Hetero nano-enzyme structures having multiple functions and their elisa applications

In this study, nanoparticles having multiple functions were synthesized using ruthenium based photosensitive amino acid monomers by cross-linking technique (ANADOLUCA) and the use of synthesized nanoparticles in ELISA (Enzyme-Linked ImmunoSorbent Assay) applications was investigated. For this purpose, firstly, cross-linked nano structures were prepared by microemulsion polymerization technique at room temperature under daylight and nitrogen atmosphere using horseradish peroxidase (HRP), alkaline phosphatase (ALP) enzymes and bis (2-2'-bipyridyl) (MATyr)2 rutenium (II) photosensitive monomers. Then, these structures were characterized by transmission electron microscopy (TEM), atomic force microscopy (AFM), zeta potential measurements. In the second part of the study, kinetic studies of the nanoenzymes were carried out. The obtained results showed that hetero nano-enzyme exhibited maximum activity towards tetramethylbenzidine (TMB) and paranitrophenyl phosphate (p-NPP) as substrates at pH 5 and pH 10, respectively. The kinetic parameters Km and Vmax values were 26 mM and 1,24 x 10-2 U for TMB and 0,516 mM, 1,70 x 10-4 U for p-NPP. The free enzymes denaturate at high temperatures. However, hetero nano-enzyme structure has not loss its activity. The reusability studies showed that there is no significant loss of activity of the nanostructure washed with ethanol even after 15 cycle. In the final step of the study, nano enzyme structure was conjugated with antibody and ELISA applications were carried out.