oxobutyrate, oxaloacetate as samples. Fluorescence excitation and emission were performed at nm and nm, respectively, for experiments requiring high sensitivity.

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The background intensity was subtracted using the value of the no pyruvate control for all sample readings.

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Bakowski D, Parekh AB. Contact Us to ask a question, provide feedback, or report a problem. Properties of the Amplex UltraRed assay. Akplex made some changes to EPA.

Therefore, the enzymatic reactions are favorable in acidic buffer, while jltra fluorogenic reaction is more efficient in a neutral or weakly basic solution. The side reaction between H 2 O 2 and pyruvate can be effectively suppressed in acidic buffer pH 5.


Group members share an kltra in encouraging computational toxicology usage and exposure science in helping to implement EPA’s mission of protecting human health and the environment. Recently, we reported a sensitive enzymatic fluorimetric assay for the determination of intracellular pyruvate based on the stoichiometric oxidization of non-fluorescent Amplex Red acetyl-3, 7-dihydroxyphenoxazine by H 2 O 2 in the presence of horseradish peroxidase HRP to brightly fluorescent resorufin [ 3 ].

The importance of sodium pyruvate in assessing damage produced by hydrogen peroxide. For bacterial pyruvate oxidase, one unit will produce 1. To overcome this problem, we examined this fluorimetric assay with the new dye Amplex UltraRed Cat. The document below is a presentation from one of the monthly meetings of EPA’s Computational Toxicology Communities of Practice, a group composed of hundreds of people from over 50 public and private sector organizations.

The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form.

Amplex UltraRed enhances the sensitivity of fluorimetric pyruvate detection

The pH optimum of HRP is in the range of 6. Support Center Support Center. Open in ajplex separate window. Pyruvate is produced by pyruvate kinase EC2. We also tested the stability of Amplex UltraRed. Pyruvate, which is produced in the final step of glycolysis, participates in anabolic metabolism, catabolic metabolism, and signal transduction.


The excitation wavelength is nm.

Lamprecht W, Heinz F. Academic Press; New York: Error bars are not shown because their sizes are smaller than the symbols.

Amplex UltraRed enhances the sensitivity of fluorimetric pyruvate detection

Concentrations of metabolites in animal tissues. A simple statistical parameter for use in evaluation and validation of high throughput screening assays.

The linearity of these data are excellent, as judged by the coefficients of determination of 0. For these reasons, the reactions should be performed at pH 7—8. Each point is the mean of triplicate measurements. However, the absorption and fluorescence of resorufin are pH-dependent. Springer-Verlag; Berlin; New York: Please review our privacy policy.

Analysis of High-throughput screening data from Amplex UltraRed (AUR) Thyroperoxidase (TPO) assays

Please note that all information on the EPA website may not be up to date, and transactions and inquiries submitted to the EPA website may not be processed or responded to until appropriations are enacted. Ultrra sensitive fluorimetric assay for pyruvate. Therefore, we recommend fresh preparations of Amplex UltraRed aqueous solutions prior to assays.