User:Linh N Le/Notebook/2009/06/22

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To Do
12-5
 * Andy is back! He wants to try the kinesin again, maybe his expertise will allow us to succeed
 * Make BRB80

Tubulin

 * Stumbled upon a site that details how they get tubulin from cow's brains

BRB80

 * Since Andy and Koch are the only people in the lab that know how to make a buffer (I actually dont know if Ant can), we waited til Andy came back to try to make BRB80
 * Not to mention the fact that we are also running out of the stuff we got from Cytoskeleton (new shipment will be in this week)
 * Andy is looking over our new fancy pH meter
 * He is starting the calibration process
 * BRB stands for Britton-Robinson Buffer
 * It is a "universal" buffer whose ph changes linearly with the amount of alkali that is added to it
 * It consists of:
 * .04M H3BO3 (Boric Acid)
 * .04M H3PO4 (Phosphoric Acid)
 * .04M CH3COOH (Acetic Acid)
 * The stuff we use is also known as PEM
 * 0.1 M PIPES (pH 6.95)
 * Piperazinediethanesulfonic acid C8H18N2O6S2
 * 2 mM EGTA
 * ethylene glycol tetraacetic acid C14H24N2O10
 * 1 mM MgSO4
 * Magnesium Sulfate
 * Also can make it as
 * 100 mM Pipes,pH 6.9
 * 1 mM MgCl2 (magnesium chloride)
 * 1 mM EGTA

Ph Titrations

 * Recalling what little i remember from freshman chemistry, Andy has instructed me to create 40uL of a "normal" base of NaoH
 * Normal meaning 1M soln
 * So, we look up the molar mass of NaOH
 * MM 39.9971 g/mol
 * If I want a 1 mol soln, i need 39.9971 g in 1L
 * Means I need 3.9971g in a dL
 * Means I need .39971g in a ml
 * Means I need .039971g in a ul
 * Meaning I need 1.598g in a 40ul soln
 * Thats wrong, new train of thought
 * Lets try to make .4L of 1M soln
 * I need 15.99884 g for .4L of water (4 deca liters)
 * divide by 10, 1.599884g for a 40ml

Making the Buffer

 * We measured out the following
 * 1mM EGTA (0.076g)
 * 80mM PIPES (4.83g)
 * 1mM MgCl2 (.2mL)
 * For a 200mL stock of the Buffer (which Andy hates naming, but I will name KLB)
 * We titrated it with NaOH to raise the Ph to 6.89
 * Used about 26ml of NaOH to get there
 * Added an entry to the Kochlab inventory page, under the VWR fridge category (Steve Koch 02:07, 23 June 2009 (EDT): Thank you! Excellent notes, as always.  I really appreciate it.)


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