Possibilities in melanoma diagnostics using surface-enhanced Raman scattering biosensor

Possibilities in melanoma diagnostics using surface-enhanced Raman scattering biosensor

In a recent study published in Journal of Physical Chemistry C, researchers demonstrated an approach to detect tyrosinase activity (TYR), an important biomarker for the diagnosis of melanoma, using an unprecedented transportable surface-enhanced Raman scattering biosensor.

Study: Surface-Enhanced Raman Scattering Biosensor Based on Self-Assembled Gold Nanorod Arrays for Rapid and Sensitive Detection of Tyrosinase. Image credit: Africa Studio/Shutterstock.com

The surface-enhanced Raman scattering biosensor, fabricated on a glass chip, is developed on dopamine (DA)-functionalized Au nanorod (Au NR) arrays that act as capture substrates and 4-mercaptophenylboronic acid (4-MPBA)-modified silver nanoparticles (Ag NPs) that form the surface-enhanced Raman scattering biosensor probe. Since detection of TYR activity in biological samples is essential for clinical melanoma diagnosis, the proposed approach with various advantages of sensitivity, portability and reproducibility may be beneficial for melanoma diagnosis.

Tyrosinase and its role in melanin synthesis

Tyrosinase, a type III dinuclear copper-containing metalloenzyme, plays an important role in the biosynthesis of melanin. As a result, its inhibition can prevent the development of several skin diseases. Furthermore, this human enzyme characteristic of melanogenesis shows thorough catalytic enzyme activity in vivo and catalyzes the hydroxylation of L-3,4-dihydroxyphenylalanine and oxidation of dopamine (DA) to dopaquinone (DQ). Consequently, aberrant TYR expression can cause skin conditions such as melasma, vitiligo, and the most lethal manifestation of skin cancer, melanoma.

Since TYR is a pivotal biomarker for melanoma diagnosis, various methods, including the electrochemical technique, colorimetric method, high-performance liquid chromatography, and fluorescence, have been tried and tested to detect TYR in biological samples so far.

The inherent limitations of these techniques, such as relatively low sensitivity, susceptibility to interference, and unreasonable equipment, make the surface-enhanced Raman scattering biosensor a highly desirable alternative approach.

Surface-enhanced Raman scattering biosensors use a powerful non-destructive, ultra-sensitive detection technique that utilizes the molecular interaction between molecules adsorbed on nanometallic planes and the electromagnetic field present in the resonance region.

The present study developed a portable surface-enhanced Raman scattering biosensor to detect TYR activity synthesized on the Au NR arrays. The efficient detection of TYR activity results in a successful melanoma diagnosis.

Au NRs and their preparation

To achieve a significant nanorod yield, excellent uniformity, and minimum amount of impurities, Au NRs were prepared using a more advanced method. Initially, 0.025 ml of a 10 mM HAuCl4 solution and 0.1 M CTAB in a volume of 1 mL were combined to create a seed solution. After 30 minutes of stirring, 0.8 ml of newly produced NaBH was added4 at 10 mM was added to the mixture.

The seed solution generated was undisturbed at 30°C for two hours. A total of 7 g of CTAB and 1.234 g of NaOL were mixed in 250 mL of H2O at 50 ⁰C to produce the growth solution. 18 ml of a 4 mM AgNO3 solution was added when the solution was cooled to 30°C under constant stirring.

After adding 250 ml of 1 mM HAuCl4 and 90 minutes of stirring, the solution gradually turned from orange to colorless. After stirring for 15 minutes, 2.1 ml of a 37°C HCl solution was added. The culture solution mentioned above was mixed for 30 seconds with 64 mM AA in a volume of 1.25 ml before being injected with 0.8 ml of gold seed mixture, which was then allowed to stand for 48 hours at 30 °C.

The transfer and compaction at three-phase interfaces driven by the Marangoni reaction formed the basis for synthesizing the Au NR arrays.

The successful synthesis of Ag NPs was achieved by the Lee and Meisel method, which included the reduction of AgNO3 by citrate in an aqueous phase.

Detection of TYR in the serum sample

The serum samples were centrifuged before analysis at 1000 rpm for 10 minutes to collect the supernatant and reduce the other substances that would prevent the detection of TYR in the sample, thereby interfering with the successful melanoma diagnosis.

A Raman spectrometer with 785 nm excitation, 50X objective lens and an exposure time of 10 s was then used to derive Raman spectra.

The TYR solution was mixed with different concentrations of inhibitors and incubated for 15 min at 37 °C to achieve inhibition. Later, the surface-enhanced Raman scattering measurements were performed on this inhibitor-treated TYR solution by introducing the surface-enhanced Raman scattering biosensor.

New biosensor efficiently detects TYR activity

The experimental results of the present work revealed that the recoveries were in the range of 96.85-98.74% when known amounts of different concentrations of TYR were added to the serum.

The method studied was further used to screen TYR inhibitors, and an inverse proportional relationship was determined where an increase in the inhibitor concentration resulted in a weaker TYR activity.

These results indicated that the surface-enhanced Raman scattering biosensor can be used for quantitative analysis of TYR activity, an important biomarker for melanoma diagnosis and screening of TYR inhibitors.

Conclusion

The state-of-the-art portable surface-enhanced Raman scattering biosensor presented in the current work is fabricated on a glass chip and is based on Au NR arrays.

This new approach to detect TYR activity in biological samples necessary for preclinical melanoma diagnosis has a competitive advantage over its precursors due to its multiple advantages, including sensitivity, portability, and reproducibility.

This technique has a wide linear detection range that can effectively assess 0.0001 U/mL TYR activity without any complex nanofabrication. The proposed method also eliminates the accumulation of nanoparticles and has a convenient structure, making it a preferred choice for the diagnosis of melanoma.

References

Minling Li et al. (2022) Surface-enhanced Raman scattering biosensor based on self-assembled gold nanorod arrays for rapid and sensitive detection of tyrosinase. Journal of Physical Chemistry C. https://pubs.acs.org/doi/10.1021/acs.jpcc.2c03408

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