Matt Erman, #4, Off to San Francisco

Since this is my last week out in California, and not much has changed since the last blog post (I am still screening), I decided to do something different for this blog; since  I am going into San Francisco for the day, I thought it would be a good idea to write about what I am doing at different points in the day. Enjoy!

12:29
First stop in San Fran is at this Burrito place in the Mission Neighborhood. It's supposed to have the best burritos in the city, and I would have to agree. It's amazing. Easily the best burrito I have ever had. Chopotle didn't have a chance.

1:36
Currently sitting under a tree in Union Square drinking a lemonade. It's weird how that right in the middle of the city there is this little enclave of nature. Might just chill here for a while it maybe go and shop a bit. There are a lot of stores surrounding the park, ranging from the extremely fancy ones with armed guards outside their doors to some that don't even have doors.

2:20
Finally made it over to the bay.  Decided to take a little break, so I'm sitting on a dock on the bay, watching the tide roll away.
I was talking to my mom earlier and she told me that today in New Jersey it was 90+ with terrible humidity. That must suck, since right now it's 78 with nearly no humidity and a cool breeze. If there is one thing that I deffenitly don't miss from New Jersey it's the weather.

A really pretty panorama of the city.

3:00
Ended up at the extremely busy and extremely touristy pier 39. It is kind of like a mix of a boardwalk and times square. Best part of the place had to be the sea lions that sit on docks next to the pier. There weren't too many, but that's because most of the sea lions go north to Alaska to the summer (I totally am a sea lion expert and didn't just read a sign).

3:50
Currently sitting again by the bay, this time with a great view of both Alcatraz and the Golden Gate Bridge. I find it strange how people were ok with a maximum security prison only a mile off the shores of a major city. I guess people just assumed that it was impossible to escape the island. I would have gone to visit Alcatraz, but a sign near the Alcatraz pier said that the next available tour was for August 16. I don't feel like waiting that long.

Look! It's the Golden Gate Bridge! It's so Gold(ish).

4:30
Made it over to Lombard street, AKA the crookedest street in America. It was a pain to get up here, as the hill is so steep that instead of walking up a road and being at a 45 degree angle, the city had to install stairs so it is easier to get up the hill. Once I got the the crooked part, I realized the street was less of a street and more of an amusement park ride, as there was a line of cars stretching for a few blocks, waiting to be able to drive down the street.

Look at how crooked it is! And the view of the city is really nice too.

5:30
It's time for my visit to San Francisco to come to a close. I saw everything that I wanted to see, except for the Full House house, but it is too far away from anything else to get there. All in all, I really enjoyed my day in San Francisco.

Bridgid, Entry #4, The Doctor is (F)in

Hi everyone!

Hope that everyone’s summer in the lab is going well! This past week (the week of the 11^th, sorry by last entry was a bit delayed) was my fourth week in the lab. I can’t believe that I am already more or about half way done my time with my lab, the days are flying by! This week was slightly more of a down week in the lab, running the standard procedures for new groups of embryos. (It was also a shorten week for me because we had a last minute family issue that I needed to care of, which required a shortened day on Wednesday and off on Thursday.)

This week on Monday, I started out the week with another day of imaging.  This time, however, I imaged the 24 hpf embryos.  It took a large part of the day, but I ended up recovering 20/25 embryos from the in situ for imaging.  Since the 24hpf embryos are more developed and have more of a defined figure (aka look more like an actual fish), they were in my opinion, much easier and much more interesting to image than the 18 hpf.  I honestly don’t know why, but personally I guess it made a different when they were more developed, maybe because they looked prettier (and more recognizable) in the pictures, but also because they were much easier to position.  After I finished imaging in the afternoon, I started the first part of the DNA extraction for the other half (#9-19) of the 18 hpf embryos. Because the third step in the procedure for the DNA extraction is to incubate the embryos over night in the water bath, I wanted to make sure that I started today, so that I would have the DNA ready tomorrow. 

