Cell biology labwork - 4PMBCBP9

Goal(s)

In these practicals the student learns eucaryote cell culture techniques. Qualitative and quantitative analysis methods to evaluate cellular function are taught.

Content(s)

During these practicals 2 main experiments are proposed: Transfection and Intracellular pH assessment

1. Transfection: Eucaryote HeLa cells are cultivated for this experiment. A transitory transfection is done using plasmids that allow fluorescence localisation in the cell using targeted proteins or organelles. Transfected cells are observed and analysed using fluorescence microscopy. Images are aquired and interpreted.
2. Intracellular pH: Intracellular pH is assessed using yeast expressing a pH sensitive fluorescent protein (pHluorin), targeted either in the cytosol or in the mitochondria. In addition, a membrane permeant fluorophore is used to measure intravacuolar pH. A calibration curve is established. The cell responses to glucose addition is investigated, in the presence of various inhibitors. Quantitative data are recorded and analyzed to measure intracellular pH gradients, in relation to cell physiology.

Basics in molecular and cell biology