In my previous blog post, I discussed the work I did last
Thursday and Friday in which I made a new growth medium for the co-cultured
TMSC’s/CSSC’s and lysed BCE cells. This brings me to the work I completed on
Monday, Tuesday, and Wednesday.
When I arrived at the lab on Monday, I immediately checked the stem cell's growth. Overall, they looked good. As seen in the pictures below, some of the big clumps of cells
(which are the TMSC’s/CSSC’s) have anchored themselves on the bottom of the
wells, and have begun to migrate towards the extra cellular matrix of the lysed
BCE cells. In most of the wells,
however, the stem cells did not seem anchored or migrating yet; this is ok
because it is still early, so by the end of the week they should be. After this, I changed the growth medium of
the cells. This can be a tedious process
because since many of the stem cells are still free floating in the growth
medium, they can be mistakenly sucked out in the process of taking the old
medium out. To prevent this, I have to
turn the well plates at an angle so that many of the stem cells will drift
towards the bottom, and then, using a small micropipette, very slowly take out
the old medium. If there are many free-floating
stem cells, then I leave about 1/3 of the old medium in the well plates to
ensure they are not taken out. When I
finished changing the medium of all the cells, I spent the rest of the day
taking pictures of the cells using a special camera-microscope. This took longer than expected because after one round of pictures, I uploaded them onto my
computer, only to find that the lighting on the pictures was totally off and
therefore the cells were barely visible.
With the help of Dr. Du, I changed the settings on the microscope so
that this would not happen again, and then proceeded to take four pictures of
cells from each of the eight wells.
Specifically, we were looking to take pictures of the initial stages of
the stem cells being anchored to the bottom of the well plates as well as some
of the stem cells that were beginning to migrate away from their central bundle. I will take
another series of pictures on Thursday or Friday to see how the cells have
developed.
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A group of TMSC's that are beginning to migrate
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Tuesday was one of the lighter days I have had in the lab in
terms of work. Besides briefly checking
on the stem cells a couple times throughout the day, I used the free time to
work on my poster as well as continue to research some of the topics that my
experiment is about, particularly the biology of corneal endothelial cells and
current treatments for a loss of cellularity in these cells. I am also
preparing for a presentation of my project for my lab mates which I will
present during our lab meeting next Tuesday.
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| A staining of TMSC's |
Yesterday (Wednesday), I spent the majority of my day in the
confocal microscope room. I worked with Ye on one of my other projects which involves
immunostaining TMSC’s which enables us to locate certain proteins.
Specifically, we were testing for the presence of 3 proteins: Fibulin, Prox-1,
and Ve-Cad, which were stained 3 different colors. The microscope is very complex and is very
expensive ($350,000) so I am always a little nervous using it. Most of the staining turned out great,
however one of the samples was very blurry. After playing around with some of
the settings regarding lighting and composure on the microscope, Ye and I
determined that we must have not used enough of the fluorescent antibodies for
that one sample. Nevertheless, the majority of the pictures turned out great,
and I included on the left.
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| The intimidating sign that is purposely placed right above the confocal microscope. |
Outside of the lab, I keep myself busy between doing summer
work for school, babysitting my cousins, and exploring Pittsburgh. On Wednesday night, I biked down to one of
Pittsburgh’s biggest parks, Schenley Park, to enjoy some live music. The weather here during the day has not been
the best; it has rained every single day this week and is supposed to for the remainder of the week into Sunday. My time here has flown by and I cannot believe next week will be my last.
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