electro-optical l-format (eolf) polarization of fluorescence module

Polarization Toolbox

Electro-Optical L-Format (EOLF) Polarization of Fluorescence Module

Add Anisotropy and Circularly Polarized Luminescence Easily

Traditional L-Format Anisotropy

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:

Anisotropy with New EOLF

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:

EOLF Advantages

• No Gain Correction Factor (G-Factor)

• No polarizer movement

• Rapid switching between polarization states supports high speed kinetic experiments

EOLF Chamber, PEM version

EOLF Chamber in a DM 45 Spectrofluorimeter

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

Photon Counter Gating

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

-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

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

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

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

CPL with Polarized Excitation

Use circular polarized excitation to photoselect one enantiomer in a racemic mixture

US Patent Pending

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

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