# Metallic Nanoparticles Obtained via \"Green\" Synthesis as a Platform for Biosensor Construction
## 1 Introduction  
Green synthesis of metallic nanoparticles (NPs) offers better manipulation, stabilization and control over crystal growth due to slower kinetics. Furthermore, green synthesis processes are rapid, eco-friendly and cost-effective [1; 2; 3]. Metallic NPs consisting of one metal, metal oxides or a composite of several metals play a significant role in nanotechnology due to their potential utilization in industry and medicine [3; 4; 5; 6; 7]. NPs of noble metals possess low cytotoxicity, enable easy modification of their surfaces, have straightforward synthesis processes and excellent biocompatibility [8; 9; 10]. Such advantages make metallic NPs obtained by green synthesis (gNPs) prospective for applications in biological analysis, drug delivery and imaging, environmental monitoring, industrial catalysis and electronic devices [11; 12; 13; 14; 15; 16; 17; 18; 19]. A special role of these gNPs is related to antimicrobial and antibiofilm agents [19; 20; 21; 22; 23; 24; 25]. Many plants [11; 12; 21; 24; 25; 26], algae [4], microorganisms [1; 3; 7; 8; 9; 10; 14; 18; 23; 27; 28; 29; 30], as well as redox-imbalanced mammalian cells and systems [31] are known to produce nanostructured mineral crystals and metallic NPs with properties similar to chemically-synthesized materials. Biological agents, including polysaccharides, polypeptides, DNA, enzymes secreted by cells [24; 32; 33; 34; 35; 36; 37; 38; 39; 40; 41] and purified enzymes [42; 43] are able to reduce noble metal ions to gain metallic NPs. Besides the practical aim of gNPs production, investigations on the green synthesis processes will be useful for elucidating the mechanisms of cells' responses to stress caused by metal ions. In many studies, the size and morphology of gNPs are considered as their basic parameters. Scanning electron microscopy (SEM) can afford information on the size, distribution and shape of the gNPs. However, the drying and contrasting processes alter the characteristics of the tested NPs and lead to imaging faults or artifacts [32]. Transmission electron microscopy (TEM) is a high-tech approach in the characterization of gNPs, and is more powerful and competitive than SEM. It can be used to determine the particle size, dispersion and aggregation in aqueous environment with a high spatial resolution that can reveal details of crystal structure. Since many gNPs are irregular in shape and exhibit a tendency to form large particles, it is difficult to conclude definitely whether their final form was caused by polydispersity or agglomeration. The most suitable method for characterization of gNPs is a dynamic light scattering (DLS) technique, which can provide data on sphere hydrodynamic diameters of homogeneous NPs in an aqueous phase [44]. However, this method cannot provide information on particles with a heterogeneous size distribution. Therefore, the diversity and ambiguity of gNPs are the reason why it is necessary to use multiple techniques, for instance a combination of TEM and DLS, for valid characterization of gNPs [32; 44]. The exact mechanism of gNP synthesis using biological agents has not been elucidated to date. However, it has been suggested that various biomolecules are involved in this process [24; 38; 39; 40; 41; 42; 43; 44; 45; 46; 47; 48]. Capping and stabilizing agents (polysaccharides, polypeptides and other bioorganic compounds) prevent the gNPs from further growth and agglomeration in resulted colloidal solution [45; 46; 47; 48; 49; 50; 51]. Extracellular synthesis of gNPs usually occurs when microbial cells secrete reductive enzymes on the surface of cell wall or outside the cell [43; 45; 46; 47; 48]. This process mediated through different enzymes, including hydrogenases [40], nitrate reductase [48], phenol-oxidizing enzymes (lacases, tyrosinases, and Mn-peroxidase) [43], as well as adhesins [49], has been described. In addition to the reducing extracellular metabolites, the cell wall of microorganisms seems to play an important role in the intracellular synthesis of NPs [8; 27; 44; 45; 46; 47].