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Blog Archive: June 2003
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June 30, 2003 - restriction enzyme digest plasmid analysis; refreezing the HeLa cells
-4:23 pm: Today we did the final analysis of our Qiagen Maxi isolated plasmid DNA by doing a double digest with HindIII and Not1 endonucleases. We haven't presented the results to the professor yet, but our own rudimentary understanding of the gel seems to indicate that we did get some useful plasmids and that the DNA itself was decently pure! In any case, that work took about 4 hours to do.... After that i decided to put my HeLa cell line back into cold storage, just for practice.
-tomorrow we are going to begin anew with plasmid isolation... this time using M9 (aka minimal) media to shock the bacteria into making lots of plasmid.. Hopefully we get a better yield than with LB


June 29, 2003 - the weather reverts to ideal
-4:26 pm: Yes! The weather has returned to just about normal, with highs in the 70s. The extended forecast predicts similar weather for at least two weeks. So happy!
-got up today at 7am... i suppose body used to getting up in time for 8am class... how unfortunate. Went to RSF for mini-workout (bike and b-ball). Decent shooting day... although my lay-up skills were on the shitty side. I returned home to eat brunch... then took a nap until just about an hour ago! Yay...
-promising news: three militant Palestinians groups (Hamas, Islamic Jihad and Fatah/Al Aqsa Matyr Brigades) agreed today to halt attacks on Israel today for three months, in exchange for Israeli military withdrawal from the Gaza Stip. Such news comes as a surprise to me, but it is a very pleasant surprise. At least now both sides seem willing to try peace. I really hope that neither side decides to ruin this by launching attacks, but i have some strong feelings that one side may falter.


June 28, 2003 - almost 7 straight hours of Halo
-11:07 pm: Weeeeeee.. i went over to Yeh's to play some co-op Halo... Some fear involved since i'm don't play FPS, but it was fun anyway. The controls take some getting used to, especially if you're used to Metroid Prime controls (and vice versa, many ppl also complain that Metroid Prime's controls suck and take getting used to)... The only thing i really didn't like was jumping control. Control misgivings aside, the game itself is very fun, especially controlling various vehicles. As the title says, we played co-op mode for almost 7 hours, getting through about 60% of the game. By the end of the 6th level if was feeling way too dizzy to continue.. i would have gone on, but alas damn dizziness. I hope to play through to the end, mainly because the storyline is particularly interesting and i wanna know what happens.
-The heat wasn't too bad today, in fact walking home from Yeh's around 9, it was actually kinda cold and i shivered a bit. Hopefully the weather stabilizes into a nice constant temperature in the low 70s.


June 27, 2003 - Analysis of plasmid DNA
-9:00 pm: Today was a relatively short day in lab, probably because we knew exactly what we were going to do... and we didn't have to wait around for someone to tell us. Basically we analyzed the samples we got from yesterday. Mainly some spectrophotometry and agarose gel electrophoresis. The results? Spectrophotometry says that we got some DNA with some protein contamination. Electrophoresis confirms the isolation of DNA. Yay! we got DNA... BUT we will not know if it is the correct DNA, until we run some more electrophoresis with restriction enzyme digested samples..
-Even though we did get some DNA, the prof. wasn't satisfied with the amount. She suspects that the wrong media was used mass-propagate the bacteria (or actually the plasmid). Next week we are going to repeat the whole process, using M9 media instead of LB...  At times the process is boring, but this process is the foundation of molecular biology!


June 26, 2003 - plasmid isolation
-9:12 pm: We spent a long time today watching stuff drip in the attempt to isolate plasmid DNA from our cultures. Basically spent the entire day doing that.. hopefully tomorrow we will find out if we actually got any DNA... and whether or not those pellets we saw were actually DNA.
-What a drastic change in the weather!! It was really cold last week... and now it's blazing hot. Freakin last week we had temperatures in the high 60s... today's high was 95! VERY uncomfortable outside...


