Objective and Purpose: In lab 6, we are trying figure out how the microplastics (polypropylene) take effects on the Tetrahymena on the movement speed, concentration and the changing of the direction. We also cut the plastics and count the cells of the Tetrahymena , calculate the average concentration of my own sample. We practiced the skill of the micropipettes, dissecting microscope and compound microscope for the future experiment. Describe the potential effect of plastics on soil microbial ecosystems, determine the dilutions required to make dilute solutions from a stock solution.

Materials: Concavity slide, Sterile container, polypropylene, Tetrahymena sample, Iodine, Compound microscope, micropippetors, 1 uL of India Ink, coverslip, dissecting microscope.

Procedure:

1. Shred/cut the polypropylene twine into small pieces with scissors.
2. Measure 0.5 of polypropylene (PP) into a sterile glass jar/beaker.
3. Add 50ml of sterile proteose-peptone-tryptone media.
4. Each group use the aliquoted culture to determined the concentration in cells/mL
5. Add 20 uL of TH to 5 uL of Iodine on a Petri Plate lid. Mix up and down by pipetting. Add 3 separated 5ul drops to a slide.
6. Observe without the coverslip by 4x or 10x objective, record and counts the cell/ML.
7. Take out another concavity slide and coverslip.
8. Transfer 20 uL Tetrahymena sample by micropippetors onto the concavity slide and mix with 5uL Iodine Ink, put the coverslip quickly.
9. Put Vaseline around the coverslip to make sure it is stable.
10. Observe under the compound microscope by 400x lens every ten minutes and record the number of vacuoles of 10 different cells.
11. Answer the QTM and discuss with team members.
12. Clean up the lab table.

Data：

Storage: I cleaned up the concavity slide, Sterile container, coverslips under water and put on the tissue. Plug out the microscope and put back the micropippetors.

Conclusion and Future Goal: The vacuoles become more clearly to see and the cells gradually move slower by the time passed. I tried to count the vacuoles accurately, but the Tetrahymena moved super fast and I couldn’t count vacuoles in accuracy in the first 10 minutes. We calculated the concentration of the cells, even though I didn’t try the dilute because I didn’t have enough time, but I got a good observation data of the number of the cells. We cut the polypropylene twine into small pieces with scissors at the beginning of the class and practiced the skills of use micropippetors and compound/dissecting microscope in the future lab/experiments.