Bioconjugation of Meldrum's acid activated furan: A detergent compatible assay for protein quantitation

A simple yet efficient assay for the quantitation of proteins ranging from plasma proteins to purified proteins from whole cell lysate, based on the bioconjugation reaction between protein and Meldrum's acid Activated Furan (MAF) is described. This easy to use, sensitive method is based on the...

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Bibliographic Details
Published in:Analytical biochemistry 2023-02, Vol.662, p.114998-114998, Article 114998
Main Authors: Ajayan, Kalyani, S, Sainath, Sadik, Ajmal, Nair, Manu Mohanan, Nair, Anju M., S, Karthika K., Vijayakumar, Anagha, Nair, Sudarslal Sadasivan, Nair, Bipin, Chandran R, Prakash, Nair, Sobha Vijayan
Format: Article
Language:English
Subjects:
BSA
Colorimetric assay
DASA
Detergent compatibility
Detergents
Dioxanes - pharmacology
Meldrum's acid activated furan
Protein quantitation
Proteins
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Summary:A simple yet efficient assay for the quantitation of proteins ranging from plasma proteins to purified proteins from whole cell lysate, based on the bioconjugation reaction between protein and Meldrum's acid Activated Furan (MAF) is described. This easy to use, sensitive method is based on the conjugation of amine functionalities present on the protein with MAF to form the corresponding Donor Acceptor Stenhouse Adducts (DASAs) with characteristic absorption in the visible region. The reaction is rapid as well as reproducible and shows a proportionate increase in color change over a broad range of protein concentration. The assay was found to be sensitive up to 0.125 mg/mL concentration of the protein and was compatible with most of the commonly employed detergents and isolation protocols which makes it ideal for the estimation of protein samples containing detergents. Another striking feature of this protocol is its tolerance towards other major interference contributors such as chelating agents, reducing agents, carbohydrates and protease inhibitors. [Display omitted] •A simple yet efficient protein quantitation assay using Meldrum's acid activated furan [MAF] reagent was developed.•The assay exploits the bio conjugate chemistry between the proteins and MAF, which facilitates their quantitation by colorimetric methods.•MAF assay render the accuracy levels similar to the other existing techniques, markedly with better tolerance towards detergents and reducing agents.•Quantitation of the protein content from various sources employing the MAF assay afforded results comparable to the existing protocols, thereby confirming its efficacy.
ISSN:0003-2697
1096-0309
DOI:10.1016/j.ab.2022.114998