On Tuesday morning, I came in and got right to work on the rest of the DNA extraction procedure for the last half of the 18hpf embryos.  Once I had extracted the DNA, I measured the DNA on the Nanovue.  Because I finished a little earlier than expected, I was able to go with Abby and Patrizia to the Influx machine at Penn. The Influx is very similar to the cell sorter (JACS) that we used last week, but it sorts the cells more gently, so that hopefully we will have cells on the slides after the cytosine and fixes the problem that we had last week.  Having never used the Influx before, we just used a control group for the cell line, lmo2, that we were sorting, to make sure that it gave us cells at the end that were visible before we ran the actual experiment.  After we took the cells sorted from the Influx to the cytospin, we were so happy to see that there were beautiful, clear cells on each of the slides!!! I spent part of the afternoon with Patrizia looking at the slides and the morphology of the cells.  It was awesome to see so many different types of blood cell lineages at different stages of maturation under the microscope.  I honestly could have spent hours looking and identifying all the cells, I thought it was that interesting.  Also, it was so great to know that the Influx worked to sort the cells and gave us the results that Patrizia wanted! In addition to imaging the slides, I also ran the PCR for the samples that I did the DNA extraction for that morning so that they would be ready for sequencing this week. 

Because of a family issue, I only came in Wednesday for a few hours before heading home.  In that time, I ran a gel with my PCR from yesterday and then started to cut some of the bands from the gel for purification.  Patrizia said that she would happily finish anything that I didn’t get to so that the PCR could be sent for sequencing.  Unfortunately, when Patrizia went to finish cutting the gel, she noticed that the marker and some of the PCR products had faded and that the sequencing would most likely not go well.  Since the results from the first 8 samples came back with an unreadable sequences anyway, we decided that it would just be better to re-run all the PCR for all 19 samples of the lmo2 18hpf DNA.

All day Friday was focused on re-running the PCR for all 19 samples of the lmo2 18hpf and then running the samples on a gel to prepare for sequencing. Also, because something might have been weird with the gel that I prepared (might have been too cold or hot when I added the ethidium bromide), Patrizia wanted me to watch her prepare the gel to see if I noticed anything different.  On a different note, this is the first time that I have mentioned it, but because CHOP has so many summer students in labs across the campus, they have put together a really nice seminar series that features all types of doctors from all different departments at CHOP. Although most of the kids who attend the lectures are in college, I am lucky enough to attend the weekly sessions on Fridays with Abby.  Each seminar runs about an hour long, with each doctor talking a little bit about their field and then talking a little bit about their journey that led to who they are now, along with giving career advice. The talks are always so informative and it is really cool to get a background in all different types cancer research that are going on at CHOP and the innovative techniques that so many of these doctors are creating and using to make huge strides in the cancer community.  The career advice is also extremely valuable, especially for someone who is only in high school! That afternoon, I helped Abby with some imaging of a specific transcription factor, cmyb, in the MLL-GAS7 fish.  Patrizia and I also prepared a gel to run my PCR, but unfortunately when I went to take out the gel from the model, it ripped. Because it was almost time for the lab meeting and we wouldn’t get the sequences until Tuesday regardless of if we ran the gel today or Monday, we decided that it would be better to not rush and finishing running the gel on Monday.  Friday finished up with a lab meeting, where I got to learn more about the research that the other post docs in my lab are conducting and Abby and I also did a short presentation about our research this week!

Soo, Entry #4: Last Week

Since it is the last week of my internship in Dr. Bassett's lab, I am finishing up my works and re-organizing the results so that other people can later on easily understand my work. 

I first went back to working with Gephi and generated brain network images with different modularity values. Small modularity value detects many small communities, whereas greater modularity value senses larger groups. The figures below are images of same subject and scan, but they have different modularity.

(modularity = 0.2)

(modularity = 1)

(modularity = 5)

After finalizing the gephi images, I revised my MatLab code so that I can save the values that I've used to generate the boxplots. I think the most important task that I would have to complete before I leave is to make sure that all results and data that I've worked with are saved and accessible. 

