The NanoBiophysics Core Facility offers a Spectrofluorimetric System from PTI.
The System includes QuantaMaster spectrofluorometer, which is a sophisticated multifunctional instrument. It has dual-emission detection in the UV-VIS spectral range, and with the included calcite polarizing prisms, accurate fluorescence anisotropy measurements can be made, along with the capability of monitoring anisotropy changes as a function of time. It also has a sample holder where the temperature is controlled and changed using the Peltier effect. The range of set point temperatures on the Peltier controller is – 20º C to + 100º C.
The instrument also includes the ability to perform fluorescence lifetime measurements. Three LED excitation sources (at 280, 295, and 370 nm) are included; these wavelengths are appropriate for the excitation of tyrosine and/or tryptophan in intrinsic protein fluorescence work (280,295), and, for example, exogenous probes such as acrylodan (370).
The spectrofluorometer has double excitation monochromator for enhancing the spectral purity of the excitation band. PTI double monochromators have a unique ability, and that is that with the appropriate options, it can also function as a single-pass monochromator. The instrument will make use of this capability to its maximum, because it will be supplied with two arc lamp housings, so that users will have the choice (for excitation wavelengths in the 250-600 nm range) of using either single-pass or double-pass excitation (depending on the needs dictated by the sample) without having to move the lamp housing from one double monochromator entrance port to another.
A programmable excitation shutter is part of the spectrofluorometer. This device is useful for minimizing photobleaching between data acquisitions or even between the taking of data points in a long time based scan.
A solid sample holder is also included in the package. With this holder, the user has both translational and rotational control over the positioning of the sample relative to the incoming exciting radiation. Thin films, samples on slides, paper, irregularly-shaped solid samples, as well as cuvettes can be accommodated by the holder. For the study of very turbid solutions, this holder does an excellent job of redirecting scattered exciting light away from the emission axis; this scattered radiation, if not efficiently removed, would contaminate the fluorescence spectrum of the sample with a high, non-constant background.
The spectrofluorometer includes a near-infrared fluorescence option. The main components of this option are the following:
a) a 1200 gr/mm, 600 nm blaze excitation grating (placed in a turret with the second
300 nm blaze grating in the double excitation monochromator;
b) a 600 gr/mm, 1000 nm blaze emission grating (placed in a turret with the 400 nm
blaze grating in one of the emission monochromators;
c) a thermoelectrically-cooled InGaAs detector (spectral range 500-1700 nm);
d) a lock-in amplifier and chopper for noise suppression;
Two common applications where this option finds applicability is in the study of novel materials such as semiconductor nanoparticles (quantum dots) as well as singlet oxygen detection.
The software that comes with the spectrofluorometer also includes a FRET calculator, for the calculation of Rο , the D-A distance, as well as kDA .
Basic QuantaMaster QM-4SE Spectrofluorometer Specifications
Signal-to-noise Ratio: 10,000:1 (based on the Raman spectrum of water, excitation wavelength 350 nm, exc. and em. spectral bandwidth 5 nm, 1 s integration time, no smoothing)
Excitation Source:
Type: continuous Xe arc lamp, 75W
Spectral range: 200-2000 nm (output dependent on monochromator grating)
Monochromators:
Type: Czerny-Turner
Focal Length: 200 mm
f number: 4
Spectral bandwidth: 0 to 25 nm
Resolution: +/- 0.5 nm
Gratings: ruled, 1200 gr/mm, blazed at 300 nm (excitation) and 400 nm (emission)
Detectors:
Standard: R1527 PMT (200-680 nm)
Optional: R928 PMT (200-870 nm), InGaAs photodetector (500-1700 nm)
PMT Detection modes: photon counting, analog (user-switchable)
Reference channel photodiode (calibrated against a standard photodiode), variable-speed magnetic stirrer, filter holders, and lid-activated emission shutter are all standard.
Time-resolved Fluorescence Accessory Specifications
Excitation wavelengths: 280, 295, and 370 nm (others available)
Detection: PTI-patented stroboscopic detector
Timescale protocols: linear, arithmetic, logarithmic (menu-selectable)
Pulse width: 1.5 ns (for 280 and 295 nm LED’s)
Range of Measurable Lifetimes: ~100 ps to >1 μs (LED-dependent)
Acquisition mode: sequential or random
Lifetime Analysis: 1-to-4 exponentials, global analysis, non-exponential, micellar kinetics, lifetime distribution analysis by ESM, lifetime distribution analysis by MEM, anisotropy decays
FRET Calculator: included
For more information go to www.pti-nj.com