User:Linh N Le/Notebook/2009/06/11

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New Computer

  • We finally got in our new computer for the lab
    • A brand new Dell optiplex 960
  • I, stupidly, bought the wrong video card for it (just got the integrated stock one) and that cannot handle dual screen
    • Will need to get one that can, wont be more that $100
  • I set it up and it runs just fine
  • We wiped it and are setting up the custom xp that caleb made for us that has all the networking stuff built in
  • Currently reformatting the HUGE drives, 1hr in and its at 16%
  • I hope the RAID 1 system is easy to get back in order
  • time: 4:12pm, 67% done with reformat
    • Still have to install windows :(
  • computer done installing at 5pm
  • nothing comes up when i log in as the admin
    • The problem i think is that the install disc that koch made does not have all the drivers that this computer needs
      • I think this b/c although it is hooked up to the internet, i get no connection or response from the ethernet cable.
  • Will try the drivers cd 2morrow or dl a fresh batch and send them over via usb stick

Acetic Acid

  • while waiting on the computer, i will help brigette make MT's
  • she is mixing up the acetic acid
  • Note ok MT's: I will be using the BRB80T that Igor and I made earlier this week (monday?)
    • it has been in the fridge all this time, so i think its still good
    • mixed it up using the vortexer (koch mentioned that taxol can come out of soln if it sets) and am leaving it on the counter for ~20mins to warm up before adding it to the MT's
  • Eventually i will remember the recipe for making MT's but for now I had to look it up
    • As of this monday, its 2ul of tubulin and 198ul of BRB80T (premixed)
  • Tubulin + brb80t mixed up
    • Andy is messing with the microscope right now...he has the translator stage off, so I wont bother trying to look at the MT's until he gets back and gives me the ok
  • looked at the MT's, no antifade. They are there. They fade in ~.25s
  • Now to add acetic acid to the MT's
    • Brigette made .273M Acetic Acid soln (w/DI H20)
    • She wants to add 3ul of this soln to the ~200ul of tublules (technically only 195 since i took 5ul to look under the scope)
    • Scratch that. We made a soln of 100MT to 3ul of acetic and 100ul MT to 1.5ul acetic
  • Right now, brigette is looking at the 3ul aa soln
    • They are still photobleaching quite fast, so it is really hard to tell if the tubes are breaking apart or not
  • Now trying it with the stock of AF that brigette made last week
  • Doing
  1. 10ul of the tubes + 3ul acetic acid
  2. 2.5ul of AF
  3. 1ul of 2M dextrose
  4. 80ul of BRB80T
  • took the soln above and gave briggete and sample to look at
  • took the soln above, added 3ul more aa to the whole stock, make sample for brigette to look at
  • note: I only used maybe 1/10 of the AF cocktail today
    • refroze the aliquot, to save money, marked the aliquot with "thawed 6/11"
    • I hope AF is ok with being thawed, refrozen and thawed again
  • Brigette and I have spent the good part of an hour trying to image our new samples.
    • ~45mins after creation, I took a look a new sample of the AF+MT(3ulaa)+dex+brb80t+2ulaa mix and saw some floaties.
    • The AF didnt seem to work very well. The background went black too fast!
  • Getting frustrated at the fact we could not see anything, I went ahead and sampled out the pure tubes +1.5ul aa
    • We saw plenty of tubes, and even though they do fade quite rapidly, you can still see them faintly in the background
  • Conclusion of the day: AF does not screw up MT. AA does not screw up MT. Mixing the 2 messes everything up!
  • In a last ditch effort, Brigette added 10 more ul of MT(with 3ul of AA) to the AF mixture
    • Hopefully more MT's allows us to see some
  • On a side note: we really need another microscope as Andy came back into the lab and wants to work on the tweezers, but we are still looking at samples!
  • MT's, no antifade, with aa, can see tubes. MTs with AF, with aa, still fades badly, but not tubes. "bad" AF that doesnt stop fading, +MTs, can see tubes, but they fade fast (as we saw last week). MTs + aa +AF (whether or not its good b/c it was in the freezer, or even if I messed it up by not thawing it right) no tubes. aa+af=bad for tubes.