Transient Spectrometer

The femtosecond transient spectroscopy system is based on a mode-locked titanium sapphire oscillator pumped with a CW diode-pumped solid state laser. The oscillator output is amplified at 1 kHz repetition rate in a titanium sapphire regenerative amplifier. Part of the laser output (90%) is used to pump an optical parametric amplifier or sent through a frequency doubler for generating excitation at a desired wavelength. The remaining part of the beam is used to produce the probe pulse by generating a white light continuum.

Excitation wavelength is available in the regions between 350 - 1200 nm. A pre-excitation pulse with high energy can be obtained in the region from 770 - 850 nm for two-pulse excitation experiment. Spectral changes from 350 - 1150 nm can be recorded.

An optical delay line is used to control the time delay between the excitation and the probe pulses. The probe and reference signals are collected by a fiber-coupled spectrograph and recorded by a dual diode array detector, or a photodiode at a selected wavelength.

Spectral and kinetic information is extracted by global analysis or SVD analysis using a locally written software ASUFIT.

Single Molecule Microscopy Setup

The single molecule microscope consists of an excitation laser, a confocal microscope base, high numerical aperture objective lens, and a sensitive detection system, which operates in single photon counting mode. The optics are designed such that the observation volume is extremely small, a few femtoliters or less. Detection of single molecules is achieved by two primary modes of operation. (1) Diffusion mode: the laser is tightly focused into a liquid sample that contains a very dilute concentration of fluorophores such that there is a negligible probability of having more than one molecule to occupy this volume at any given time. Bursts of fluorescence are detected as molecules randomly diffuse through the detection volume. (2) Surface mode: Fluoresecently labeled molecules are attached to a surface and scanned through the focused laser beam. This mode allows for much longer observation times since the diffusion component is removed, although the surface chemistry creates experimental challenges. Both cw and femtosecond pulsed lasers are used with this instrument. The pulsed laser is used for lifetime measurements and multiphoton excitation. Sometimes the fluorescent light is divided into two detection channels based on wavelength for fluorescence resonance energy transfer measurements (FRET). Other common single molecule measurements include fluorescence correlation spectroscopy (FCS) and population analysis.