Matt:LabNotes/2016-2-10: Difference between revisions

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>Mzcai
(Created page with "=Polyacrylamide Gel in tube Test= * Yesterday failed to get good complete polymerization even in the tubes ==Purpose== *Test polymerization of two...")
 
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==Conclusion==
==Conclusion==
*Droplets formed after washing with 1X PBS that looked similar to "oil droplets" from mineral oil but there was no mineral oil
*Droplets formed after washing with 1X PBS that looked similar to "oil droplets" from mineral oil but there was never any mineral oil added
**Possible that free radicals from APS or TEMED caused a small amount of [https://en.wikipedia.org/wiki/Elimination_reaction_of_free_radicals depolymerization of polystyrene]

Latest revision as of 20:09, 12 February 2016

Polyacrylamide Gel in tube Test[edit]

  • Yesterday failed to get good complete polymerization even in the tubes

Purpose[edit]

  • Test polymerization of two different acrylaide:bis-acrylamide mixes (28:1 vs 199:1)
  • Test mineral oil as a sealant from oxygen

Experiment[edit]

Samples[edit]

  1. 28:1, mineral oil, open lid
  2. 28:1, open lid
  3. 28:1, closed lid
  4. 199:1, mineral oil, open lid
  5. 199:1, open lid
  6. 199:1, closed lid

Procedure[edit]

  • Following this general Matt:LabNotes/Polyacrylamide_Gel_Protocol
  • 28:1 mix is made from 9 parts 29:1 and 1 part 19:1
  • 199:1 mix is made from 9 parts 40% acrylamide (from powder) and 1 part 19:1
  • Added just a couple drops of mineral oil to cover
    • Good layer separation
  • Mix APS in PCR tubes and let sit on bench for 30min

Results/Conclusion[edit]

  • 28:1 had noticeably stronger polymerization than 199:1
  • After aspirating mineral oil good gel formed beneath
  • Open tube also has a polymerized gel but some liquid layer on top and sides that did not polymerize
    • Probably due to oxygen exposure
  • Overall open, closed, and mineral oil are all pretty comparable

Polyacrylamide Gel on culture dish[edit]

2 Conditions[edit]

  1. Try polymerizing in a vacuum chamber to minimize oxygen
    • Stiff/dense gel formed along sides
  2. Try polymerizing in a styrofoam box with CO2 from dry ice to displace oxygen

Vacuum Chamber Procedure[edit]

  • Sample A1
  • Prepare 100ul 28:1 Acrylamide mix and add to dish
  • Put in vacuum chamber, apply vacuum, and then seal
  • Leave on bench for 30min

Results[edit]

  • Stiff/dense gel formed along edges of hole
  • Center of glass has nothing
  • Guessing that vacuum caused water to evaporate

Dry Ice Procedure[edit]

  • Sample A2
  • Prepare 100ul 28:1 Acrylamide mix and add to dish
  • Put in styrofoam box with beaker of water and drop dry ice in the water
    • Leave lid of box slightly ajar
  • After 30min no noticeable polymerization so kept it in the box with dry ice for 4 more hours

Results[edit]

  • After ~5 hours the center was dry
  • The solution on edges was more viscous than before but not exactly a gel

Conclusion[edit]

  • Droplets formed after washing with 1X PBS that looked similar to "oil droplets" from mineral oil but there was never any mineral oil added