Metal-enhanced bioluminescence (MEB) is a complex photophysical event which manifests itself in manifold luminescence enhancement and depends on many parameters. The parameters include the distance between the luminescent source and the nanomaterial surface, the size and shape of nanoparticles, localized surface plasmon resonance (LSPR) peaks of the nanomaterial and the dielectric constant of the surrounding medium. Studying distance-dependent MEB in the presence of linkers of specified length may provide new insights into luminescence enhancement. In this regard, the present investigation is aimed to understand the role of ethylene diamine as a potential linker in model systems for distance-dependent metal-enhanced bioluminescence (MEB) with gold nanoparticles (AuNPs). Four different types of AuNPs (AuNP1, AuNP2, AuNP3, AuNP4) were synthesized by varying the trisodium citrate (TC) and silver nitrate (AgNO3) concentrations with maximum absorbance values of 0.99, 1.24, 1.21 and 1.38 respectively and the corresponding LSPR peaks (λmax) of 520 nm, 535 nm, 525 nm, and 525 nm. Luminescence enhancement up to 1.44-fold was observed when ethylene diamine (ED) was used as a linker in the presence of 1-N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide (EDC), ATP and AuNP1. The sample consisting of FMN, EDC and AuNP1 showed 1.3-fold luminescence enhancement. It was noted that AuNPs synthesized using AgNO3 as an additional component did not enhance luminescence in all the investigations. The suggested technique of using linkers of predetermined lengths may also prove fairly effective for studying other parameters which can influence MEB and cause sensitivity enhancement of luminescence-based biosensors
The present study describes the method of preparing reagents containing firefly luciferase (FLuc) and its substrate, D-luciferin, immobilized into gelatin gel separately or together. The addition of stabilizers dithiothreitol (DTT) and bovine serum albumin (BSA) to the reagent is a factor in achieving higher activity of reagents and their stability during storage. The use of immobilized reagents substantially simplifies the procedure of assay for microbial contamination. The mechanism of action of the reagents is based on the relationship between the intensity of the bioluminescent signal and the level of ATP contained in the solution of the lysed bacterial cells. The highest sensitivity to ATP is achieved by using immobilized FLuc or reagents containing separately immobilized FLuc and D-luciferase. The limit of detection of ATP by the developed reagents is 0.3 pM, which corresponds to 20,000 cells·mL−1. The linear response range is between 0.3 pM and 3 nM ATP. The multicomponent reagent, containing co-immobilized FLuc and D-luciferin, shows insignificantly lower sensitivity to ATP—0.6 pM. Moreover, the proposed method of producing an immobilized firefly luciferin-luciferase system holds considerable promise for the development of bioluminescent biosensors intended for the analysis of microbial contamination.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.