A spectrofluorometer is an instrument which takes advantage of fluorescent properties of some compounds in order to provide information regarding their concentration and chemical environment in a sample. A certain excitation wavelength is selected, and the emission is observed either at a single wavelength, or a scan is performed to record the intensity versus wavelength, also called an

emission spectrum The emission spectrum of a chemical element or chemical compound is the Spectrum (physical sciences), spectrum of frequencies of electromagnetic radiation emitted due to electrons making a atomic electron transition, transition from a high energ ...

. The instrument is used in

fluorescence spectroscopy Fluorescence spectroscopy (also known as fluorimetry or spectrofluorometry) is a type of electromagnetic spectroscopy that analyzes fluorescence from a sample. It involves using a beam of light, usually ultraviolet light, that excites the electro ...

Operation

Generally, spectrofluorometers use high intensity light sources to bombard a sample with as many photons as possible. This allows for the maximum number of molecules to be in an

excited state In quantum mechanics Quantum mechanics is the fundamental physical Scientific theory, theory that describes the behavior of matter and of light; its unusual characteristics typically occur at and below the scale of atoms. Reprinted, Add ...

at any one point in time. The light is either passed through a filter, selecting a fixed wavelength, or a

monochromator A monochromator is an optics, optical device that transmits a mechanically selectable narrow band of wavelengths of light or other radiation chosen from a wider range of wavelengths available at the input. The name is . Uses A device that can ...

, which allows a wavelength of interest to be selected for use as the exciting light. The emission is collected at the perpendicular to the emitted light. The emission is also either passed through a filter or a monochromator before being detected by a

photomultiplier tube Photomultiplier tubes (photomultipliers or PMTs for short) are extremely sensitive detectors of light in the ultraviolet, visible light, visible, and near-infrared ranges of the electromagnetic spectrum. They are members of the class of vacuum t ...

photodiode A photodiode is a semiconductor diode sensitive to photon radiation, such as visible light, infrared or ultraviolet radiation, X-rays and gamma rays. It produces an electrical current when it absorbs photons. This can be used for detection and me ...

, or

charge-coupled device A charge-coupled device (CCD) is an integrated circuit containing an array of linked, or coupled, capacitors. Under the control of an external circuit, each capacitor can transfer its electric charge to a neighboring capacitor. CCD sensors are a ...

detector. The signal can either be processed as digital or analog output. Systems vary greatly and a number of considerations affect the choice. The first is the signal-to-noise ratio. There are many ways to look at the signal to noise of a given system but the accepted standard is by using the Raman signal of water. Sensitivity or

detection limit The limit of detection (LOD or LoD) is the lowest signal, or the lowest corresponding quantity to be determined (or extracted) from the signal, that can be observed with a sufficient degree of confidence or statistical significance. However, the ...

is another specification to be considered, that is how little light can be measured. The standard would be

fluorescein Fluorescein is an organic compound and dye based on the xanthene tricyclic structural motif, formally belonging to Triarylmethane dye, triarylmethine dyes family. It is available as a dark orange/red powder slightly soluble in water and alcohol. ...

NaOH Sodium hydroxide, also known as lye and caustic soda, is an inorganic compound with the formula . It is a white solid ionic compound consisting of sodium cations and hydroxide anions . Sodium hydroxide is a highly corrosive base and alkali t ...

, typical values for a high end instrument are in the femtomolar range.

Auxiliary components

These systems come with many options, including: *Polarizers *Peltier temperature controllers *Cryostats *Cold Finger Dewars *Pulsed lasers for lifetime measurements *LEDs for lifetimes *Filter holders *Adjustable optics (very important) *Solid sample holders *Slide holders *Integrating spheres *Near-infrared detectors *Bilateral slits *Manual slits *Computer controlled slits *Fast switching monochromators *Filter wheels

References

{{reflist Laboratory equipment Spectrometers