BACKGROUND

Technical Field

The present disclosure is directed to fluorescence microscopy, and particularly, to a method of increasing a fluorescence microscopy signal of a fluorophore.

Description of Related Art

The “background” description provided herein is for the purpose of generally presenting the context of the disclosure. Work of the presently named inventors, to the extent it is described in this background section, as well as aspects of the description that may not otherwise qualify as prior art at the time of filing, are neither expressly nor impliedly admitted as prior art against the present invention.

Fluorescence-based microscopy is a technique used in a diverse range of fields of research from single-molecule detection to early cancer diagnosis, super-resolution display, and quantum photonics. The technique is well-known because of its excellent sensitivity and selectivity, in addition to low invasiveness. However, due to very weak emission, the use of fluorescence microscopy faces challenges, and methods are needed to enhance intrinsic emission of analytes.

To address such challenges, a strategy, known as a surface-enhanced fluorescence (SEF), had been introduced. SEF enhancement is attributed to mainly two factors; one, designing a plasmon-active tag as a driving part, and two, positioning a fluorophore in the right place to provide the enhanced fluorescence. Noble metal nanoparticles, particularly, silver nanoparticles or nanostructures, are well-known fluorescent tags, and their plasmonic properties have been found very promising in fluorescence spectroscopy. By tuning the optical properties and nanomorphology of the silver nanoparticles, the detection efficiency can be enhanced several orders of magnitude through SEF. When the fluorophore is placed in a suitable position near the plasmon-active fluorescent tag, local electromagnetic (EM) near-field distributions enhance the excitation rate as well as a radiative decay rate and thus enhance fluorescence quantum yield. Many previous studies, adjust the positioning of the fluorophores by introducing a spacer or by shaping the plasmon active tag, which leads to a complicated design and inconsistent results.

As such, there exists a need to design a facile method that will provide strong electromagnetic (EM) near-field distributions and enhanced fluorescence emission without any additional spacer.

SUMMARY

In an exemplary embodiment, a method of increasing a fluorescence signal of a fluorophore is described. The method includes dissolving the fluorophore in a solvent to form a solution and adding silver nanoparticles to the solution to form a mixture. The silver nanoparticles have a size of 3-10 nanometers (nm) and the silver nanoparticles have an irregular shape and at least one fractal structure. The method further includes at least partially coating a substrate with the mixture to form a fluorescence sample and recording a fluorescence image of the fluorescence sample. The fluorophore is adsorbed to at least one silver nanoparticle and the fluorescence signal of the fluorescence image is higher than a fluorescence signal of a fluorescence image of the fluorophore without the silver nanoparticles.

In some embodiments, the silver nanoparticles have a substantially crystalline structure.

In some embodiments, the silver nanoparticles have hydroxide (—OH) and nitrogen oxide (—NO2) functional groups on a surface.

In some embodiments, at least a portion of the silver nanoparticles have at least two fractal structures in an individual nanoparticle.

In some embodiments, at least a portion of the silver nanoparticles have at least two different lattice fringes in an individual nanoparticle.

In some embodiments, the lattice fringes are in different directions.

In some embodiments, the lattice fringes are in the same direction.

In some embodiments, a lattice fringe has a d-spacing of 0.20-0.24 nm and corresponds to the [200] crystal structure of silver.

In some embodiments, a lattice fringe has a d-spacing of 0.22-0.26 nm and corresponds to the [111] crystal structure of silver.

In some embodiments, at least a portion of the silver nanoparticles are twinned.

In some embodiments, the silver nanoparticles have an optical bandgap of 2.8-2.9 electron volts (eV).

In some embodiments, the silver nanoparticles have a maximum absorption signal at 350-400 nm.

In some embodiments, the silver nanoparticles have a maximum fluorescence signal at 420-440 nm.

In some embodiments, the silver nanoparticles have an electromagnetic field intensity of 6-60 volts per meter (V/m).

In some embodiments, the fluorophore is selected from a group consisting of a rhodamine derivative, a fluorescein derivative, a cyanine derivative, and a coumarin derivative.

In some embodiments, the fluorescence sample is irradiated with 300-330 nm light to record the fluorescence image.

In some embodiments, the fluorescence signal of the fluorescence image is 10 to 10,000 times higher than a fluorescence signal of a fluorescence image of the fluorophore without the silver nanoparticles.

In some embodiments, the silver nanoparticles are made by a method comprising dissolving sodium borohydride in water at a temperature of 5 to 15 degrees centigrade (° C.) to form a 1-3 mM sodium borohydride stock solution, dissolving silver nitrate in water at a temperature of 20 to 25° C. to form a 0.5-2 milli Molar (mM) silver nitrate stock solution, and adding the silver nitrate stock solution dropwise to the sodium borohydride stock solution at a temperature of 5 to 15° C. to form a silver nanoparticle solution. The sodium borohydride stock solution and silver nitrate stock solution are added together within 30 minutes of dissolving the sodium borohydride and silver nitrate in water.

In some embodiments, the rate of dropwise addition is 1-10 drops per second.

In some embodiments, the method includes centrifuging the silver nanoparticle solution and filtering to leave a solid and washing the solid with water to form the silver nanoparticles.

The foregoing general description of the illustrative present disclosure and the following detailed description thereof are merely exemplary aspects of the teachings of this disclosure and are not restrictive.

