User:Tara K. Luckau/Notebook/Team ConGen/2010/11/23

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Make Tris-Cl, pH 8.0

 * here we go again ... how come 'error' has to be a part of 'trial and error'?

Attempt #1 = Fail

 * used remaining amount of Tris-HCl
 * Promega Corp
 * Lot # 283523, molecular biology grade
 * Cat # H5121, 100g, MW=157.56


 * $$18.5 \mathrm{g} \bullet \tfrac{\mathrm{mol}}{157.56\mathrm{g}}\ \bullet \tfrac{\mathrm{L}}{0.5 \mathrm{mol}}\ \bullet \tfrac{1000 \mathrm{mL}}{\mathrm{L}}\ = 234.8 \mathrm{mL}$$


 * 1) Mixed 18.5g Tris-HCl with approximately 150mL NanoPure H2O
 * 2) Heated on hotplate with stirbar until dissolved
 * 3) Measured pH using electronic pH meter: pH = 4.12  :(
 * 4) added remaining 75mL (to total 234.8mL) and put in storage bottle, labelled "0.5M Tris-Cl, pH 4.12, TKL, 23 November 2010"


 * everything I read said to use HCl to bring this mixture to the desired pH (8.0), but it can only go more acidic! How do I get it more basic? Can I use NaOH? Google searches yielded nothing, neither did text messages

Attempt #2

 * found another container ... Tris
 * Fisher Scientific
 * Lot # 086040A
 * Cat # BP152-500, 500g, FW=121.14


 * $$200 \mathrm{mL} \bullet \tfrac{\mathrm{L}}{1000\mathrm{mL}}\ \bullet \tfrac{0.5 \mathrm{mol}}{\mathrm{L}}\ \bullet \tfrac{121.14 \mathrm{g}}{\mathrm{mol}}\ = 12.114 \mathrm{g}$$


 * 1) Mixed 12.12g Tris with approximately 150mL NanoPure H2O
 * 2) Heated on hotplate with stirbar until dissolved
 * 3) Measured pH using electronic pH meter; used concentrated aqueous HCl to bring pH to 8.0
 * 4) * Let sit for 1 hour (went to class)
 * 5) * pH rose to 8.16, so added HCl to bring back down
 * 6) * Let sit for another 30 min until stable at pH 8.0
 * 7) Added remaining water to bring total volume to 200mL
 * 8) * Transfered to glass bottle, half-turned screw top (for very loose fit), autoclave tape connecting bottle to cap
 * 9) * Labelled as "0.5M Tris-Cl, pH 8.0, TKL 23 November 2010"
 * 10) Autoclave: seems to have started fine (I waited for a couple minutes until it made clanking sound), but upon return 1 hour later, alarm was sounding
 * 11) * used: left autoclave, Setting #3 (Liquid, 45 min, 121°C)
 * 12) * autoclave tape was darkened, so Tris-Cl got sterilized ... considered good!
 * 13) re-pHed when cool to confirm ... pH 8.01 ... yay!


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