User:Madeleine Y. Bee/Notebook/Single Molecule Fluorescence/2013/06/26

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June 26, 2013

Notes from Literature

Looking for information on silica-coating thickness for DNA attachment procedure.

  • From: Distance and Wavelength Dependent Quenching of Molecular Fluorescence by Au@SiO2 Core-Shell Nanoparticles
    • References 8-13, 15, 26
    • "Gold nanoparticles between 10 and 20 nm in diameter are important because they are too small to scatter light significantly[...]fluorescence of molecules in close proximity to these particles is quenched."
    • "...almost complete (>90%) quenching at short separations to negligible electromagnetic coupling at a distance of 43 nm"
    • "...hydrolysis of the reactive NHS moiety on the dye is much slower in ethanol than in water." (For attachment procedure)
    • "...attachment was more efficient for particles with a thicker silica shell due to the higher surface area and the increased number of binding sites available per particle."
    • "For average dye separations above 15 nm (less than 30 dyes per particle)[...]small (<5%) decrease in fluorescence lifetime and in a 20% decrease in steady-state fluorescence relative to the fluorescence of the free dye. A very pronounced decrease in both fluorescence intensity and lifetime is observed for dye separations below 10 nm (more than 75 dyes per particle). For the highest dye loading, the steady-state fluorescence decreases by more than 90% and the fluorescence lifetime is reduced to 0.5 ns compared to 4.0 ns for the free dye in solution."
  • Solvents: ethanol for coated spheres, 2-propanol for uncoated

FCS Measurements

  • Samples prepared to be added to data and calibration graphs from 06/25/2013:
    • (1) 100pM DNA/120pM MB
    • (2) 80pM DNA/96pM MB
    • (3) 60pM DNA/72pM MB
    • (4) 40pM DNA/48pM MB
    • (5) 20pM DNA/24pM MB
    • (6) 100pM Oligo D
    • (7) 80pM Oligo D
    • (8) 60pM Oligo D
  • Data added to calibration curves from 06/25/2013, but samples will be made and run again due to inconsistencies

DNA/ThT Sample Prep

  • Another 700nM sample was made to complete the calibration curve for the DNA/ThT samples
  • Fluorescence measurements taken
  • Results still inconclusive, will be redone tomorrow