User:Klare Lazor/Notebook/Chem-496-001/2011/11/15

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Objective

  • make purple solution
  • make purple fibers

Description

green test tubes......1:2 ratio of gold to BSA with a volume of 10mL

  • test tube A (Sent off to lab for analysis labeled GA)
    • 250µL of gold 4.17mM
    • 500µL of BSA 30µM
    • 9250µL of H20
  • test tube B (Sent off to lab for anlysis labeled GB)
    • 500µL of gold 4.17mM
    • 1000µL of BSA 30µM
    • 8500µL of H20
  • don't take out trying to make solution
  • nothing happened within two hours left over night


Red test tubes......1:1 ratio of gold to BSA with a volume of 10mL

  • test tube A (Sent off to lab for analysis labeled RA)
    • 500µL of gold 4.17mM
    • 500µL of BSA 30µM
    • 9000µL of H20
  • test tube B (Sent off to lab for analysis labeled RB)
    • 1000µL of gold 4.17mM
    • 1000µL of BSA 30µM
    • 8000µL of H20
  • leave in for 50minutes and take out for 10 min


[[Note:]] Both concentrations are the same, but doubled from previous solutions or experiments.



Also today we will make

  • 1 L 50mM Tris + 1M NaCl pH 9 Filter
    • 6.05 grams of Tris + 58.4grams of Nacl and made to a volume of 1 Liter
  • 1 L 50mM Tris pH 9 Filter
    • 6.05 grams of Tris and made to a volume of 1 Liter
  • 4 L 50mM Tris pH 9 (2x)
    • 24.228 grams of Tris and made to a volume of 1 Liter
  • MW Tris--> 121.14 g/mol MW NaCl--> 58.4g/mol
  • all were calibrated to a ph of 9 using 3M HCl
  • the smaller flask were filtered for contaminants from tris and nacl


PROTEIN EXTRACTION

Sonacation of previously made cells

  • Cells that were centrifuged down and transferred to tubes, and then frozen to later thaw to aid in process of lysing. freeze/thaw has been shown to effectively release recombinant proteins located in the cytoplasm of bacteria and is recommended for the lysis of mammalian cells in some protocols.
  • they were than sonacated for 30 seconds and not for 30secounds. This process was repeated three times for each test tube of cells.
  • In principle, the high-frequency is generated electronically and the mechanical energy is transmitted to the sample via a metal probe that oscillates with high frequency. The probe is placed into the cell-containing sample and the high-frequency oscillation causes a localized low pressure region resulting in cavitation and impaction, ultimately breaking open the cells. Although the basic technology was developed over 50 years ago, newer systems permit cell disruption in smaller samples (including multiple samples under 200 µL in microplate wells) and with an increased ability to control ultrasonication parameters (wikipedia.com).
  • Then tubes were weighed and then each brought to the same weight using water. common weight was roughly ~117 grams. And, placed into a centrifuge.
  • the tubes were centrifuged for 1 hour at 18000 RPM
  • solution was placed in bags with pores and than places in 4L tris solution with ph 9. equilibrium will form with ions. placed in freezer to stabilize proteins overnight

Data

1:1 RATIO OF GOLD TO BSA RESULTS

Image:Screen_Shot_2011-11-15_at_3.49.32_PM.png


1:2 RATIO OF GOLD TO BSA RESULTS

  • After two hours in the oven at 80 degrees Celsius nothing happened
  • After twenty-four hours the solution turned purple, NO FIBERS !!

Notes

This area is for any observations or conclusions that you would like to note.


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