Tea and Crumpets: A Summer in Science

Date: 28.6.13 Time: 13:00 Location: The Dunn School of Pathology, Oxford, England Trypanosome division My final day in Oxford began with Helen, counting cells under the microscope that have been tagged with DAPI, which tags the nucleus and kinetoplast.  The cells were then counted and sorted into groups based on where they were in the life cycle.  The five groups were 1K1N, 2K1N, 2K2N, Zoid, and Other.  1K1N, 2K1N, and 2K2N refers to the number of nuclei and kinetoplast in the cell.  For example, 1K1N has one nucleus and one kinetoplast, so that cell has either just undergone cell division or has not yet begun cell division.  2K1N cells have begun dividing, as they contain two kinetoplasts, which are the first part of the cell to divide.  2K2N cells have almost completed division, as they contain both two kinetoplasts and two nuclei.  Zoids are cells where division has gone wrong, they contain only one kinetoplast and no nuclei.  Other is for any cells that have encountered ot

Date: 27.6.13 Time: 18:09 Location: The Dunn School of Pathology, Oxford, England 96 well plate, round bottom Starting off my penultimate day in Oxford, Jack and I helped Richard (who is away doing research in Africa) with his ongoing experiment.  He is studying transfection efficiency.  When cells are transfected and the new DNA is placed in the cells, cells that have not taken up the new DNA are killed with a drug that the new DNA is resistant to.  The cells are then placed in 96-well trays so that some grow, leaving samples of cells with only the new DNA.  Richard took cells and put them in trays and then created pairs for each tray with a 1:100 dilution. We looked at each tray and counted the number of wells where the cells were alive and growing.  There were many without any growth, however that should be expected as the only times we should have seen any growth was when there were over 50 wells with growth in the undiluted solution.  However, there is also potential for

Date: 26.6.13 Time: 8:14 Location: Dunn School of Pathology, Oxford, England This morning began with a deflagellation experiment with Francois.  Deflagellation is the process of removing the flagellum from the cells (in this case, leishmania) for further testing of the isolated component.  This is done by taking the cells and first centrifuging them from their media then placing them in a PBS solution.  Once in solution, the cells are counted.  The deflagellation process has proved to only provide a 7% rate of recovery of useful material for Francois so he needs to count the cells with frequency to be sure he isn't losing too much of his sample. Leishmania before deflagellation  After the cells were counted, the solution was spun down again in the super-centrifuge again at 100,000g.  The supernatant was discarded and the cells were placed in fresh solution.  The cells were then placed on special microscope slides that are divided into tiny little grids that can then be use

Date: 25.6.13 Time: 19:49 Location: St. Edmund Hall, Oxford, England This morning began with some work with Helen transfecting cells.  This is done, like before, by preparing and separating out DNA samples that are then put back into the cells by blowing holes in them using electrocution.  This was very exciting as I personally was able to electrocute the cells!! The cells are then put on microscope slides.  This is a very lengthy process and therefore took most of the morning.  We then looked at the cells under the microscope and found some good images where the protein of interest was present and well-indicated. Next, Mike gave me an overview of mitochondrial DNA (which is referred to as the kinetoplast in trypanosomes).  In human cells, there are tons of mitochondria within each cell, however trypanosomes are different.  They only have one kinetoplast.  It is believed that mitochondria were independent species such as bacteria that were floating around near the cells and on

Date: 24.6.13 Time: 17:38 Location: St. Edmund Hall, Oxford, England This morning began with a lab meeting and a presentation by Helen.  She is currently writing and hoping to publish a paper on her work with trypanosomes thus far.  She has been working with the role certain proteins play in cell division, specifically BA25 and BA35, which are two of the proteins our very own Bungo discovered.  She found that when procyclic form of the cells lack BA25 or BA35, there is no change in their splitting.  However, when the bloodstream form lacks these proteins, they turn into 2K1N cells (cells that are not able to duplicate), and eventually die.  Therefore, finding a way to remove the BA25 and BA35 proteins from the bloodstream form would provide a cure for the disease they cause.  Also, when publishing her paper, Helen would like to provide official names for the BA25 and BA35 proteins (since they were discovered in this lab they do not yet have official names besides Bungo's initia

Date: 21.6.13 Time: 14:40 Location: Dunn School of Pathology, Oxford, England Today was the graduate student symposium, where first, second, and third year students presented the work they have done thus far and work they plan on doing in the future.  First and second year students prepare posters explaining their work, while third year students give 15 minute talks with questions.   First up was Narin Hengrung who discussed his work with the structural properties of influenza polymerase.  The portion of the influenza virus he is studying is incredibly small and not much is known about its structure.  He is studying the PB1, PB2 (which are basic components), and the PA (which is an acidic component).  In order to gain more information on the structure he looked at other viruses that have similar characteristics to the influenza virus. He also discussed the many problems he encountered during his work, reminding me that you fail often in science and that acknowledging  and shar

Date: 20.6.13 Time: 13:35 Location: St. Edmund Hall, Oxford, England Chart of Eukaryote supergroups This morning one of the members of the lab from Japan explained his work to me.  There are two different types of cells, prokaryotes and eukaryotes.  Prokaryotes are simple cells such as bacteria while eukaryotes are complex, like humans and yeasts.  Chromosome splitting Orange is kinetochore When cells divide, the chromosomes must divide as well.  In order for this to happen, the chromosome pair (which looks like two graphs of a trigonal bi-pyramidal atom with two lone pairs), is recognized by two microtubules which attach to the kinetochore (a series of proteins on the chromosome), and pull the two chromosomes in opposite directions, separating them.  While the contents of the kinetochore of other eukaryotes is known, that of the Trypanosome is not.  With 90,000 genes in the Trypanosome, that's a large gap in knowledge.  It also just so happens that this member of the

Date: 19.6.13 Time: 14:22 Location: Sir William Dunn School of Pathology, Oxford, England Removing trays of boxes This morning began with a trip to the cell bank.  Stored in nine giant vats of liquid nitrogen, boxes and boxes of cells are kept at -198*C.  The contents of each vat, which can hold around 90,000 cell capsules, are logged in a computer.  (That's a maximum capacity of 810,000 cell capsules!)  While this makes locating and organizing cells much easier, glitches in the system have electronically removed cells that are actually physically present.  This then gives the cell bank moderator the difficult task of deciding whether to keep or dispose of unknown cells that can no longer be identified.  Also, when placing the cells in the bank, an entire tray, which holds 8 8x8 boxes, must be removed from the jumbo in order to locate free space, raising the temperature of the cells and compromising the effectiveness of the liquid nitrogen cooling.  (I see room for innovati

Date: 18.6.13 Time: 11:01 Location: St. Edmund Hall, Oxford, England This morning in lab began with Jack and a milk rinse of the protein membrane began yesterday.  This was to cover the membrane with antibodies so that when the other antibodies were added to the membrane, they would only stick to the proteins, sort of like saturating a solution so that none of a new solid will accidentally dissolve.   From the lab to the microscope room, the fun continued with Helen.  Here we looked at cells that had been treated with a detergent to strip away the membrane and proteins on the outside so that they appeared transparent then tagged with immuno fluorescence to indicate certain parts of the cells.  First, we examined a protein that exists right where the cells split, between the two skeletons of the cells.  This protein is present in both the human and fly form of the parasite, however the fly form does not require this protein to duplicate.  On the contrary, the human form does.  Thi