Azza Entry #2 - The Other Side of the Lab

Last week I had the opportunity of shadowing my PI, Dr. Rahul Deo, in the clinic to observe how he interacts with his patients, and the type of concerns his patients bring to him. Yesterday we had a female patient in her mid-sixties. She came to UCSF clinic seeking answers concerning her cardiovascular state. Her older brother passed away a couple months after being diagnoses with hypertrophic cardiomyopathy. Hypertrophic cardiomyopathy causes thickening of the heart muscle linings making difficult for the heart to pump blood as easily. In turn, making the heart significantly weaker. It turned out that most of her family has died from heart failure and a majority of them being from hypertrophic cardiomyopathy. Not knowing whether her relatives died from inheriting a heart disease or simply due to age and higher blood pressure. Dr. Deo has to gather genetic testings’ her relatives have done in Canada before being able to tell whether this patient has also inherited a mutation for heart failure. After meeting with the patient Dr. Deo showed me an ultra sound of the patients’ hearts and pointed out to me the different chambers of the heart and also showed me some thickening of the patient’s atrium, but he figured the thickening could have been due to high blood pressure and age.  He also showed me an EKG which is a electrocardiogram of the patient and showed that there were no obvious reason that would validate the patient of having a mutation. This was really interesting to me because not only have I been able to work in the lab, but I also got to see how Dr. Deo interacts with patients, and how patients like that influence future questions and concerns in the world of cardiology. 

Katie Entry #3: Finishing Growth Curves

Last week was my fourth week in the Cheung Lab.  It was pretty similar to the previous week because I was finishing monitoring the growth of the potentially cold-sensitive mutant MRSA strains in optimal and cold conditions.  I continued to observe the optical densities of each strain using a spectrophotometer, making growth curves using the ODs, and finding/comparing the growth rates of each strain at different temperatures .  However, this week the postdoc that I have been working with, Dhana, found a better equation for determining the growth rate of cells using ODs so I went through  my past and present work using that equation to find the doubling time of each culture and making new growth curves.

Also, some of the agar plates from the primary screening had issues growing in the incubator because of an issue with air flow on the top shelf so the agar on the plate ended up shrinking and becoming contaminated.  I re-picked each strain on agar plates to grow at this temperature to finish the primary screening. 

After I finished picking colonies and growing/observing the mutant strains in liquid media, I made a master graph to compare the growth of the 50 strains  at each temperature and to pick the most important strains (the ones that grew the slowest in cold conditions) to use during transduction.  We are going to start transduction next week using two strains so I went over the protocol and streaked the two strains that exhibited the most cold-sensitive phenotype on a plate to grow over the weekend.

Emma, Blog #4, Weeks 5 and 6. WARNING GRAPHIC IMAGES

The past two weeks have really picked up in terms of what I have been doing in the lab.

Week 5 in the lab saw four big developments:
1. Finishing NeuN immunochemistry and moving to GFAP: NeuN immunochemistry is where you stain mice brains (at sizes of 50 micrometers) to view neuron channels under a microscope. Then, my mentor Yuling can run analyses on the brains in order to compare the amount of neurons between wild-type and genetically modified mice. I have been doing this type of immunochemistry for the past two weeks, but starting next week I am moving onto GFAP, which will stain brain slices to view glial filaments. The process between NeuN and GFAP immunochemistry is almost identical, however you use different secondary antibodies that bind to the primary.
2. Cutting brain slices: In order to start running GFAP, we need brain slices to stain. My mentor taught me how to use the machine that slices the brain slices to your desired thickness (which in our case is 50 micrometers). I spent about two hours cutting over 100 slices from one brain, which we then froze in a -80C freezer until we start the immuno next week.

3. Performing perfusion on three mice: Whole animal perfusion is a way to kill an animal and then fix it with paraformaldehyde to obtain the best preservation of the brain. This is an important process, because it needs to be done correctly in order to obtain brains that are preserved well and can be worked with either for staining or western blots. To perfuse a mouse, you first sedate it with ketamine threw a intramuscular injection. Once the mouse is fully under anesthesia, you pin it down stomach up in order to cut into the stomach. After cutting through the stomach you go up through the sternum to expose the heart. You inject a butterfly needle into the heart and pump in saline, and then quickly cut the heart to kill the animal. The saline is taken up through the capillaries and flushes out all of the blood. Once the animals lungs are white, and there is no longer any blood left in its system, you switch out the saline for paraformaldehyde, which is a fixative that preserves the brain instantly.
Here are some pictures from the procedure.

4. Joining a behavioral studies project: At lab meeting, my PI Dr. Siegel announced that he needed people to help in a behavioral study that tested Dextras1's effect on memory, EEG readings, and locomotion. I am specifically working on running 30 mice through a T Maze, a paradigm that tests working memory. I run the mice through 8 trials and then analyze the results.

