Week 6 Mendelsohn lab

Hi again. This is a summary of what I did during my sixth week at the Mendelsohn lab.

I briefly worked on Sol's silk bladder augmentation project by paraffin sectioning some of his blocks. I found these paraffin blocks much more difficult to cut because the silk embedded into the bladder tissue made it harder to fully slice without it ripping apart. 

Throughout my stay at the lab, I have been paraffin sectioning many mouse embryos, not fully understanding where and how these embryos have been embedded into the paraffin wax. During the week, Katya brought an E17 pregnant mouse into the lab to remove its embryos. She removed all 13 embryos from the mouse and placed them in 1x PBS, while I prepared 13 tubes of diluted formaldehyde fixing agent. Katya showed me the following steps to dissect a mouse embryo and told me to dissect the rest of the twelve embryos. First, I removed the amniotic sac and cut a small piece of its tail to be further genotyped through PCR. Then I bisected the embryo, under the arms, removing the upper half of the embryo (because we are only looking at the lower half). After bisecting, I moved under the microscope to clean out the rest of the embryo removing everything but the bladder and kidneys. After removing the existing limbs and tail, I placed the embryos into each of the 13 formaldehyde tubes to be fixed and eventually paraffin blocked at the histology department for future sectioning. 

Mouse embryo E17

                          

                                  Dissecting microscope

Under the microscope

The next day we ran a PCR on the small pieces of embryo tail (that I previously mentioned) and the following day we ran a gel to confirm the Cre genotyping of the embryos to see which had the gene and which didn't because eventually we want to cross mouses that have Cre with mouses that have Apaf mutations. 

Gel

If you have been reading, I haven't explained what this project has been really about. Basically, the Apaf project (the one involving mouse embryos) is about the connections between the ureter and the bladder. In embryos, the ureters are joined in the nephric duct through the common nephric duct. Normally, the ureters would detach from the nephric duct and fuse with the bladder epithelium. This project is trying to analyze Apaf (Apoptotic protease activating factor, one of the major proteins that form the apoptotic regulatory network) mutants to determine whether apoptosis is required for ureter insertion.