User:Linh N Le/Notebook/2009/08/28

To Do

• Write my last official notebook entry as a KochLab member :(
• We are running out of PEM, so I would like to make some more before I go
• We got motility working yesterday, so Andy may want to continue to pursue that

Inventory

• Brian had to leave, but i did give him a crash course on how to use the inventory page
• Most of the pages are self-explanatory
• Also note that chemicals and software inventory require an update to the google docs page
• The optics section of the inventory is blank
  • My idea was to have it as a static page that just listed important opto-mechanics
  • Perhaps we can have a check in-check out system so we can keep track of who took what where and when

PEM

• Andy, Brigette and I made 205ml of PEM a month or 2 ago and its getting low
• I will try to make 200ml more and follow the exact steps I took last time
• I will need NaOH and possibly HCL to titrate the buffer
  • We should have plenty of normal soln left from earlier buffer makings
  • We have 4 tubes of NaOH but no HCL, lets hope I dont screw up the titration

Procedure

• Following what I did last time, I want the following:
  • 1mM MgCl2
  • 1mM EGTA
  • 80mM PIPES
• I am using the following chemicals:
  • 1M MgCl2 in 50mL falcon tube made 2 years ago ( we used it in the motility assay yesterday and it was fine)
    • For 1mM in 200ml, I want .2ml (200uL)
  • EGTA powder, working weight 380.35
  • For 1mM in 200ml i need .07607g
  • PIPES powder, working weight 302.37
  • For 80mM in 200ml, I need 4.83792g
• I will wash out a 250ml glass bottle with DI water then use DI water to suspend the salts
  • I will use the autoclaved DI water I made a month ago, that way we use it a up a bit since we have a fancy schmancy DI water machine
• I filled up the jar with ~150ml of DI h20, now to add the powders
• After addition of powders and washing it down with more DI water, I'm about 175ml
• Powder Concentrations used:
  • 200ul MgCl2
  • .0768g EGTA
  • 4.8387g PIPES
• Note: I used the "DI Water" in the spray bottle. It may be tap water, Andy and I both dont know who filled it up with what. Last time I made buffers I used the water in the spray bottle, hopefully it still works
• Now to titrate

Titration

• Starting Ph
• added 25.8mL of 1M NaoH to get it to a Ph of 6.89
  • Thats 12.9mM of NaoH in 200ml soln of PEM

Test Drive

• Andy is making motility stuff and is going to try my PEM, hope it all works!

DI H₂O

• I went ahead and emptied out all the squeeze bottles that are labeled DI water and replaced it with the stuff from our DI water machine