BioNukleo offers carefully assembled kits for the detection of (pyro-) phosphate by continuous UV-spectroscopic monitoring of 5-bromouridine phosphorolysis (PUB module, see also Eilert et
al. 2022, DOI: 10.1021/acs.analchem.1c05356). The PUB module allows quantitative phosphate detection by observing the shift in UV spectrum when
5-bromouridine and phosphate are converted to 5-bromouracil by a pyrimidine phoshphorylase. Key properties of the PUB phosphate detection system are:
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Easy to use
Very few simple sample handling steps and detection by standard spectroscopic equipment.
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Sensitive
Phosphate concentrations of 100 µM to >5 mM can readily be detected. Lower phosphate concentrations down to around 2 µM can be detected by providing 5-bromouridine in excess and
measuring absorbance differences between a baseline and a near-equilibrium state.
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Selective
Highly selective to inorganic phosphate and exhibits no cross-reactivity with other phosphate species.
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Scalable
Compatible with different reaction volumes, reaction vessels and sizes of analysis equipment. Detection can be carried out in 1 ml
cuvettes in a photometer, 200 µl multiwell plates in a platereader or even in 1 µl droplets assessed via nanodrop.
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Versatile
Functioning at a broad range of experimental conditions and applicable to a variety of experimental scenarios and reactions of interest.
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HT-suited
Compatible with parallelized experimentation, for instance in assays in multiwell plates.
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Quick
No long sample incubation times and readout in under 30 mins.
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Continuous
Enables continuous monitoring of live reactions.
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Broad temperature range
Not limited by single assay temperature requirements.
The extended version of this kit - including an inorganic pyrophosphatase (hydrolyzing pyrophosphate to orthophosphate) - enables the quantification of pyrophosphate in a fully
analogous fashion.
Available upon request. Please contact us for further information.
