Pdh activity kit

Manufactured by Abcam

The PDH activity kit is a tool used to measure the activity of pyruvate dehydrogenase (PDH), a key enzyme complex involved in cellular energy metabolism. The kit provides the necessary reagents and protocols to quantify PDH activity in various biological samples.

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Lab products found in correlation

Anti actin monoclonal antibody, by Merck Group (3 mentions) A cocktail of protease inhibitors complete cocktail, by Roche (3 mentions) Immun star hrp substrate kit, by Bio-Rad (3 mentions) Trypsin, by Merck Group (3 mentions) Sodium pantothenate, by Merck Group (2 mentions) Aconitase activity kit, by Abcam (2 mentions) Nfs1 antibodies, by Santa Cruz Biotechnology (2 mentions) Anti tau clone ht7, by Thermo Fisher Scientific (2 mentions) Anti isd11, by Thermo Fisher Scientific (2 mentions) Anti pank1, by Thermo Fisher Scientific (2 mentions) Anti pank3, by Thermo Fisher Scientific (2 mentions) Hoechst 3342, by Thermo Fisher Scientific (2 mentions) Anti lipoic acid, by Merck Group (2 mentions) Pantethine, by Merck Group (1 mentions) Pantothenate, by Merck Group (1 mentions) Vitamin e, by Merck Group (1 mentions) Luperox, by Merck Group (1 mentions) Thiamine, by Merck Group (1 mentions) Prussian blue, by Merck Group (1 mentions) Bodipy 581 591 c11, by Thermo Fisher Scientific (1 mentions)

5 protocols using pdh activity kit

Antibody-based Mitochondrial Protein Analysis

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Monoclonal anti-actin antibody, Prussian Blue, pantothenate, pantethine, vitamin E, L-carnitine, thiamine, Luperox^® and trypsin were purchased from Sigma Chemical Co. (St. Louis, MO). Anti-mitochondrial acyl carrier protein (mtACP), anti-NF-Y, anti-FOXN4 and anti-hnRNPA/B were purchased from Invitrogen/Molecular Probes (Eugene, OR). Anti-phospho-PGC1α was purchased from RD systems. NFS1 antibody and Omega 3 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA). Anti-PANK2, anti-PGC1α, complex 1 activity kit and PDH activity kit were purchased from Abcam (Cambridge, UK). Anti-TFAM was purchased from Cell Signaling. BODIPY^® 581/591 C11 was purchased from Thermo-Fisher (Waltham, MA). A cocktail of protease inhibitors (complete cocktail) was purchased from Boehringer Mannheim (Indianapolis, IN). The Immun Star HRP substrate kit was from Bio-Rad Laboratories Inc. (Hercules, CA).

Álvarez-Córdoba M., Reche-López D., Cilleros-Holgado P., Talaverón-Rey M., Villalón-García I., Povea-Cabello S., Suárez-Rivero J.M., Suárez-Carrillo A., Munuera-Cabeza M., Piñero-Pérez R, & Sánchez-Alcázar J.A. (2022). Therapeutic approach with commercial supplements for pantothenate kinase-associated neurodegeneration with residual PANK2 expression levels. Orphanet Journal of Rare Diseases, 17, 311.

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Pyruvate Dehydrogenase Activity Assay

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PDH activity was assessed using a commercially available PDH activity kit from Abcam (Catalog No: ab109902). In the assay, the PDH enzyme was immunocaptured within the wells of the microplate and activity was determined by following the reduction of NAD+ to NADH, coupled to the reduction of a reporter dye to yield a colored reaction product with an increase in absorbance at 450 nm at room temperature. Hearts were homogenized in ice-cold PBS containing protease and phosphatase inhibitors. Protein concentration was determined using a BCA protein assay and each sample was adjusted to 200 μg of protein. The assay was then performed to the manufacturer’s specifications in kinetic mode for 15 minutes at room temperature. The rate (ΔmOD/min) was then calculated by using the following formula:

\frac{R a t e = (A b s o r b a n c e_{t 2} - A b s o r b a n c e_{t 1})}{t i m e (m i n u t e s)}

Shimada B.K., Boyman L., Huang W., Zhu J., Yang Y., Chen F., Kane M.A., Yadava N., Zou L., Lederer W.J., Polster B.M, & Chao W. (2021). Pyruvate-Driven Oxidative Phosphorylation is Downregulated in Sepsis-Induced Cardiomyopathy: A Study of Mitochondrial Proteome. Shock (Augusta, Ga.), 57(4), 553-564.

