Optical Background Noise Research Articles

Fundamental flow cytometer properties governing sensitivity and resolution

The fundamental properties of a flow cytometer that govern its capacity to detect and resolve dim fluorescent particles are 1) the efficiency with which it can convert a photon emitted by a fluorochrome into a photoelectron at the photocathode of the photomultiplier tube and 2) the amount of optical background noise that is collected along with the fluorescence emission from the particle. Either of these properties alone is insufficient to predict an instrument's performance. Thus, to characterize a flow cytometer, both of these properties need to be determined. To determine these properties, one must not only consider the optical characteristics of the instruments but also understand the impact of the signal processing on the data collected. After the collection efficiency and the optical background noise level are determined, it is possible to predict the instrument's performance. A specific flow cytometer performance level is not unique to only one pair of these properties. To achieve a specific level of instrument performance, whole families of pairs of these properties are possible.

Sensitive absorbance detection method for capillary electrophoresis based on laser wave-mixing

Forward-scattering four-wave mixing is demonstrated as a sensitive absorbance detection method for capillary electrophoresis, using an argon ion laser operating at 457.9 nm. Since this four-wave mixing laser technique utilizes only two input laser beams, it offers important advantages, including ease of optical alignment, high wave-mixing efficiency and low excitation power requirements. In addition, since the analytical signal is a laser-like coherent beam, highly efficient optical signal detection can be performed with minimum optical background noise. Excellent detection sensitivity and short absorption path lengths, and hence, small detector probe volumes, are some of the useful features this absorbance detection method offers for on-column detection of both fluorescing and non-fluorescing analytes in capillary electrophoresis and liquid chromatography. Preliminary “detected” concentration detection limit of 8.5·10−8M, mass detection limit of 13 amol and an absorbance-unit detection limit of 1.35·10−5 AU are determined for dabsyl-glycine using this absorbance detection method.

Doppler-Free Spectroscopy Based on Phase Conjugation by Degenerate Four-Wave Mixing in Hollow Cathode Discharge

Optical phase conjugation in the hollow cathode lamp by degnerate four-wave mixing is a novel analytical laser spectroscopic method in which a laser beam is generated as the signal beam. With the use of a demountable hollow cathode discharge cell and a continuous-wave dye laser, the signal beam is clearly visible to the naked eye when 1 µg of sodium is placed inside the cathode cavity. Since the signal beam is a time-reversed replica of the probe laser beam, highly efficient optical signal detection can be performed with minimum optical background noise. Since the signal is Doppler-free with negligible Lorentzian broadening (30 Torr discharge pressure), it provides excellent spectral resolution. Sodium D2 hyperfine lines of 3s 2S1/2 ( F' = 2) → 3p 2P3/2 ( F = 3) and 3s 2S1/2 ( F' = 1) → 3p 2P3/2 ( F = 0) were resolved. In addition to excellent spectral resolution and ease of signal detection, the technique also offers unique properties of optical phase conjugation.

The quantitatives assessment of fluorescence on thin media chromatograms : A theoretical study

Owing to inherent wavelength conversion in fluorescence analysis, the measuring signal is much less affected by optical background noise than is the case in absorptiometric measurements by both transmittance or reflectance procedures. This explains the higher sensitivity and accuracy obtained in fluorescence analysis. A further advantage of fluorimetry is the much wider range of concentrations over which the response is nearly linear. Measurements from either side of the medium are shown to be nearly equivalent in this respect. More light is, however, available if the measurements are carried out from the illuminated side.

Recent advances in the photodensitometric evaluation of thin-media chromatograms

The article brings a survey of recent developments in theory and instrumentation for the quantitative evaluation of thin-media chromatograms by photodensitometric measurements. Optical and electrical noise as factors limiting the performance of the method are discussed and attention is called to the possible use of semiconductor photodetectors. Double-beam scanning in its various modifications is considered the best way of reducing the effect of the optical background noise and flying spot scanning is shown to remove errors due to irregular geometry and non-uniform density profiles of the zones. The paper ends with the brief description of a newly developed instrument incorporating these features.