Metallothionein (MT) is a low-molecular mass protein playing an essential role in homeostasis of heavy metal ions. Its relation with formation and progression of a tumour disease is discussed in this article. Here, we propose a new methodological approach for visualization of MT on PVDF membranes after dot- and electroblotting by using a commercial mouse monoclonal antibody E9 and polyclonal chicken antibodies. The optimized procedure was as follows. We dotted 1 microL sample volume on PVDF membrane and let it to dry. Then, we blocked the membrane surface with 2% BSA in PBS for 30 min. After that, the membrane was incubated in chicken primary antibody (diluted 1:500), washed, and incubated in rabbit-anti-chicken secondary antibody conjugated with horseradish peroxidase. To visualize the interaction, we used 3-aminoethyl-9-carbazole. Under these conditions, we estimated detection limit as 3 pg of MT per 1 microL. The optimal approach was further utilized for detection of MT level in two human fibroblast cell lines and in blood serum obtained from children with medulloblastoma. The results were in good agreement with differential pulse voltammetry-Brdicka reaction.
- MeSH
- dítě MeSH
- fibroblasty metabolismus MeSH
- imunoblotting metody MeSH
- kadmium farmakologie MeSH
- kolodium MeSH
- kur domácí imunologie MeSH
- lidé MeSH
- meduloblastom MeSH
- metalothionein analýza krev metabolismus MeSH
- nádorové buněčné linie metabolismus účinky léků MeSH
- polyvinyly MeSH
- protilátky diagnostické užití metabolismus MeSH
- transformované buněčné linie MeSH
- western blotting metody MeSH
- zvířata MeSH
- Check Tag
- dítě MeSH
- lidé MeSH
- zvířata MeSH
Acta dermato-venereologica, ISSN 0001-5555 suppl. 213, May 2003
47 s. : tab., il. ; 30 cm
- MeSH
- antipruritika MeSH
- epidermolytická ichtyóza MeSH
- ichtyóza MeSH
- kolodium MeSH
- pruritus terapie MeSH
- Geografické názvy
- Švédsko MeSH
- Konspekt
- Patologie. Klinická medicína
- NLK Obory
- dermatovenerologie