June 25, 2003 - a long, productive day
-11:26 pm: the first week of IB131 (human anatomy) is over. The class has been so far, as much as expected... long lists of things to memorize. Prof. Gilson also likes the lecture at a very fast pace, perhaps on par with Prof Schekman's pace... Wow... at least human anatomy is as terribly complicated as vesicle trafficking or whatever.
-IB131 begins at 8am, however i went to lab in LSA at 7:30 am to take my transformed bacteria LB plates out of the incubator. Succesful transformation! Actual colonies and the negative control had no colonies, meaning the plates were properly made. After that i headed off to IB131 to learn about the bones of the skull and the vertebral column
-After IB131 headed promptly back to LSA. Prof. informs us how to begin a "starter" culture. The starter culture is a mini 5mL culture of LB to grow up a lot of bacteria from a single colony. We autoclave the appropriate containers and proceed with the starter culture in the "shake" room. We complete the work at about 12:45 pm... the culture needed to go for 8 hrs
-Ate lunch from newly opened Quizno's. Pretty good sandwich although a bit on the pricy side. The toasted aspect was new to me... seems to deliver what it promises in the commercials.
-at lunches end, i decide to make some more LB broth to help out another student in the lab. Another 2L of the stinky stuff. 2 hr lag time before anything else productive happens. At 3:45, we are instructed about the main bacteria culture, a large 1 L culture used to make 1 g of plasmid. We were supposed to use the LB we made on Monday, however the LB became turbid... meaning it was contaminated with bacteria!!! UGH... Fortunately the other student did not need the LB i made earlier in the day, and we decide to use that.instead. Some more autoclaving of appropriate containers. At that point it was 5:00 pm... and 3 hrs, 45 min before the starter culture would be finished.
-I rush home to change into "RSF" clothes to join my lab partners for workout. We shoot hoops and play a game of 2on2  for about 1.5 hrs, after which we decide to try our hands at Ping-pong... Ah good old ping pong from the days in Freeborn Hall. The two years that have elapsed since then showed.. and i was terrible at it. I did win the HORSE game while we played basketball... but oh well.
After only about 10 minutes of ping-pong we move onto bikes. A pleasant surprise during RSF was running into three good old friends. Saw Andre and Nam from Hoover High school and Salman from freshman year Freeborn Hall. I hadn't seen them for so long! It was definitely nice seeing them again!
-By the time we were all pooped out from the RSF, it was already 7:45, so i decide to head back to LSA to start up the 1L cultures. More autoclaving... final preparation of LB (injection of appropriate antibiotics) and finally pouring of the 5mL starter cultures into the main cultures!! Yay! After some difficulty with the shakers (the damn things weren't set to the right temperature) i finally finish the day at 9:30 pm.
-Ever since i started working in the lab.. my life has been very, very busy... Very stark contrast to the boredom i endured the first week after school ended. Being busy has definitely been much more satisfying to me though... i can't wait to see if our bacteria cultures turn out correctly tomorrow!


June 24, 2003 - more LB-agar plates
-6:20 pm: as the title implies... made more LB-agar plates today. fun, especially popping bubbles with the flame. Also transformed some bacteria and plated them on our plates from yesterday.... Have to come into lab at 7:30 am tomorrow to take them out of the incubator.  Although much more happened today, i'm about to step out, so cutting entry short here :p


June 23, 2003 - the weekend and making LB-Agar plates
-5:54 pm: My brief time in the Los Angeles area was very fun. Good food, playing basketball, watching/playing videogames, etc.. nothing could be better. Brother got nearly $1700 for graduating from HS... lucky him!  I got back to Berkeley around 4:30 yesterday, left at 12:00 pm on friday. Damn SF-Oakland baseball game made the Colliseum BART station extremely crowded yesterday... imagine 10 car BART trains completely full - standing room only. Ugh.
-Lab on friday was frankly pointless for me.. didn't anything but received some more literature relevant to projects
-today we began to prepare for bacteria experiments. First we made 2 L of Luria Bertani (LB) broth. Then we made some LB-enriched agar plates (those pale-yellow plates from Bio1A). Pretty fun making the plates themselves.. but making the broth was a pretty stinky affair (damn autolyzed yeast extracts). Tomorrow we are going to grow some bacteria cuz the lab is out of a particular plasmid.


