title: Enhancement of enzyme-activated 1,2-dioxetane chemiluminescence in membrane-based assays.
abstract: Processes for enhancing the chemiluminescent signal generated by enzyme-activated 1,2-dioxetanes have been developed for membrane-based assays in which detection is done using a charge-coupled device (CCD) camera or X-ray film. The enhancement is demonstrated using slot-blots of biotinylated lambda DNA in conjunction with an avidin-alkaline phosphatase conjugate. For detection with a CCD camera, the nylon membrane is dried after processing and incubation in dioxetane substrate solution and heated to temperatures of 50 to 80 degrees C during detection. Up to a 100-fold signal increase is obtained using this enhancement process compared to the conventional detection procedure, in which the blot is kept saturated with substrate solution in a sealed plastic bag during detection. For detection with X-ray film, a fivefold increase in signal intensity is realized by drying the membrane before exposure to the film. These enhancement processes greatly reduce the time required for detection in membrane-based assays.
authors.affiliation: Corporate New Business Development Division, DuPont Company, Incorporated, Wilmington, Delaware 19880.
authors.forename: R K
authors.initials: RK
authors.lastname: Kobos
authors.forename: B A
authors.initials: BA
authors.lastname: Blue
authors.forename: C W
authors.initials: CW
authors.lastname: Robertson
authors.forename: L A
authors.initials: LA
authors.lastname: Kielhorn
mesh_terms: D006571:Heterocyclic Compounds / Q000737:chemistry*; D006573:Heterocyclic Compounds, 1-Ring; D008163:Luminescent Measurements; D012815:Signal Processing, Computer-Assisted; D013696:Temperature; D014962:X-Ray Film
chemical_list: D006571:Heterocyclic Compounds; D006573:Heterocyclic Compounds, 1-Ring; C079142:1,2-dioxetane