Team:UCLA/Notebook/Materials/19 August 2015

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Attempt and Fail of spinning HFIP Silk

• • 30% wt/vol HFIP dope, as instructed in the recombinant spider silk protocol

component	volume	total mass
lyophilized silk	N/A	134 mg
HFIP	447 uL	N/A
Tamura (2.970 mg/mL)	90.24 uL	0.268 mg

• OBSERVATIONS
  • When the Tamura was added, small liquid bubbles appeared on the surface of the HFIP. They looked kind of like oil bubbles on water
    • I think that the bubbles are water that isn't mixing with the HFIP
      • This makes no sense though -- Wikipedia states that HFIP is miscible with water
      • Other sources corroborate that HFIP should be miscible with water. After all, the main point of HFIP is that it's crazy strong at hydrogen bonding
      • Alternatively, maybe the bubbles are protein aggregates? They look so liquid though

• • After about an hour or so, there were two pieces that were still solid in the liquid
    • I poked at them with a pipette tip to try to push them and dissolve them
      • They behaved exactly like gels

• • After about another hour, the solid pieces still weren't dissolving, and the Tamura was just sitting on top of the solution.
  • I vortexed to try to dissolve the solid pieces and to mix the Tamura in. The same protocol by the Lewis group states that vortexing to mix the dope is ok

• • After about another hour, I tried to pipette the dope to place into the syringe
  • the damn dope had gelled up and I could not pipette it.

• POSSIBLE CAUSES OF THE GELATION
  • vortexing
  • the fact that I used a pipette tip to mix the dope around
  • addition of water in the Tamura

• THINGS TO TRY FOR NEXT TIME
  • Chop up the lyophilized silk into smaller pieces
    • I placed large chunks of silk into the HFIP today

Concentration Dialysis

• Started at 3:00 PM
  • Aim for 17 hours on the concentration dialysis, so end it at 8:00 AM tomorrow
  • I'm doing 17 hours because so far, even 19 hours has caused the silk in the cassette to gel.
  • ANOTHER NOTE: I accidentally made the concentration solution 100 g PEG in 1000mL water, instead of 100 g PEG in 900mL water, as the protocol states. So I fucked up, this causes less osmotic pressure on the cassette, so honestly I won't be able to tell if it was the lower osmotic pressure or the shorter concentration time that makes a difference tomorrow.

• ENDED at 8:10 AM the next day
  • Enormous differences!
  • The dope was so much easier to remove from the cassette
  • I believe that I removed most, if not all of the silk in the cassette (~3 mL)
  • It's not as viscous as the dope usually is when extracted from the cassette.
  • Pipetted 100 uL to test the concentration.
    • There was 0.014 grams in dried silk, meaning 14mg/100uL = 140mg/mL = 14% w/v.