June 19, 2003 - the centrifuge of death
-7:37 pm:  wow. today was probably the scariest day of lab ever.  We were preparing some delipidated FCS which required the use of a centrifuge to separate liquid phases. So we set everything up, made sure the the centrifuge was balanced, and started up the centrifuge. A little shaking at 800 RPM, some more at 2100 RPM... the centrifuge is stopped just make sure everything is OK... Stuff seems to check out alright. We put the solution back in and start up. The same shaking at 800 and 2100... we decide to go on. No shaking and the centrifuge reaches max at 3500. All green.... for 5 seconds... a sudden pop noise. We needed to check what the noise was... As the centrifuge slows down, it begins to violently shake. I thought it was going to blow.. people in the lab start arguing over how to best handle the situation... The centrifuge mercifully stops and we open it up to inspect the damage. The glass containing the liquid is completely shattered. I mean really shattered, it looked like fine grain sand. After much talk and clean up... we decide the culprit was probably leaving the stir bar inside...
-The most interesting thing about the experience with the centrifuge was that that centrifuge was a low speed one... The professor told us about how she's see ones that walked around when they were unbalanced. Yikes! Hopefully we have no centrifuge disasters in the future.
-A NOTE TO MY LOYAL READERS (YOU RULE!!) This blog won't be updated again until Monday as i'm going home to LA this weekend (it's my bro's HS graduation party!!)


June 18, 2003 - another looong day
-9:02 pm: the days in lab are starting to get really long. I came around 10:15 and didn't leave until 8pm. Wow. Yesterday i had the impression that we were only going to watch how to do immunostaining... however we actually did the immunostaining all the way from fixation to mounting the coverslips onto slides. Probably the most frustrating part was micropipetting 1 microliter of antibody solution. I became really frustrated with the P10 micropipetman; the stupid thing didn't seem to like me. OH well... we didn't get to look at our immuno stains today, but we will be doing that first thing tomorrow.
-Because we were in lab so long today, we did not get to go play basketball at the RSF as we had planned yesterday.. BOO!! I'm hoping we can go tomorrow... it probably won't be a long day, but who knows.


June 17, 2003 - watching more of the GAG chain assay
-9:19 pm: for mostly just watching the rest of the GAG chain assay, i spent quite a long time in lab today.. from about 10:45 to 7:15. Most of the time i was either watching someone else do experiments or chatting away. The GAG chain assay was finished up by transferring precipitated GAG chains to scintillation vessels. This actually takes a long time because it involves the filtration of liquid from 64 tubes! I also watched people prepare coverslips for cell binding and immunostaining. Tomorrow we're going to watch the immunostaining process as well as see our results from the GAG chain assay. I actually did some lab work today... but it was simple routine cell culture.
June 17, 2003 - the stupidity of Bush and Sharon
-12:08 am: i'm pretty tired of hearing all about the Bush and Sharon intentions to rid Israel/Palestine of terrorism by eliminating Hamas. What utter stupidity. What these idiots do not understand is that Hamas is much more than a terrorist group. Before you get me wrong, i do not condone their terrorist acts,  but you must consider what other things that Hamas does. Hamas provides the most basic of human services to many Palestinians including food, potable water, economic aid and schools. This is why their support base is so strong and why the general Palestinian public really likes Hamas. Targetting Hamas only enrages the Palestinian public; Israel is targetting basic services. The real culprit is the lack of Palestinian government. Government should be providing these basic services.. only then can Hamas be safely "eliminated." So at this point, attacking Hamas only makes things worse... an enraged Palestinian public can only be more willing to produce more suicide bombers.
-There are a few Republican Senators who in recent days have suggested that the U.S. military actively engage Hamas... I really hope that this does not happen. Not only would it makes things worse for both Palestinians and Israelis, but would only serve to further inflame deeply held Arab hatred for the U.S.


