## 1 Introduction  
Raman spectroscopy is a branch of vibrational spectroscopy that allows highly sensitive structural identification of various chemical and biological materials based on their unique vibrational characteristics, all without destroying the sample. Raman spectroscopy is an effective tool for analytical studies, but the low intensity of Raman signals is a major disadvantage of the method [1]. Surface-enhanced Raman scattering (SERS) is a commonly used technique to enhance the signal, that allows the analysis of low-concentration samples or even the detection of a single molecule. The SERS effect can occur when the analyte is in close vicinity of a nanoscale-structured metal surface. By using metal surfaces with optimal parameters, the intensity of Raman signals can be enhanced by several orders of magnitude [2, 3, 4]. It is now accepted that two types of mechanisms play a role in an increase in the intensity of Raman signals: the electromagnetic (EM) enhancement [5] derived from electromagnetic fields generated by plasmon excitation of metal particles serving as SERS substrates, and the chemical amplification [6, 7], which allows the target molecule to transfer electrons to the metal particles, often leading to the formation of a chemical bond between the metallic surface and the molecule [2]. For SERS measurements, plasmonic metal substrates made of silver, gold, or copper are used in most cases [8], which are synthesized by various methods. SERS active agents can be divided into three categories: (1) metal nanoparticles (MNPs) in suspension, (2)MNPs immobilized on solid substrates, (3) nanostructured surfaces fabricated directly on solid substrates by using nanolithography and template synthesis [9]. Manufacturing technologies allow tuning of the size and shape of the nanoparticles, which can also greatly influence the degree of enhancement [10; 11]. To produce substrates with optimal parameters providing maximum SERS enhancement [12], it is necessary to understand how the material, size, shape, and arrangement of nanoparticles, or the excitation source, affect SERS efficiency [13; 14; 15]. If the particle size is too small, the light scattering properties will decrease, resulting in a decrease in SERS enhancement. As the particle size increases, the SERS effect also increases as the number of available free electrons is higher, which plays an important role in the excitation of plasmons [8]. In our previous work, examining the effect of size and distance of Au and Ag nanoparticles generated by heat treatment of a thin metallic layer produced by vacuum evaporation, we found that the SERS effect increases linearly with particle size, as larger particles function as larger scattering centers [12]. In addition, by reducing the distance between the particles, hot spots will be located closer to each other, leading to the formation of highly concentrated EM fields, which results in the enhancement of Raman signals [12; 16; 17; 18]. To effectively optimize nanoparticles for SERS, the plasmon wavelength of the particle must be considered as well. The localized surface plasmon resonance (LSPR) peak of the plasmonic material must be close to the wavelength of the excitation source or the inelastically scattered light. The LSPR wavelength of a plasmonic material depends on the material, size, and shape of the particle. In the case of gold and silver, the plasmon wavelength is between 500 and 800 nm, and 400 and 700 nm, respectively. Furthermore, the plasmon peak shows a redshift with increasing size of the nanostructures and distance between the particles. If the wavelength of the excitation source is far from the LSPR peak of the plasmonic material, the extent of EF also decreases [12; 13]. In addition to the parameters of the nanoislands, the adsorption of the analyte to their surface should play an important role in the SERS process.