Isolation of metallothionein from cells derived from aggressive form of high‐grade prostate carcinoma using paramagnetic antibody‐modified microbeads off‐line coupled with electrochemical and electrophoretic analysis

Abstract

Prostate cancer with altered zinc(II) cell metabolism is the second most frequently diagnosed cancer in developed countries. The alterations of zinc(II) metabolism can influence metabolism of other metal ions and can also be associated with the expression and translation of metal-binding proteins including metallothioneins. The aim of this article was to optimize immunoseparation protocol based on paramagnetic beads conjugated with protein G for the isolation of metallothionein. Isolated metallothionein was determined by differential pulse voltammetry Brdicka reaction and SDS-PAGE. Optimal conditions: antigen-binding time - 60 min, temperature - 70°C, and buffer composition and pH - acetate buffer, pH 4.3, were determined. Under the optimized conditions, lysates from 22Rv1 prostate cancer cells treated with various concentrations of cadmium(II) and copper(II) ions were analyzed. We observed strong correlation in all experimental groups and all lysate types (r>0.83 at p<0.041) between metallothionein concentration related to viability and concentration of copper(II) ions and cadmium(II) ions in medium. Moreover, the results were compared with standard sample preparation as heat treatment and SDS-PAGE analysis.

Metallothioneins and zinc in cancer diagnosis and therapy

“…MT can serve as a prognostic marker in central nervous system tumors of childhood and adolescence (Rickert and Paulus, 2005;Rickert, 2004), osteosarcoma (Trieb and Kotz, 2001), breast cancer (Woolston et al, 2010;Jin et al, 2004;Gomulkiewicz et al, 2010), pancreatic islet cell tumors (Tomita, 2002), and tongue squamous-cell carcinoma (Theocharis et al, 2011). MT can also serve as a serum tumor marker in prostate cancer Masarik et al, 2011), head and neck tumors , childhood solid tumors , and melanoma . Expression of MT may help to distinguish between benign and malignant tumor, as shown in thyroid tumors (Krolicka et al, 2010), prostatic lesions (El Sharkawy et al, 2006), GI stromal tumors, and gastric carcinomas (Soo et al, 2011).…”

Sharka: The Past, The Present and The Future

“…Therefore, methods of nanotechnology take advantage of nanoparticles on the basis of surface modifications possibility, which can significantly improve the quality of isolated RNA. Magnetic nanoparticles have been used for isolation and purification of nucleic acids; the use of magnetic nanoparticles provides very rapid and simple isolation of nucleic acids [449,450,451,452,453,454,455,456]. In addition, magnetic nanoparticled surface may be variously modified (these modifications include for example the introduction of silanol, epoxide, diol and carboxyl groups respectively).…”

Improved Electrochemical Detection of Zinc Ions Using Electrode Modified with Electrochemically Reduced Graphene Oxide

“…Zinc has these important roles and effects mainly as a co-factor of numerous proteins, therefore it is not surprising that metallomics and proteomics of zinc-containing proteins are emerging fields of science [6,7,8]. From these, metallothioneins are highlighted as maintainers and transporters of these proteins and their importance in zinc metabolism belongs to the interest of numerous researchers [9,10,11,12,13,14,15,16,17,18]. …”