My confocal images!


















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Day 27 and 28: Assays, secondary antibody incubation and splitting cells: The assays: I ran assays for all of the OGD plates for Hoechst, MTT and LDH and the results were as follows: Secondary antibody incubation for immunofluorescence: I had to incubate the coverslips with secondary antibodies against the TOMM20 (which would be an indicator of mitochondrial morphology and presence) and Drp1 (which would be an indicator of fission. TOMM 20 were going to appear red whilst Drp1 was going to appear green on the confocal microscope. I incubated the coverslips in darkness for the rest of the day. At the end of the day, I mounted the coverslips onto slides which contained DAPI which would mark the nucleus and this would appear blue on the confocal microscope. Mounting the coverslips onto the slides: The slides were labelled with their relevant characteristic and the cloverslips were placed on top. The coverslips were secured in place with nailvarnish to the micro...
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Week 4: Inhibiting mitochondrial fission using p110 peptide varients in an attempt to prolong cell survival during neonatal hypoxic ischaemia

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The theory behind the experiment: - There is an increase in mitochondrial mitophagy during neonatal hypoxic ischaemia. The cell becomes stressed as a result of the reduced oxygen in the environment. Reactive oxygen species (ROS) being formed after electrons react with molecular oxygen in the electron transport chain. The mitochondria which have become damaged will undergo the process of mitochondrial fission, mediated by the proteins Drp1, MFF, MID49/51 and Fis1. The mitochondria which have accumulated damage will have their damaged features taken to just one area in the mitochondrion. Once fission has occurred and one mitochondrion has become two, the mitochondrion which had accumulated its damage will undergo mitochondrial mitophagy. - As a result, less ATP is available for the cell but more ROS is produced. In addition, the mitochondrial membrane also becomes leaky releasing apoptosis stimulating factors and cytochrome c, therefore stimulating apoptosis and exaggerating th...
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Day 15: Simply splitting cells I split cells from the two flasks that I left incubating last week and combined them into two flasks but with different media. One flask contained the media that I had previously used before doing my own oxygen glucose deprivation experiments while the other contained a different medium. On Monday (day 16) I will compare the confluence of the cells in both flasks and hopefully will be conducting more cell culture experiments next week!
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