Deriving a sub-nanomolar affinity peptide from TAP to enable smFRET analysis of RNA polymerase II complexes.

Our capability to visualize protein complexes such as RNA polymerase II (pol II) by single-molecule imaging techniques has largely been hampered by the absence of a simple bio-orthogonal approach for selective labeling with a fluorescent probe. Here, we modify the existing calmodulin-binding peptide (CBP) in the widely used Tandem Affinity Purification (TAP) tag to endow it with a high affinity for calmodulin (CaM) and use dye-CaM to conduct site-specific labeling of pol II. To demonstrate the single molecule applicability of this approach, we labeled the C-terminus of the Rpb9 subunit of pol II with donor-CaM and a site in TFIIF with an acceptor to generate a FRET (fluorescence resonance energy transfer) pair in the pol II-TFIIF complex.

Improved axial point spread function in a two-frequency laser scanning confocal fluorescence microscope.

A two-frequency laser scanning confocal fluorescence microscope (TF-LSCFM) based on intensity modulated fluorescence signal detection was proposed. The specimen-induced spherical aberration and scattering effect were suppressed intrinsically, and high image contrast was presented due to heterodyne interference. An improved axial point spread function in a TF-LSCFM compared with a conventional laser scanning confocal fluorescence microscope was demonstrated and discussed.

Measurement of the surface effect of a small scattering object in a highly scattering medium by use of diffuse photon-pairs density wave.

The surface effect close to the boundary of a small light-scattering object in a highly scattering medium is experimentally demonstrated. This is the first attempt to measure the surface effect of a small spherical scattering object in 1% intralipid solution by use of developed diffuse photon-pairs density wave (DPPDW) in terms of the amplitude and phase detection. Theoretically, the surface effect of a small scattering object in turbid media is localized close to the boundary according to the perturbation theory, concerning an inhomogeneous distribution of the diffusion coefficient in the frequency-domain diffusion equation.

Zeeman laser scanning confocal microscope and its ability on reduction of specimen-induced spherical aberration.

The spherical aberration induced by refractive-index mismatch results in the degradation on the quality of sectioning images in conventional confocal laser scanning microscope (CLSM). In this research, we have derived the theory of image formation in a Zeeman laser scanning confocal microscope (ZLSCM) and conducted experiments in order to verify the ability of reducing spherical aberration in ZLSCM. A Zeeman laser is used as the light source and produces the linearly polarized photon-pairs (LPPP) laser beam.

Dual-frequency heterodyne ellipsometer for characterizing generalized elliptically birefringent media.

This research proposed a dual-frequency heterodyne ellipsometer (DHE) in which a dual-frequency collinearly polarized laser beam with equal amplitude and zero phase difference between p- and s-polarizations is setup. It is based on the polarizer-sample-analyzer, PSA configuration of the conventional ellipsometer. DHE enables to characterize a generalized elliptical phase retarder by treating it as the combination of a linear phase retarder and a polarization rotator.

Differential phase decoder in a polarized optical heterodyne interferometer.

A differential-phase decoder (DPD) together with a polarization common-path optical heterodyne interferometer is set up. Based on this interferometric configuration and a novel balanced-detector scheme, the performance of the quantum-noise-limited differential-phase decoder is demonstrated and analyzed. The minimum-detectable differential phase is on the order of 10(-7) rad/sqrt Hz when a 2.

Degrees of polarization and coherence of paired linear polarized laser beam by scattering glass plates measured using optical coherent ellipsometer.

An accurate optical coherent ellipsometer (OCE) is proposed and setup in which a two-frequency paired linear polarized laser beam is integrated with a common-path heterodyne interferometer. This OCE is able to precisely measure the optical properties of scattering specimen by measuring ellipsometric parameters (Psi, Delta). In the mean time the degree of polarization P, and degree of coherence Chi of incident two-frequency linear polarized laser beam are measured too.

Measurement of diffuse photon-pairs density wave in a multiple-scattering medium.

As a continuation of the previously developed theory of a diffuse photon-pairs density wave (DPPDW) [Appl. Opt.44, 1416-1425 (2005)APOPAI0003-693510.

Determination of retardation parameters of multiple-order wave plate using a phase-sensitive heterodyne ellipsometer.

To characterize the linear birefringence of a multiple-order wave plate (MWP), an oblique incidence is one of the methods available. Multiple reflections in the MWP are produced, and oscillations in the phase retardation measurement versus the oblique incident angle are then measured. Therefore, an antireflection coated MWP is required to avoid oscillation of the phase retardation measurement.

Properties of a diffused photon-pair density wave in a multiple-scattering medium.

A novel diffused photon-pair density wave (DPPDW) composed of correlated polarized photon pairs at different temporal frequencies and orthogonal linearly polarized states is proposed. A theory of DPPDWs is developed. A DPPDW selected by coherence gating and polarization gating that satisfies the diffusion equation has been verified experimentally.