Schematic of the experimental setup described in the text. Red (darker) solid and dotted lines denote the IR laser beam paths. The blue (lighter) solid line denotes the illumination light path.
Schematic of illumination and optical trap pathways with polarization states. Green corresponds to the illumination light and red corresponds to the trapping laser. Double-headed arrows/ellipses show the polarization state at various points in the illumination pathway, and black arrows correspond to s- and p-polarization axes with respect to the dichroic mirrors. Abbreviations: LED = light-emitting diode; LCF = laser clean-up filter; P = polarizer; QWP = quarter-waveplate; DM = dichroic mirror; DP = DIC prism; Cond. = condenser; Obj. = objective.
Calculated change of polarization state after reflection as a function of wavelength. Ψ (red solid line) gives the counterclockwise rotation with respect to the abscissa and tan χ = b/a (dotted black line) gives the ratio of the semi-minor axis to the semi-major axis (ellipticity). The inset shows the coordinate system with respect to the three variables. The four ellipses above show the elliptical polarization state and rotation after reflection at several wavelengths: left to right: 447, 467, 488, and 505 nm. Data regarding reflectance and phase shift for s- and p-polarizations upon reflection were kindly provided by the manufacturer (Chroma Technologies).
DIC images of microtubules fixed to a coverslip with varying degrees of compensation for the described polarization and optical rotation effects. From left to right, top to bottom: No compensation, only QWPs included, only LCF included, both QWPs and LCF included. All images are enhanced by background subtraction and represent the average of 10 frames at a video rate of 25 Hz. Image contrast was adjusted in each image individually for optimal visibility of the microtubules.