June 16, 2003 - HeLa cells
-9:49 pm: having nothing to do right while waiting for the washing machines to finish their cycle, i got a little curious about the HeLa cells that are used not only in the lab i'm in right now but all over the entire world. Apparently i was wrong about when the cell line was established... it was not established in the 1970s but was actually established in 1951. They were cervical cancer cells taken from a 31-year-old woman named Henrietta Lacks (hence the name HeLa) at Johns Hopkins University Hospital. Despite their importance in cancer research and general cell biology research, they are quite the controversial little cells. Apparently neither Henrietta Lacks nor her surviving family ever gave permission for her cells to be used for research (Henrietta Lacks died 8 months after her cells were taken, initially only to diagnose them as malignant). Her family found out about the use of her cells more than twenty years after her death, but by then the cells had already been distributed worldwide. And to this day, the family has not received one bit of monetary compensation. The controversy does not end with the lack of authorization or monetary compensation. After the establishment of the HeLa cell line, many other cell lines for other tissue cancers were also supposedly established. People began to do experiments on the properties of various tissues using the various cell lines. However, the HeLa cells were apparently too good at proliferating and could easily overtake a tissue culture contaminated with them. Due to lack of proper sterile technique, many of the "other" cell lines that had been established after HeLa, were not actually other cell types but additional HeLa cell lines. Thus there was much controversy in the 70s and 80s over all experiments that were done with the supposedly "other" cell lines, but were actually done with HeLa cells. Research on lung epithelial cells, intestine epithelial cells, etc. was all rendered fraudulent and the millions of dollars that were poured into those research experiments were wasted. Some scientists refused to acknowledge the obviously egregious errors. Journals took no corrective action, believing that the problem was too widespread (and thus, as one author i read suggested, could be ignored). I guess there are two lessons to be learned from the HeLa experience. One lesson was the ethics involved.. now permission is always sought to use tissue or cells for research. While that lesson has been learned.. the other lesson is one of sterile/aseptic techniques... we should all use them! I suppose i should cite where i got all this info.. since i really only paraphrased what someone else wrote: Masters, John R.  HeLa Cells 50 years on: the good, the bad and the ugly. Nature Reviews 2: 315-319. Online PDF text available at http://www.lifesci.texas.edu/research/huibregtselab/HeLae.pdf
June 16, 2003 - watching the pulse-chase experiment
-6:45 pm: today wasn't terribly exciting since all we could do was to watch how the experiments were done. Since the experiments involve the use of radioactive reagents, we are not allowed to handle them until we have been trained to do so (that will be sometime in july). Nevertheless it was interesting to see just how complex the experiment is, especially with all the controls that were necessary. When it comes time for us to actually do the experiments we'll probably have loads of fun incorporating all the techniques we've learned so far.... It was all there: proper micropipetting, sterile solution preparation, sterile cell culture.. just about everything except for gel electrophoresis. Tomorrow the experiments will be finished up by measuring radioactivity levels and we will see exactly what effect the level of cholesterol has on constitutive secretion


June 15, 2003 - the inevitable
-11:18 pm: so the Spurs finally won the NBA championship... woohoo... it was definitely the ugliest basketball series i've ever seen in my entire life. i don't think that the Spurs or the Nets ever broke 100 points and i think the Spurs breached 90 only once... dang. At least David Robinson gets to finish out his NBA career on the highest note possible.
-did some rearranging of the left side bar thingie on this webpage.. added links to other blogs. E-mail me to remove the link if you don't approve or if you want yours linked there too.
June 15, 2003 - why are Sundays always so unproductive?
-3:43 pm: Sundays seem to always be a complete waste of time for me... i never get anything done, although technically right now i really don't have anything to do (perhaps though i should be reviewing for that lovely GRE in september). So yeah after that Simpsons marathon last night, i proceeded to surf the web for a few more hours. Then Star Wars Episode II came up on HBO, and even though the movie is terrible (OMG that acting) i couldn't help but watch it... so i ended up going to sleep at 4 am. Fun. Subsequently i didn't get up until about 12:30 pm today... Woohoo... so a fansub of Gundam Seed episode 36 was released during my beauty sleep so i proceed to start the d/l. I eat the leftovers from last night (leftovers are always nasty...) while waiting for the d/l to finish. Impressively the download takes only an hour and a half (impressive in the sense that the file is almost 200 MB) and proceed to watch! Interesting episode about how one character catches up to the events of the last episode, although i didn't have any fun mecha action.
-The experiments we were going to do in lab today were moved to tomorrow (i guess ppl didn't want to come in on the weekend which is understandable), so nothing to report about that.
June 15, 2003 - 6 hrs of The Simpsons
-12:04 am: Wow.. just finished watching 6 consecutive hours of The Simpsons. I think i don't want to hear that theme song for at least a week now. Besides that unproductivity, the rest of the day was pretty much the same.. doing jack shit... It'll end by next week though, as summer school starts up.


