Extracellular Pyrophosphate Quantification

Extracellular PPi levels were measured using the differential adsorption of uridine diphospho-(6-³H) glucose (GE Healthcare, Buc, France), and its reaction product 6-phospho-(6-³H) gluconate on activated charcoal, as previously described [44 (link)]. The standard concentrations, ranging from 10 to 400 pmol of ePPi, were included in each assay. After adsorption of the reaction mixture on charcoal, and centrifugation at 14,000 rcf for 10 min, 100 μL of supernatant were removed carefully and counted for radioactivity in 5 mL of Bio-Safe II (Research Products International Corp., Mt. Prospect, IL, USA). Results were expressed as picomoles of ePPi per microgram of total cell proteins.

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Velot É., Elkhoury K., Kahn C., Kempf H., Linder M., Arab-Tehrany E, & Bianchi A. (2022). Efficient TGF-β1 Delivery to Articular Chondrocytes In Vitro Using Agro-Based Liposomes. International Journal of Molecular Sciences, 23(5), 2864.

Publication 2022

Bio-Safe II

Activated charcoal Adsorption Assay Cell Centrifugation Charcoal Gluconate Glucose Ml ii Proteins Radioactivity Uridine

Corresponding Organization :

Other organizations : Université de Lorraine, Centre National de la Recherche Scientifique, Ingénierie Moléculaire et Physiopathologie Articulaire, Laboratoire des Biomolécules

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Variable analysis

independent variables

Extracellular PPi levels

dependent variables

Radioactivity of the supernatant

control variables

Standard concentrations of ePPi, ranging from 10 to 400 pmol
Adsorption of the reaction mixture on charcoal
Centrifugation at 14,000 rcf for 10 min

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

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