Using of chicken antibodies for metallothionein detection in human blood serum and cadmium-treated tumour cell lines after dot- and electroblotting
Language English Country Germany Media print
Document type Journal Article
- MeSH
- Child MeSH
- Fibroblasts metabolism MeSH
- Immunoblotting methods MeSH
- Cadmium pharmacology MeSH
- Collodion MeSH
- Chickens immunology MeSH
- Humans MeSH
- Medulloblastoma MeSH
- Metallothionein analysis blood metabolism MeSH
- Cell Line, Tumor drug effects metabolism MeSH
- Polyvinyls MeSH
- Antibodies * metabolism MeSH
- Cell Line, Transformed MeSH
- Blotting, Western methods MeSH
- Animals MeSH
- Check Tag
- Child MeSH
- Humans MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Names of Substances
- Cadmium MeSH
- Collodion MeSH
- Metallothionein MeSH
- polyvinylidene fluoride MeSH Browser
- Polyvinyls MeSH
- Antibodies * MeSH
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.
References provided by Crossref.org
ELISA-like Analysis of Cisplatinated DNA Using Magnetic Separation
Serum metallothioneins in childhood tumours-a potential prognostic marker