Prussian Blue, a blue coordination polymer has a very promising future in the realm of biomedicine. Its nanoparticles, known as catalytic labels or nanozymes. They exhibit remarkable peroxidase-like properties and serve as effective antioxidants. Demand for synthesizing Prussian Blue nanoparticles with customizable sizes is on the rise. This article unveil a novel approach to synthesizing Prussian Blue nanoparticles. A synthesis of Prussian Blue nanoparticles by reducing an equimolar mixture of FeCl3 and K3[Fe(CN)6] with hydrogen peroxide in different water-alcohol mixtures is demonstrated for the first time. Alcohols with a lower dielectric constant (propanol-1, isopropyl alcohol, and tert-butanol) contribute to an increase in nanoparticle size, particularly at mole fractions of 0.02–0.05 and beyond. Conversely, alcohols with a higher dielectric constant (ethanol, methanol, ethylene glycol, and propylene glycol, excluding glycerol) demonstrate the ability to decrease nanoparticle size at mole fractions of 0.2–0.26 and higher. A scalable and reproducible method for preparing 30–40 nm Prussian Blue nanoparticles using 79.2% ethylene glycol as a solvent is presented. The proposed mechanism behind the effect of ethylene glycol involves the limitation of both growth and secondary aggregation of Prussian Blue nanoparticles. These synthesized nanoparticles prove their efficiency as catalytic labels in a model vertical flow immunoassay designed to detect antibodies against SARS-CoV-2.

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