Monstervoorbereiding voor metabolomics: een methode om celmonsters voor te bereiden op metabolietprofilering

Metabolomics is the study to identify and quantify metabolites present within the cells. To prepare the sample, begin by seeding breast cancer cells along with growth medium in two plates labeled as test and control. Incubate for three days at 37 degrees Celsius. Now, remove the medium and add fresh medium with estradiol in the test plate and plain medium in the control plate.

The estradiol binds to estrogen receptor alpha in breast cancer cells to induce certain expressions, causing a change in the metabolite composition. Incubate the plates for the desired duration. Replace the medium with ice cold phosphate buffered saline or PBS to wash the cells and then add ice cold acetone. The ice cold reagents stop the action of proteases and thus prevent protein degradation.

Now use a scraper to detach the cells from the plates, and transfer them to tubes with the help of a pipette. Take 20 microliters of this cell suspension onto a hemocytometer and count the number of cells. Store the samples at minus 80 degrees Celsius until used for further analysis. In the following protocol, we prepare samples from MCF-7 cells for gas chromatography and mass spectrometry.

Using MCF-7 cells cultured