User:Michael S. Bible/Notebook/581/2014/10/08

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Procedures

R6G Fluorescence: Calibration and Measurement

  1. Make stock concentrations (both groups can use the same solutions)
    • Initial stock: 2500μM, secondary stock: 500μM
    • 0.10μM
      • Made with 500μM solution
    • 0.50μM
      • Made with 500μM solution
    • 1.0μM
      • Made with 500uM solution
    • 1.5uM
      • Made with 500μM solution
    • 2.0μM
      • Made with 2500μM solution
    • 0.75μM
      • Made with 500μM solution
    • 1.2μM
      • Made with 500μM solution

Thanks to Madeleine for making initial 2500μM solution of R6G.

  1. Take UV-Vis and Fluorescence spectra of these samples
    • Fluorimeter Settings:
      • 500nm excitation
      • 515-700nm scan range
      • 10.0nm slit width
      • 200nm/min scan speed
  2. Make a calibration curve based on UV-Vis.
    • Compare your data to some published values
  3. Make a calibration curve based on the fluorescence.
    • In order to do this, you'll need to measure the area under the fluorescence curve, not just the fluorescence peak height.

Calculations

To calculate necessary amounts of stock solutions to add to enough water to have a total of 10 mL of calibration stock solutions:

  • C1*V1 = C2*V2
    • secondary stock: 500μM
      • (2500μM)*x = (500μM)*(10mL) → x = 2 mL 2500μM R6G
    • 0.1μM
      • (500μM)*x = (0.1μM)*(10mL) → x = 2 μL 500μM R6G
    • 0.5μM
      • (500μM)*x = (0.5μM)*(10mL) → x = 10 μL 500μM R6G
    • 0.6μM
      • (500μM)*x = (0.6μM)*(10mL) → x = 12 μL 500μM R6G
    • 0.75μM
      • (500μM)*x = (0.75μM)*(10mL) → x = 15 μL 500μM R6G
    • 0.85μM
      • (500μM)*x = (0.85μM)*(10mL) → x = 17 μL 500μM R6G
    • 1.0μM
      • (500μM)*x = (1.0μM)*(10mL) → x = 20 μL 500μM R6G
    • 1.2μM
      • (500μM)*x = (1.2μM)*(10mL) → x = 24 μL 500μM R6G
    • 1.5μM
      • (500μM)*x = (1.5μM)*(10mL) → x = 30 μL 500μM R6G
    • 2.0μM
      • (500μM)*x = (2.0μM)*(10mL) → x = 40 μL 500μM R6G

Data

  • Add data and results here...

Notes

Add notes here.




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