June 13, 2003 - Short Day
-3:02 pm: Yay! my culture did not have any bacterial contamination! We had a really short day in lab today as all we did was to split the culture into two dishes for experiments later on. I'm coming into lab on Sunday to begin the work on the cholesterol experiments.  As lab was short today... such will be the length of this entry.... :P


June 12, 2003 - Sterile Tissue Culture and Reviving Long Dormant Cells
-4:34 pm: After learning about all the techniques dealing with dead stuff (gels and the like) we moved on today to handling live cells!! We primarily learned how to properly thaw frozen cells and how to transfer them to appropriate growth medium all under sterile conditions. After observing the veteran student prepare a cell culture we each had to perform the same procedure. We will see tomorrow if we were successful in preventing microbial contamination. The type of cells we transferred from cryopreservation to growth medium were HeLa cells, a cancer cell line that has been in existence since the 1970s. The cells were cryopreserved at -196 degrees C (77 K, the boiling point of liquid nitrogen) in 10% DMSO (dimethyl sulfoxide). The cells that we used have been frozen for almost 8 years! After quickly thawing them, the cells were transferred to DMEM (Delbecco's modified Eagle's Medium) containing 4.5% glucose. Being sterile about it the whole time proved very interesting... nothing was allowed to touch anything else etc etc. I was pretty nervous about being sterile because one mistake meant the entire hood would be contiminated at great inconvenience to the lab. I don't think i contaminated anything... but we'll see tomorrow.


June 11, 2003 - Finishing the western; Cholesterol
-3:59 pm: We began today rather late (around 12). We finished off the western by staining our nitrocellulose papers with a red dye, revealing the transferred BSA. After discussing the results we moved on to learning about what experiments we are going to do to learn tissue culture. We are specifically going to be studying whether or not cholesterol levels play a role in vesicle formation from the trans golgi. It's a very cool experiment to run because it will teach us many techniques including sterile procedures and some pulse chase. The pulse chase portion is actually something called the GAG chain assay and it involves measuring the radioactivity from radiolabeled GAGs (glycosaminoglycans). Basically GAGs are produced in the trans-golgi. We can created GAG enriched trans-golgi and vesicles by adding a membrane-permeant xylose analog which initiates synthesis. If we pulse radioactive sulfate into the cells, a pulse of radioactive GAGs is produced. You can move the pulse along the pathway by adding non-radioactive sulfate (the chase). So using this GAG chain assay and varying the cholesterol levels we can determine just how much of a role that cholesterol plays in vesicle formation. Today we were told all about this and the ideas behind it, and we get our official start tomorrow.
-The weather here has been disturbingly cold. I've not seen the sun in a long time and wondering when the heck actual summer weather is going to arrive. It feels like January outside; although at least it is not raining. hmm weather.com projects mid 70s for the weekend... but they also projected that for the past few days.... damn you weather, damn you!


June 10, 2003 - Western Blot
-3:47 pm: They tend to describe many experiments in MCB classes and a very common one is Western blotting. They make it sound so simple... it's always "so you just do a Western blot of this" and blah blah... etc. What they don't say is that the blotting itself is actually done eletrically (so it's not actually blotting per se).. and it involves the construction of a weird sandwich of filter paper, polyacrylamide gel, nitrocellulose paper, and more filter paper. Nothing terribly tricky about actually constructing the sandwich, although my freakin gel ripped (a small unimportant corner of it). I'm also wondering if i cut out a big enough piece of nitrocellulose paper... we will find out tomorrow when we see the results and stain the paper. So yeah since we're not actually doing any experiments yet; we can't probe our paper with antibodies for a few reasons: 1) the blot was mainly practice, 2) antibodies are expensive, and 3) BSA is too uninteresting.
-Finished watching the first two seasons of Gundam Seed (Episodes 1-26). It's a very interesting anime with some seriously screwed up characters who all seem to like to use each other for their own means. It's definitely better than my first Gundam series (which also had some really messed up characters and one really annoying Relena Peacecraft... erggg), Gundam Wing, and i look forward to watching the third season and on. Miss Relena does have a counter-part in Gundam Seed (in the extremely annoying sense), the revenge-obsessed Frey Alstar.