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Measuring Pyruvate Dehydrogenase Activity

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PDH activity was assessed using a commercially available PDH activity kit from Abcam (Catalog No: ab109902). In the assay, the PDH enzyme was immunocaptured within the wells of the microplate and activity was determined by following the reduction of NAD+ to NADH, coupled to the reduction of a reporter dye to yield a colored reaction product with an increase in absorbance at 450 nm at room temperature. Hearts were homogenized in ice-cold PBS containing protease and phosphatase inhibitors. Protein concentration was determined using a BCA protein assay and each sample was adjusted to 200 μg of protein. The assay was then performed to the manufacturer's specifications in kinetic mode for 15 min at room temperature. The rate (ΔmOD/min) was then calculated by using the following formula:

\frac{R a t e = (A b s o r b a n c e_{t 2} - A b s o r b a n c e l_{t 1})}{t i m e (\min u t e s)}

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Immunohistochemistry Reagents and Antibodies

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Monoclonal Anti-actin antibody, anti-fatty acid synthase (FAS), Prussian Blue, sodium pantothenate and trypsin were purchased from Sigma Chemical Co. (St. Louis, MO). Anti-mitochondrial acyl carrier protein (mtACP), anti-aminoadipate semialdehyde dehydrogenase phosphopantetheinyl transferase (AASDHPPT), anti-NADH dehydrogenase [ubiquinone] 1 alpha subcomplex subunit 9 (NDUFA9), anti-NADH-ubiquinone oxidoreductase chain 1 (MTND1), DAPI and Hoechst 3342, were purchased from Invitrogen/Molecular Probes (Eugene, OR). NFS1 antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA). Anti-PANK2, complex 1 activity kit, PDH activity kit and aconitase activity kit were purchased from Abcam (Cambridge, UK). Anti-mitochondrial 10-formyltetrahydrofolate dehydrogenase (ALDH1L2), anti-cytosolic 10-formyltetrahydrofolate dehydrogenase (ALDH1L1), anti-alpha-aminoadipic semialdehyde synthase, mitochondrial (AASS), anti-LYRM4, anti-Tau clone HT7, anti-ISD11, anti-PANK1 and anti-PANK3 were purchased from Thermo-Fisher (Waltham, MA). Anti-lipoic acid was acquired from Merck (Darmstadt, Germany). A cocktail of protease inhibitors (complete cocktail) was purchased from Boehringer Mannheim (Indianapolis, IN). The Immun Star HRP substrate kit was from Bio-Rad Laboratories Inc. (Hercules, CA).

Álvarez-Córdoba M., Talaverón-Rey M., Villalón-García I., Povea-Cabello S., Suárez-Rivero J.M., Suárez-Carrillo A., Munuera-Cabeza M., Salas J.J, & Sánchez-Alcázar J.A. (2021). Down regulation of the expression of mitochondrial phosphopantetheinyl-proteins in pantothenate kinase-associated neurodegeneration: pathophysiological consequences and therapeutic perspectives. Orphanet Journal of Rare Diseases, 16, 201.

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Monoclonal Antibody-Based Mitochondrial Profiling

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Monoclonal Anti-actin antibody, anti-fatty acid synthase (FAS), Prussian Blue, sodium pantothenate and trypsin were purchased from Sigma Chemical Co. (St. Louis, MO). Anti-mitochondrial acyl carrier protein (mtACP), anti-aminoadipate semialdehyde dehydrogenase phosphopantetheinyl transferase (AASDHPPT), anti-NADH dehydrogenase [ubiquinone] 1 alpha subcomplex subunit 9 (NDUFA9), anti-NADH-ubiquinone oxidoreductase chain 1 (MTND1), DAPI and Hoechst 3342, were purchased from Invitrogen/Molecular Probes (Eugene, OR). NFS1 antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA). Anti-PANK2, complex 1 activity kit, PDH activity kit and aconitase activity kit were purchased from Abcam (Cambridge, UK). Antimitochondrial 10-formyltetrahydrofolate dehydrogenase (ALDH1L2), anti-cytosolic 10-formyltetrahydrofolate dehydrogenase (ALDH1L1), anti-alpha-aminoadipic semialdehyde synthase, mitochondrial (AASS), anti-LYRM4, anti-Tau clone HT7, anti-ISD11, anti-PANK1 and anti-PANK3 were purchased from Thermo-Fisher (Waltham, MA). Anti-lipoic acid was acquired from Merck (Darmstadt, Germany). A cocktail of protease inhibitors (complete cocktail) was purchased from Boehringer Mannheim (Indianapolis, IN). The Immun Star HRP substrate kit was from Bio-Rad Laboratories Inc. (Hercules, CA).

Álvarez-Córdoba M., Talaverón-Rey M., Villalón-García I., Povea-Cabello S., Suárez-Rivero J.M., Suárez-Carrillo A., Munuera-Cabeza M., Salas J.J., & Sánchez‐Alcázar J.A. (2021). Down Regulation Of The Expression Of Mitochondrial Phosphopantetheinyl-Proteins In Pantothenate Kinase-Associated Neurodegeneration: Pathophysiological Consequences and Therapeutic Perspectives.

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