Electro-Optical L-Format (EOLF) Polarization of Fluorescence Module
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Transcript of Electro-Optical L-Format (EOLF) Polarization of Fluorescence Module
![Page 1: Electro-Optical L-Format (EOLF) Polarization of Fluorescence Module](https://reader035.fdocuments.us/reader035/viewer/2022062317/5a4d1b1d7f8b9ab059994310/html5/thumbnails/1.jpg)
Polarization Toolbox
Electro-Optical L-Format (EOLF) Polarization of Fluorescence Module
Add Anisotropy and Circularly Polarized Luminescence Easily
![Page 2: Electro-Optical L-Format (EOLF) Polarization of Fluorescence Module](https://reader035.fdocuments.us/reader035/viewer/2022062317/5a4d1b1d7f8b9ab059994310/html5/thumbnails/2.jpg)
Traditional L-Format Anisotropy
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Traditional Calculation of Anisotropy and Polarization
G =IHV
IHH
P =IV V – GIVH
IV V + GIVH
IV V – GIVH
IV V + 2GIVH
r =
Gain Correction:
Anisotropy:
Polarization:
![Page 4: Electro-Optical L-Format (EOLF) Polarization of Fluorescence Module](https://reader035.fdocuments.us/reader035/viewer/2022062317/5a4d1b1d7f8b9ab059994310/html5/thumbnails/4.jpg)
Anisotropy with New EOLF
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Calculation of Anisotropy and Polarization using an EOLF Module
P =IV V – IVH
IV V + IVH
IV V – IVH
IV V + 2IVH
r =Anisotropy:
Polarization:
![Page 6: Electro-Optical L-Format (EOLF) Polarization of Fluorescence Module](https://reader035.fdocuments.us/reader035/viewer/2022062317/5a4d1b1d7f8b9ab059994310/html5/thumbnails/6.jpg)
EOLF Advantages
• No Gain Correction Factor (G-Factor)
• No polarizer movement
• Rapid switching between polarization states supports high speed kinetic experiments
![Page 7: Electro-Optical L-Format (EOLF) Polarization of Fluorescence Module](https://reader035.fdocuments.us/reader035/viewer/2022062317/5a4d1b1d7f8b9ab059994310/html5/thumbnails/7.jpg)
EOLF Chamber, PEM version
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EOLF Chamber in a DM 45 Spectrofluorimeter
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Comparison of PEM and LCVR Properties
Property PEM LCVR
Optical Aperture 17 mm 18 mm
Optical Material Supracil Nematic Liquid Crystal
Operating Frequency 50 KHz 10 Hz or Lower
Wavelength Range 170 - 1400 nm 340 to 2000 nm
Waveform of Retardance Changes Sinusoidal
Square wave (IR and IL)
Static Retardance No Yes
Adjusting Retardance LCPL = RCPL (only one voltage)
LCPL and RCPL must be set independently (two voltages)
Kinetic Measurements milliseconds seconds
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Photon Counter Gating
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Excitation Anisotropy Spectrum of Rhodamine
-0.2
0.0
0.2
0.4
340 360 380 400 420 440 460 480 500 520 540
Pola
rizat
ion
or A
niso
tropy
Excitation Wavelength, nm
Polarization
Anisotropy
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-0.2
0.0
0.2
0.4
280 300 320 340 360 380 400 420 440 460
Pola
rizat
ion
Excitation Wavelength, nm
Fluorescein Spectra as a Function of Temperature
Increasing temperature
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EOLF Also Supports CPL
Circularly Polarized Luminescence (CPL)
• Differential emission of left and right circularly polarized light
• Fluorescence analog of circular dichroism (CD)
• Sensitive to environmental changes of chiral fluorophores
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Left and Right Polarized Emission Spectra
0
10000
20000
30000
570 580 590 600 610 620 630
Intensity, counts
Emission Wavelength, nm
Left polarized
Right polarized
Europium(III) tris(trifluoromethylhydroxymethylene)-d-camphorate in DMSO
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CPL Spectrum of Europium Compound
g lu
m
-0.8
-0.6
-0.4
-0.2
0.0
570 580 590 600 610 620 630
Emission Wavelength, nm
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CPL with Polarized Excitation
Use circular polarized excitation to photoselect one enantiomer in a racemic mixture
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US Patent Pending
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EOLF Adds Complementary Techniques to a Single Optical
BenchMeasurement Selectivity Sensitivity
Absorbance Chromophore Ground state environment
Fluorescence Fluorophore Excited state environment
POF (anisotropy) Fluorophore Excited state mobility
CD Chiral chromophore Ground state symmetry
CPL Chiral fluorophore Excited state symmetry
FDCD (fluorescence detected circular dichroism)
Chiral fluorophore Ground state symmetry
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With One EOLF, Measure:
• Conformation characterization of proteins and other biological macromolecules
• Equilibrium binding studies of macromolecules
• Characterization of metal ligand complexes
• Characterization of entantiameric mixtures