June 9, 2003 - DNA (agarose) gel
-7:26 pm: Today's lab training was fairly straightforward. First we made our second set of polyacrylamide gel and it was definitely much easier the second time around. After we were done with that and our gels stored properly, we moved onto agarose gels. The method used to make agarose gels is far simpler than that for polyacrylamide gels; it simply involves microwaving a solution of agarose containing ethidium bromide and pipetting some onto a glass plate with the plastic comb. While the gels were setting, we made our DNA solutions, then we micropipetted the solutions into the wells created by the comb. Then it was simply putting the gels into the electrophoresis box and turning the switch on (well not THAT simple.. ). While waiting for the gels to run, we were shown how to properly clean just about everything and anything in the lab. Glass beakers, plastic cylinder, micropipette tips... etc. We were even taught how to make what i named "super-clean" water. Tomorrow we will run some more SDS-PAGE and perform a Western blot.
-Yesterday I journied to the newly opened Trader Joe's at the El Cerrito Plaza. A $26.86 well spent on this week's food. I'm fairly happy that they've opened a store that i can reach without having a car.. although having to BART there limits the amount i can buy.. oh well.


June 7, 2003 - Return to old habits
-5:30 pm: After watching 12 episodes of Mobile Suit Gundam Seed and Scrapped Princess it was inevitable that i would return to 1st and 2nd year of college habit of always listening to my small collection of anime themes/J-pop...oh my... of course the catchy themes from both series have been added... i tend to like "Invoke" (MS Gundam Seed) more just cuz it has no English in it unlike "Little Wing" (SP). As for the anime series themselves it's kind of a toss up which one's better. They're in totally different genres so it's probably unfair to compare them. Scrapped Princess's storyline is more original though, as Gundam Seed's plot tends to be along the lines of older MS Gundam series. Gundam Seed, being Gundam, definitely has more action and the animation itself seems to be of higher quality, but i think the characters of Scrapped Princess are far more interesting (although Kira Yamato from Gundam Seed is a real head-case).


June 6, 2003 - Running SDS-PAGE
-6:18 pm: SDS-PAGE is a nice protein separating technique that is often described in many biology courses as extremely useful. Today we got to run our samples of BSA through the gels we made yesterday, and though it was pretty challenging and fun, it involved quite a bit of waiting. The first step was to take the gels and assemble the electrophoresis device. Making a device that did not leak was pretty difficult and took quite a bit of effort. After the devices were all set-up and filled with running buffer we loaded the protein samples into the lanes (also a pretty difficult step). The devices were turned on and we had to wait two hours before proceeding to the staining step. After the two hours, (we went out to eat lunch and snooze around some shops), we came back to disassemble the devices and transfer the gels to staining dishes. Removing the gels from the glass plates was fairly nerve-wracking. The gels are extremely thin and so the fear of ripping them was quite high. After transferring to the dish, the gels were stained with Coomassie Brilliant Blue (CBB) for 2 hours.(we played Hangman to kill the time.... sigh). After the two hours were mercifully up, the CBB was removed and destaining solution added. Tomorrow we are going to dry our gels and see how our BSA samples turned out. I personally can't wait to see if my gel turned out correctly! On Monday we are going to make more gels (practice makes perfect!) and that time we will transfer protein to nitrocellulose paper instead of staining. I feel so lucky to be able to learn all of these techniques. We've learned in just one week what is normally taught in MCB130L in four. And plus we've had to prepare our own solutions, which is usually not the case in course lab work.  I really wish i had tried all of this much earlier in my college career, but i suppose it's better late than never...
June 6, 2003 - The past
-12:09 am - my brother 's last days in high school are zooming by right now. He will officially graduate on the 18th.  I warned him yesterday on AIM to enjoy these finals moments with his friends, for it is very possible he may never see some of them again. I look back on the same days of my high school experience and regret that in those last days i did not do more with my friends. I did not even say goodbye to most of them and have not seen a single one for almost three years. Of course there has been the occasional e-mail here and there, but it's not the same, and can never be the same as talking face to face. I look back on those fun times in high school and wonder what happened. I miss the days of sitting in the Lower Quad eating lunch, bashing our teachers, and talking about TV, sports and the like. I miss our after school Friday basketball games. I miss going out to the theatre and movie theatre. Most of all I miss the company. Perhaps one day in the future we will all see each other again, but it will be very awkward. Everyone will have changed...
-Looking back on what happened to my friendships from high school, i can only wonder what will happen to my friendships here at Cal. Will they suffer the same fate? To some degree they must suffer inevitably since we would not be in the same places. It is quite depressing to be thinking about this, but my brother's upcoming graduation has stirred up old regrets about my own last days before high school graduation.
"True friendship is like sound health; the value of it is seldom known until it be lost"
-Charles Caleb Colton