Week 2

This week brought about another new project, as well as the start of my behavioral testing,

1. GFAP: This week I ran immuno for GFAP on brain's, and it went very well. My slices were all in tact with no breakage, which gave my PI the confidence to give me a project....
2. PROJECT: I got a project I am working on with one other high school student Meghan for the duration of my stay. I am using GFAP immuno staining to analyze if ketamine exposure in adolescent mice produces any differences in glial filaments when compared to wild mice. This may not seem related to schizophrenia, the labs main focus, however schizophrenia is a NMDA antagonist and is thought to have a correlation to the disease. I will start immuno next week for this project.
3. Behavioral testing: I ran my first trials on all 30 mice this week in the T maze. It took me the full day, but it was actually pretty fun. This was the first time I got to interact with the mice completely alone, which made me nervous at first but in the end it was fine. Next week I will have two more testing days before I can start analysis.
3. Clinical trials with my PI: Dr. Siegel not only runs a lab, but he meets with patients every Wednesday afternoon at UPenn's hospital. I was able to shadow him for the whole afternoon. Unfortunately, I cannot divulge much information because of HIPAA, but it was a great experience and I got to actually meet people dealing with schizophrenia. It was a really moving experience and made me glad that I was working in a lab that was trying to help people like them live a normal life again.

Jay Ha #2 growing e coli and collecting data

This week had been full of bacteria. 

Now that I am sitting in front of my laptop trying to get my weekly journal stuff done, I wish I had taken more photos. I guess I will take more photos next week. 

This is one of the few photos I took before autoclaving these bacteria and saying goodbye to my weekly production of lovely bacteria . As you can see, there are many plates, and all of these plates contain somewhere between 0 to tens of thousands of colonies of E Coli bacteria that I had grown. YaY!

My lab works started with production of more phosphate buffer solution and LB Broth. It wasnt that hard as I had to put some pills or powdery materials into the beaker using precise scientific scales and mixing them with DI water and shake them and put them in sonic machine that evenly mixes the solution. All solutions prepared had to be autoclaved to create a sterile environment. The phosphate buffer solution would be used in serial dilution method of E Coli culture to prepare E Coli of varying concentration. Some of the LB Broth, or the source of food for bacteria, are used to directly culture E Coli population in a shaking incubator set to 37.5 degrees Celcius, and other portions of LB Broth are mixed with solidifying agent so that they would be prepared to plates as seen in the picture above for the grown E Coli cultures to be tested for their growth pattern. 

Plates were prepared under bio safe fume hood. Making forty plates is tedious but does not take that long time. When the E Coli culture that used to be frozen is defrozen and incubated and all prepared, they are serially diluted to the factors of 1, 10^-2, 10^-4, ....., 10^-8. This is done by adding 0.1 mL of E Coli culture or serially diluted cultures into 9.9 mL of phosphate buffer. 

Then, these serially diluted solutions containing E Coli are spread onto the plates using glassware that spreads the culture onto the plates. Five sets of plates are prepared on every two hour intervals for 20 hours and put in the incubator to be counted in next 24 hours. After 24 hour intervals these plates are counted for the number of colonies of bacteria that had grown overnight. 

OD measurements using spectroscopy machine are taken in varying time intervals to see the correlation between OD measurements and CFU. 

I have prepared and counted the bacteria for five sets of two hours intervals everyday and had conducted this for four days total. 

First sets of data were discarded because bacteria were poorly cultured because the incubator was partially broken, so the data taken from third and fourth days are below. 

Time (h)	OD at 600nm	Count	Dilution Factor	CFU/mL
0	-0.0048	335	1	3.35E+03
2	-0.0018	301	1	3.01E+03
4	-0.0063	512	1	5.12E+03
6	-0.0223	45	100	45000
8	-0.0021	309	100	309000
10	0.0123	61	10000	6.10E+06
12	0.771	165	1000000	1650000000
13	0.8125
14	0.4506	290	1000000	2900000000
15	0.8439
16	0.9864	161	1000000	1610000000
17	1.0974
18	1.1149	301	1000000	3010000000
19	1.196
20	1.208	261	1000000	2610000000

Here are three graphs from data collected this week. 

Next week, I will be conducting research that deals with disinfection that had been catalysed through oxidation processes. Sounds fun. lol