Field of research: Cell Biology
Lab: Derrick Brazill
School: Manhattan Hunter Science High School
Year: 10th/11th Grade
Determining how Over Expression of PakD affects Dictyostelium discoideum in Under the Agarose Folic Acid Chemotaxis Assays
Good health is a very important aspect of life and being able to stay healthy allows organisms to live longer. However, there are illnesses that jeopardize our good health and it is important to understand these diseases. For example, there isn’t much we know of about cancer metastasis; this is when cancer cells spread from a primary tumor to form secondary tumors in the body. Understanding chemotaxis, the phenomenon in which cells move to a certain chemical in their environment, helps to understand cancer metastasis.
Dr. Brazill’s lab studies Dictyostelium discoideum, single-celled amoebas, which are good model organisms. The chemotactic characteristics, speed, directionality and chemotactic index, of D. discoideum are studied in the lab, to see what protein are involved in folic acid chemotaxis. Under the agarose folic acid chemotaxis assays were used to compare the speed, directionality and chemotactic index of PakD over expressing (OE) cells to control wildtype (WT) cells.
In performing the folic acid chemotaxis assay, time-lapse microscopy was performed on WT and PakDOE cells as they moved underneath the agarose towards higher concentrations of folic acid. Images are taken every 30 seconds using an inverted microscope and Metamorph Software. Afterwards, those images were further analyzed using Image J and Microsoft Excel Software.
For each experiment, 15 cells of each cell type were manually tracked to find their speeds, directionalities and chemotactic indices. The averages for each experiment were compiled and graphed to determine the average speed, directionality and chemotactic index for both cell types. T-tests were run to see whether the two groups had statistically different results. Since the P values, a measure of significant difference, for the findings were over 0.05, there was no significant difference. Therefore PakD is not involved in folic acid chemotaxis in D. discoideum.
This was a great experience and I hope that I can further continue my research.