June 5, 2003 - Making a polyacrylamide gel
-7:02 pm: We did not "do" a Western blot per se, we simply observed how to "block" (using milk interestingly enough) the nitrocellulose paper. Blocking prevents non-specific binding of antibodies (which are very expensive). After observing the blocking process, we were assigned the task of making some solutions of BSA which we are going to run through SDS-PAGE. The most exciting (and long) part about today was actually making the gels. Back in Bio1a the gel was was already made for us, but those prefabricated gels do not have enough lanes for our purposes (we are running 12 lanes at once). Thus we had to make the gels ourselves, something that proved both interesting and challenging. First a veteran lab student made the acrylamide solutions, then we had to form the gels from them. The hardest part was creating the seal at the bottom of the mold... we used an acrylamide solution that polymerized really fast, so pipetting the liquid before it solidified proved pretty difficult. My first attempt failed (the stuff polymerized in the pipette tip) and even the second time i wasn't sure if i got a good seal. Fortunately i did get a good seal and making the rest of the gel was relatively easy. Tomorrow we are going to actually run our gels with the BSA samples.
-In other news, a post-doc in the lab got mad at us for using his bench space (we didn't know it was his.. oops) and started accusing the student who was helping us of stealing buffer solution. We were also unknowingly using his micropipettemen.... I now know who to avoid. Everyone else in the lab is pretty friendly and definitely far more patient with us than i would ever be hehehe.


June 4, 2003 - Bradford Protein Assay
-8:22 pm: So the goal of today's lab lesson was to learn how to properly micropipette. And boy did we have a LOT of practice. Basically we did something called the Bradford protein assay. The assay involves a dye that changes color when it binds protein, and based on the amount of color change and comparison to standardized concentration-absorbance curves, one can determine the concentration of protein in solution. We actually didn't do any protein concentration measurements, instead we prepared solutions for 3 equivalent standard curves. This involved making around 50 separate solutions using some insane number of micropipette tips (of course proper technique is to use a different tip for each solution)! The idea was to test our micropipetting ability; each of the standard curves should come out the same. My hand hurts from depressing the micropipette so much although it was pretty fun. I used to be pretty bad a micropipetting, but after doing it around 300 times today, i think i can probably micropipette with the best of them! Yay! I passed the "test" and get ot move onto Western blotting tomorrow.... oooooooh.


