Transcript Trouble-shooting. (also a science) Things that didn’t go as expected • Gold color did not disappear over 2 hour time rocking at room temperature •

Trouble-shooting.
(also a science)
Things that didn’t go as expected
• Gold color did not disappear
over 2 hour time rocking at
room temperature
• After reduction and silver step,
purple color did not develop
• Are there nanowires? What
happened??
Clear – what it is supposed to look like!
Gold – after two hours of rocking
Dark orange – something’s not right
“Low hanging fruit”
• Phage titered correctly? Concentrations
calculated correctly?
– T/R lab, John and Bridget went through all of your
notebooks looking for these calculations
– This is why you always right neatly and make sure
to read directions! Only one group remarked on
the color change that took place.
– End result: even if there were slight calculation
errors, all the phage added into the reaction
mixture is in the correct order of magnitude
Too much gold for the ascorbic acid?
• Calculated the number of gold particles (based on
molarity of gold solution)
• Calculated the number of nucleation sites (based on
concentration of phage and number of p8 proteins)
• Phage reduces the gold that binds to the mutated p8
protein
• Calculated amount of ascorbic acid able to reduce the
extra free gold in solution (if there weren’t enough
phage)
• End result: reducing power is still in excess – this
shouldn’t be a problem
Problem of “cutting corners * 1000”
• If we are an order of magnitude off in terms of
phage and an order of magnitude off in terms of
gold, does this make the experiment just fail?
• End result: Maybe, but nothing so drastic. This
“cutting corners * 1000” is more likely to have an
effect on the nanoscale and create nanoparticles
instead of nanowires…it isn’t something that
would change how we’d see the color change
Contamination?
• Belcher Lab has had issues with E4 contamination
in the past
– Remember E4?
• Normally perform a DNA sequencing analysis to
check on this
– How could this experiment be done?
• End result: phage contamination would not
change the goldclear or the purple color
change, so this isn’t contributing to our problem
pH?
• What if the pH of both the water and TBS is
off?
– Nanopure water vs. DI water
• End result: pH of both Belcher solutions and
20.109 solutions were similar (within 0.01)
What about the other solutions?
• Hmm, that gold looks sort of dark…
• Absorbance on a spectrophotometer
• Gold solution used T/R was significantly darker
(error in solution-making?)
• Bridget re-made gold solution for W/F lab
• Is this it??
Gold color didn’t disappear. Again. Now
what?
• What about TBS? I mean, it’s a standard
buffer, right?
• Apparently not. . .
Ionic Strength
General Equation:
or
Biorad 1xTBS:
pH = 7.5
0.020M TBS
0.5M NaCl
Rockland 1xTBS:
pH = 7.5
0.1M TBS(HCl)
0.15M NaCl
Therefore,
IBiorad TBS = 0.518
IRockland TBS = 0.241
Qualitatively increased ionic strength results in:
• Increased ionic charge shielding
• Decreased ionic mobility (and diffusivity)
These factors should:
•Decrease the ability of gold ions to penetrate CTAB bilayer
• A high [NaCl] might lead to Na+ binding at p8