In research, you have a lot of long days. Last week, we had a particularly long day. We were finishing an experiment that involved treating mice with leukemia with a drug that we hope might be a potential therapy for Juvenile Myelomonocytic Leukemia. The mice get leukemia because they express an inducible knock-in gene that can be turned on in the mouse’s hematopoietic cells. Unfortunately, the leukemia these mice get is mild and takes a long time to manifest itself, and isn’t fatal like it is in humans. As a result, we decided to treat these mice with the drug over a period of 4 months, alternating 2 weeks on the drug and 2 weeks off. To complete this experiment, we wanted to harvest tissues from these mice to assess the extent of the leukemia that they have.
After sacrificing the mice, we had to harvest the spleens and weigh them (the most profound phenotype of the leukemia in these mice is splenomegaly, or enlarged spleen). We also used some splenic tissue for histological analysis and saved the rest for progenitor assays and flow analysis (which I’ll describe below). We also harvested some liver tissue for histological analysis as well as the hind leg bones (for the bone marrow). We also isolated bone marrow cells for the progenitor assays and flow analysis.
After isolating the spleen cells and bone marrow cells, we had to count them … and count them … and count them some more. This is really the longest and the most tedious part of the process. We have to count them so we can plate equivalent numbers in the progenitor assays.
For the progenitor assays, we plate equivalent numbers of cells from each mouse (some treated with the drug, and the others treated with vehicle as a control) in solid gel that kind of looks like red Jello. The plates will have different doses of a growth factor called GM-CSF, which stimulates leukemia cells to grow faster than normal cells. We let these plates grow for a week before counting the number of colonies that are formed (which will be another long day). Our hope is that cells from the mice treated with the drug will grow less than those from the vehicle-treated mice, which would suggest that the drug was effective at killing the leukemia cells.
Finally, we used the remaining bone marrow and spleen cells for flow analysis. To do this we have to stain the cells with antibodies that recognize cell surface markers that characterize particular types of white blood cells. We are particularly interested in macrophages and neutrophils, which are increased in the leukemia we study. The antibodies are conjugated to fluorescent molecules that can be detected as different colors when stimulated with a laser in a flow analysis machine. This allows us to count the numbers of each type of cell. We hope that the drug-treated mice will have lower numbers of macrophages and neutrophils, indicating that their leukemia was reduced by the drug.
It was a long day, but so far the data looks promising. We of course have to repeat the experiment, which means there will be many more long days in the future.
Cheers!
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First of all I never thought for a second that someone with an undergrad degree in Linguistics would venture in to medicine. You just gave me hope… at least a little. Also I just learnt about physician scientist by reading your blog. So thanks for enlightening me today. Hope your days are fun so you don’t see how long they are.
Comment by Yaa — April 14, 2011 @ 2:21 am
Thanks for your comment. I feel very fortunate the MD/PhD program “took a chance” on a Linguistics major like me, and I’ve tried to work hard and make the most of the opportunity. I will say this though: a strong background in humanities is very important for developing effective communications skills in general and strong writing ability in particular, which is essential for the practice of medicine, and which I think has become greatly acknoweledged at this point. It is also very important for a career in research, which I don’t think receives enough emphasis or appreciation. In research, you can have great ideas and insights into the molecular mechanisms of disease, but it is all for nothing if you cannot communicate these ideas clearly and succinctly in the form of manuscripts and grants.
Good luck with your future endeavours!
Comment by Charles Goodwin — April 14, 2011 @ 8:11 am
Well said Charles.
Comment by Indianapolis Jiu Jitsu — September 17, 2011 @ 8:37 pm