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R S Handley

Publications and source records attributed to R S Handley.

18 recordsLinked to original sources
PubMed1999

Sequential chemiluminescent detection of target DNAs without stripping and reprobing.

We present a simple method for sequential chemiluminescent detections of two different DNA loci on a single Southern blot. First, an enzyme-linked DNA probe for a unique sequence is detected with a horse-radish peroxidase (HRP) substrate followed by the detection of another enzyme-linked DNA probe for a different unique sequence with an alkaline phosphatase (AP) substrate that simultaneously inhibits the chemiluminescence generated by HRP. Such sequential detection steps eliminate the need to strip and reprobe blots and can be performed with no intervening steps.

Alkaline Phosphatase↗
PubMed1992

A rapid and simple chemiluminescent assay for Escherichia coli beta-galactosidase.

We describe a chemiluminescent assay for E. coli beta-galactosidase using Lumi-Gal 530, a commercial formulation containing a stable phenylgalactose-substituted dioxetane as the substrate. Removal of the galactose moiety leads to the generation of an unstable dioxetane which decomposes to provide the observed chemiluminescence which is measured with a luminometer. Advantages of the assay are that it is simple, inexpensive and has 20-fold greater sensitivity than the standard spectrophotometric assay. Additional advantages are that the dioxetane is quite stable in the commercial formulation, and beta-galactosidase functions efficiently and is not degraded during the course of an assay. As luminometers are becoming commonplace in molecular biology laboratories, this assay provides a preferable alternative to the spectrophotometric assay.

Biotechnology↗
PubMedDate not supplied

CCD camera imaging for the chemiluminescent detection of enzymes using new ultrasensitive reagents.

We have designed and constructed an inexpensive imaging system based on charge-coupled device (CCD) technology and utilized it to demonstrate the sensitivity and rapid detection possible with Lumigen chemiluminescent reagents. We also report the development of two new chemiluminescent reagents, Lumi-Phos Plus and Lumigen PS. Lumi-Phos Plus is an enhanced formulation for the rapid detection of alkaline phosphatase on membranes and in solution. It provides excellent images in blotting applications with exposures of under a minute. Lumigen PS represents a new generation of peroxidase detection reagents. The wide dynamic range with excellent linearity, higher signal and lower background than other chemiluminescent reagents make Lumigen PS of unsurpassed value in enzyme-linked immunoassays and nucleic acid probe assays using HRP conjugates.

Alkaline Phosphatase↗
PubMedDate not supplied

Chemiluminescent acridan phosphate labelling compounds for detection in gels.

We have developed a chemiluminescent acridan phosphate labelling compound which produces a flash of light upon chemical triggering. Sequential treatment of the label with acid and a strong base, one of which contains a peroxide, initiates light emission as a rapid flash lasting 1--2 s. Labelling of analytes with these compounds permits their direct detection in a non-enzymatic assay format. Representative compounds have been linked to BSA as a model protein analyte and detected by chemiluminescence assay. Light intensity correlated with the amount of BSA over six orders of magnitude, permitting the detection of 50 amol of protein in solution. We have also demonstrated for the first time that labelled proteins can be separated by electrophoresis without destruction of the label and that chemiluminescence can be produced and detected directly in the gel. The ease, speed and sensitivity of detection by this new method should enable the development of simpler assays for a wide variety of analytes.

Acridines↗