Application of Western Blotting to the Identification of Metallothionein Binding Proteins

✍ Scribed by Cecilia M. Whitacre

• Publisher: Elsevier Science
• Year: 1996
• Tongue: English
• Weight: 84 KB
• Volume: 234
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1006/abio.1996.0056

✦ Synopsis

vidual test tubes (9). Using only every other well, a complete separation of the 14 CO 2 spots was achieved. SAMDC or ODC in attached cells could also be measured; however, due to low cellular enzyme activity the signal obtained was rather low and required extensive exposure times of ca. 7 days on the Phosphor-Imager screen (data not shown).

In conclusion, we have developed a system for the detection of decarboxylation reactions within surfaceattached cells. This method allows the study, e.g., of mitochondrial function in tissue culture cells without the need to detach the cells from their solid support, which may introduce undefined changes of cell surface FIG. 3. Phosphor-Imager analysis of an assay for inhibition of soluproperties as well as of the metabolic status of the cells. ble rat liver ODC by aminooxy-propanamine derivatives. Rat liver

The method can also be applied to detection of cell-free ODC (1 mg/well, sp act 32 pmol CO 2 released from ornithine/mg decarboxylation reactions in a multiwell format. protein and min) was incubated for 1 h at 37ЊC after addition of inhibitor (row A wells 1