It never turns out well.

I went in to SLU yesterday to check on my students plates, and, if everything looked as well as I expected, to take the first timepoint.

Stop right there and review that last sentence. Do you see what my mistake was? You might say it was "going in to SLU yesterday". You might say "doing the timepoints for the students". I wouldn't necessarily disagree. But the real mistake was that I expected everything to look good.

Things did not look good. They looked quite bad. One after the other I found contamination, plates that weren't labeled, plates that didn't appear to have anything in them… so after I ended up throwing out ¾ of the plates, I began to wonder: are they all really not getting it? Or… is it me? I mean, I knew some of them were having difficulty, but I though some were OK.  But the evidence was to the contrary. As I see it, there are three possibilities:

1. I am a bad teacher.
2. My students are not coming to class prepared.
3. The experiment is flawed.

Let's work through each possibility:

1. I am a bad teacher. According to my TA I'm going over and above what has been done in previous semesters. I give my students detailed protocols; I describe visually the protocol; and I demonstrate for them how to do it. I am covering all different learning styles. Plus, I've gotten positive reviews from other classes I've taught and student I've mentored. That still doesn't necessarily rule out this possibility, but it does seem to indicate that the shortages aren't on my end.

2. My students are not coming to class prepared. Well, that is true. I am racking my brain to come up with a way to combat this particular problem, and would welcome any suggestions. However, even so, once they get to class they have everything they need to complete the experiment, so while this fact is annoying it shouldn't necessarily result in such a catastrophe of having to discard 75% of the work.

3. The experiment is flawed. I know it isn't inherently flawed because it has been done before, successfully. However, I haven't done it recently, and I haven't done it at SLU, so perhaps this possibility should be explored further. I decided to take some cells from my TA and set up the experiment myself- just to make sure that it could be done. My TA kept protesting that she would do it, but you know the saying… if you want it done right, do it yourself.

I would have preferred to start from scratch, but I didn't have time to make media. So I took her media and filtered it, and went to get all the dishes and supplies I needed. It was actually a useful learning experience for me, since I had never personally used the TC facilities at SLU, having imported all my cells from MRU where I do my research, and handing them directly to the TA who has been maintaining them since. So I'm gathering all the supplies, and that is when I noticed that the 12 well plates that the class had used for the growth curves were collagen coated. F*** f*** f*** f*** f***. I don't know for sure, having never actually directly compared, but I have a feeling that this is perhaps some of the reason the cells are not growing well and do not look right. I am extremely angry about this for several reasons.

>It will, no matter what, screw up the results. Hopefully not badly enough that my class won't still learn something.
>My TA didn't notice, when I asked her to find 12 well TC plates, that the box was labeled, in big letters, "COLLAGEN". Or, perhaps, she didn't realize it was important.
>I didn't notice it during class, when everyone was plating their cells.
>I'm not sure if I should be mad at myself- for not specifically checking to make sure these were the correct plates. But really, who would think that there would be collagen coated plates, and not plain old TC plates, in the supply closet? SLU has not much money, and I would never expect them to have the more expensive plates just laying around up for grabs.

At the end of the day, it is nobody's fault but my own. I have relied too heavily on my TA, and on protocols supplied to me by past-professors of the course. It is good that I am in there, getting my hands dirty, so to speak. I am turning over a new leaf. It took a Sunday visit to the lab for me to realize just how bad things were, but the semester is still young.

I'm getting ready to teach my first class at SLU. It is my first experience working completely independently to teach a class, and there is so much work that goes into it.

First, of course, I have to design the syllabus. When I say design I mean two things: First, figure out what to teach, when. Second: literally type it up in a coherent format for distribution. These are two different things but both have to be done well, and they are providing me with quite a challenge.

To be honest, though, I am not starting completely from scratch. This is a course that has been taught before, and I have several years worth of old course materials as a resource. I started with the old syllabi and tried to revamp them completely. The problem is that as a lab course, with experiments that will run over multiple days or even weeks, fitting everything in is like putting together a puzzle. I was advised to keep out one or two exercises that were often problematic to execute, always confusing to understand, and not really integral to the goals of the class. Seemed like good advice, so I eliminated those. I was also advised not to add anything new: leave the vacancy; give myself some breathing room my first semester out. Seemed like good advice, so I decided to reorder the remaining exercises. The next challenge has to do with budget: there isn't much of one. I can't start introducing all these newfangled techniques when we don't have the reagents or equipment for them.

The question becomes, then: How do I revamp an old class with very little money and a very limited number of ways to achieve the goals set out in the course description? I have been working on this puzzle for two weeks now, and have finally come to the conclusion that if it ain't broke, don't fix it. I have just taken last years' syllabus, added my name and contact info to the top, and retitled it for Fall 09.I have realized that I can update the class by focusing on the lectures and assignments, and leaving the labs alone.

I'm not done yet, though. That was only one fourth of the overall syllabus. Next topic for discussion: deciding what kinds of assignments to give, and how much each assignment will be worth.

For my non-scientist readers some background:

Many of us use cells that we grow in special little dishes for our research. These can be different types of cells that are isolated from different organisms. They can be healthy 'normal' cells (although they aren't exactly normal, since they are growing in a dish in a lab somewhere and not in their natural environment- the body). I use cancer cell lines, which means someone at some point took a tumor out of a patient, isolated the cancerous cells, and those tumor cells have been happily growing in our labs ever since. A primary cell line is one that has been very recently isolated- days old. Established cell lines have been growing for years or even decades. Some of these cancer cell lines have survived long past the patient. The T47D breast cancer cell line shown in the picture above was isolated sometime before 1980, and perhaps even as early as 1974. This technique of cell culture is a well established practice, but that doesn't mean it is foolproof. Cells in culture don't always behave the way you want or expect them to, and that can be very frustrating. Case in point:

I work with a lot of different 'primary' –oma cell lines. I use the word 'primary' with some caution, because although our manager of the tissue culture facility goes down to the OR once or twice a week to get new tumors from which to establish these cells, by the time I get them they are really at a passage past the point that qualifies them for still being primary, yet they aren't immortalized either. This is significant because although they do continue to grow for some time, at any unpredictable and random point in time they will reach a crisis and just up and die. It has nothing to do with passage number; and the frustrating thing about it is that they all seem to die at the same time. I will come in one day having plated mass quantities of cells in preparation for big experiment, and lo and behold all my cells are dead.

This has happened to me several times, and I am quite frustrated about it because it makes my progress on this project sporadic and sloooooooooooooooooooow. Several of us are working with these cell lines, each investigating our own angle, and this happens to each of us, but each at different points. I can rule out equipment failures, I can rule out media problems. It isn't contamination. It isn't that they are overgrowing, because that will actually cause this same problem so I am meticulous to the point of being anal about preventing that particular phenomenon. I have been doing tissue culture for 16 years now, and this is the first time I've had this problem, and it is driving me batty. My only other option is mycoplasma, so I am testing for that now, but even treatment with Ciprofloxacin doesn't help.

It makes me want to give up. I am incredibly discouraged. My PI keeps telling me not to wait for the cells, but since all of my experiments hinge on having these cells to work with, I don't really think there is any other option. Grrr. Meanwhile, I'm a cell miser. I don't throw anything away; I'll freeze the cell pellets, or freeze lysates at -80 and stock them away for future use. It isn't my favorite way to work, but at this point, it is the only way. Sigh. Maybe it is time to find a new model system. I'm thinking zebrafish.