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Alizarin Red S for Online Pyrophosphate Detection Identified by a Rapid Screening Method.

Jens Fischbach¹, Qiuting Loh², Frank F Bier³

¹Division Molecular Biotechnology and Functional Genomics, Technical University of Applied Sciences Wildau, Hochschulring 1, 15745 Wildau, Germany.

Scientific Reports

|March 25, 2017

Summary

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Alizarin Red S is a novel fluorescent dye for detecting pyrophosphate in enzymatic assays like LAMP and PCR. This inexpensive, non-toxic dye offers superior performance for pyrophosphate detection in various biological applications.

Area of Science:

Biochemistry
Molecular Biology
Analytical Chemistry

Background:

Pyrophosphate detection is crucial for various enzymatic assays.
Existing methods for pyrophosphate detection can be limited in sensitivity or applicability.
There is a need for robust and accessible fluorescent dyes for real-time pyrophosphate monitoring.

Purpose of the Study:

To identify and evaluate fluorescent dyes for online pyrophosphate detection in enzymatic assays.
To assess the suitability of Alizarin Red S for loop-mediated isothermal amplification (LAMP) and polymerase chain reaction (PCR) assays.
To develop a screening method for discovering new pyrophosphate detection compounds.

Main Methods:

Iterative screening of selected fluorescent dyes for enzyme compatibility.

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Evaluation of inorganic pyrophosphate sensitivity using manganese as a quencher.

Optimization of detection conditions for online monitoring in LAMP and PCR assays.

Main Results:

Alizarin Red S was identified as a selective and effective fluorescent dye for pyrophosphate detection.
The dye demonstrated superior performance in LAMP and PCR assays compared to other tested compounds.
Alizarin Red S exhibits a red-shifted spectrum, long shelf life, and low toxicity.

Conclusions:

Alizarin Red S is a valuable tool for online pyrophosphate detection in nucleic acid amplification assays.
The dye's properties make it suitable for a broader range of enzymatic assays not limited to nucleic acid generation.
The developed screening method can facilitate the discovery of novel pyrophosphate detection agents.