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Week 2

BMS Away Day
Firth Court Entrance

At the beginning of the week I attended the Biomedical Science (BMS) research away day, which was held in Firth Hall. The day consisted of numerous talks and poster presentations given by various members of the four research areas which exist within the BMS department at the University of Sheffield. Namely, cell biology and cancer, developmental biology and disease, neuroscience, and stem cell and regenerative medicine. The day was really interesting as it allowed me to explore the work that is going on within the department; in addition to giving me an indication as to how to approach poster and flash talk presentations when I (hopefully) become a PhD student.  

Laboratory Work

Experiment 1: Microscopy
This week I had my first go at microscopy; using standard transmitted light before moving on to stain some samples and use fluorescence. However, I wasn't able to gain any results from these latter experiments as our cells were contaminated. As the fluorescent dye itself appeared to be working fine, we are hoping to simply repeat the staining procedure and hopefully get some usable data next week.
The Microscope

Candida Hyphae Visualized under Transmitted Light using 150X Objective

 Experiment 2: Caspofungin Concentration Sensitivity Plates
This experiment was very similar to that of the cell wall sensitivity plate assay; however, this week we simply focused on only one chemical agent (caspofungin) and tested a broader range of concentrations.  

 Experiment 3: Alcian Blue Assay Take Two
I ran through the alcian blue staining experiment again on Friday, this time including 3 biological repeats for each Candida strain in both yeast and hyphal growth conditions. This meant having 12 tubes in total, which equated to somewhat of a lengthy cell counting session! Although I did get results for both yeast and hyphal cells, I decided not to conduct statistics on the latter, as they did not pellet very well and re-suspended almost immediately following centrifugation. This made it very hard to actually collect supernatant following alcian blue incubation, which made the relevance of the results questionable. This was in stark contrast to using yeast cells, which didn't seem to cause any trouble at all. As alcian blue binds to phosphomannan, I used my results to infer differences in phosphomannan content between wild-type and mutant strains.
The loss of apm4 increases phosphomannan content in C. albicans. Yeast cells from wild-type (WT) and apm4Δ/Δ mutant strains of C. albicans were incubated with 30µg/mL alcian blue. Values plotted are the mean and standard deviation obtained from 3 refreshed cultures. *p = 0.0117 (unpaired t test, n=3)


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