June 3, 2003 - Memorable Moments before college
-8:22 pm: i'm currently in the nostalgic mode, remembering things from before college. The following is a brief summary of the most memorable things from school years past (6th through 12th grade)
6th Grade: Space Camp and Washington D.C. -got carsick on both trips (yikes). Sprained arm at space camp, but enjoyed "living among the trees." Watched July 4th fireworks from the National Mall
7th: Renaissance Faire. Dressed up as monk for school Renaissance Faire, had a disastrous time making raisin bread
8th: Yosemite Trip: Spent 6 days in Yosemite Institute program. no shower, Spider Cave, wet Mist Trail, crossing "lava" in Tuolumne Grove
9th: frequently absent biology teacher (i think she was high or stoned most of the time), sue-happy math teacher (sued my high school over asbestos), overly nice english teacher (she NEVER got mad), fun Spanish class (Spanish Jeopardy is so fun)
10th: Mr. Crosby's class (a very terrifying place), first AP class (chem), damn PSATs
11th: Mrs. Lester's vocab lists (sooooooo looooooong), first exposure to physics (wow), damn SATs
12th: whacky math class (was it really a class?), seeing plays ( i miss seeing plays), sarcastic english teacher (bless his soul)
June 3, 2003 -
From paper to AutoClave, Apt. Construction
-4:09 pm: Just got back from research lab. Today we learned the basic of solution preparation, both simple salt solutions and pH specific buffers, and how to sterilize the solutions. In old chem and bio labs all of this was done for you; we didn't have to worry about weighing stuff and mixing with the magnetic stir bars. And pH-meters? what pH-meter? Seems like silly stuff to learn, but it's important. We cannot use the classic best-friend lab report excuse of bad chemicals etc... cuz we've made the chemicals ourselves. And since this is a lab dealing with mammalian cell cultures, sterilization is extremely important. Speaking of which, my favorite part about today was playing with the AutoClave machine. This machine sterilizes the solutions by heating them up for 20 minutes to 250 degrees F under 190 PSI pressure (high pressure prevents the solutions from boiling.... if you remember your thermo!!). The least fun part today was adjusting the pH of my Tris buffer to 8.0. I had to use 12 M HCl, which is particularly nasty stuff if gets on anything. I was so afraid of acid burn i had to use two hands on the Pasteur pipette... i should probably get used to it, overcome the fear, and come to grips with the fact that i will inevitably burn myself sometime...weeeeeeee. oh well. Tomorrow i get to re-learn how to do micropipetting. All this stuff might seem pretty boring to have to go through and do, but i was thoroughly enjoying all of it (except for the 12M HCl). Perhaps it was exciting because i hadn't done anything meaningful for an entire week, but nevertheless it was pretty fun. I'm looking forward to when we move past the basics and begin experimenting on cell biology. I'm particularly interested in experiments on vesicle budding and fusion, as that was one of my favorite parts from MCB130. w00t... synaptotagmin! (if you know what that is your either 1) a nerd,  2) an MCB-CDB major, 3) me).
-Non-research related blog: Aaaaaah... so the pounding of the exterior walls of the building continues... IT IS SO DAMN ANNOYING. They sound like they're going to bust through the interior wall at any moment. The worst is that there is absolutely nothing i can do about it. the management ppl of this building are unapproachable, money-loving, unethical and especially bitchy. I really don't know why we didn't look for another place to live... oh yeah cuz moving and shit is so damn annoying too. Annoying enough to tolerate these management ***holes? i'm starting to reconsider.


June 2, 2003 - First Day of Lab
-10:20 pm: Yay! So I actually did something other than watch TV today... I officially began my research position in a protein trafficking lab. Being totally inexperienced, we began today with the absolute basics: lab safety and solution preparation. Nothing terribly interesting about lab safety although I was intrigued by the Geiger-counter in the lab, since i had never seen one before. Beep... Beep... ... ... Beep! Beep! Beep! hahah. For those that don't know, radioactivity is used to study protein trafficking. The specific process is called pulse-chase; it involves exposing cells to radioactive amino acids for brief pulses and tracking the incorporated radioactive amino acids as they move through the cell's pathways. Solution preparation was all on paper... we had a series of solutions we had to describe how to make from powder and stock. Much more difficult and tricky than it seems. Actual preparation will be tomorrow.


June 1,  2003 - Watching Panzer Dragoon Orta
-9:57 pm: So the TV in my apartment is now uber special. It has the privilege of having displayed games from all three next-gen consoles! Yeh came over with his brand new Microsoft Xbox to play Sega's Panzer Dragoon Orta. He only has a small 6" TV at his place, and needless to say the game probably looks better on a 28" Sony Wega. The game itself looks pretty fun. It involves shooting enemies with a genetically engineered dragon in order to protect the genetically engineered heroin, Orta.  I've watched Yeh play through almost the entire game, the last boss being too uber-difficult to beat.
-In other news, my boredom should officially end tomorrow as my research position officially begins at 2pm. I'm very excited, to say the least, as anything at this point beats watching TV all day. The last week has definitely been a great waste of time... i did not do anything even remotely close to being productive.



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