Establishment of the optically pumped chemiluminescence technique for diagnostics

Abstract

A new chemiluminescence procedure was established that eliminates the need to use a separate oxidant such as hydrogen peroxide for chemiluminescence detection reactions. In the procedure called optically pumped chemiluminescence (OPC), red light energy Is used by a photosensitive dye to oxidize luminol and generate blue light. By pulsing the excitation light and measuring chemiluminescence during the off time, light scattering background was eliminated and sensitivity enhanced 10-fold. The OPC reaction mechanism was determined to be type I (direct reaction of excited dye with substrate), and the dye catalyst Is recycled In the reaction. Of the popular photosensitive dyes tested, methylene blue showed the best detection limit In the OPC system. A methylene blue dye derivative was synthesized and covalently attached to protein. Antibody proteins were labeled with up to 16 dyes per antibody. An a-fetoproteln Immunoassay was developed having a detection limit of 17 pg. This assay had sensitivity of 1.5 ng of alphafetoproteln/mL of plasma and showed good correlation (/? = 0.98). The OPC catalysts are more stable and less temperature sensitive than enzyme catalysts, and the substrate solution needed Is very stable. Thus, OPC may be valuable In applications where convenience and reagent stability are Important.

Light‐emitting‐diode‐induced chemiluminescence detection for capillary electrophoresis

“…This might also be originated from the rapid decay of the CL signal before detection. The time for RF migrating from irritation window to the detection window is much longer than the half-life time of the CL, about 1 s according to Motsenbocker et al [33].…”

Chemiluminescence from luminol solution after illumination of a 355 nm pulse laser

“…On the other hand, it was found that the type II mechanism could be neglected because there was no CL yielded even at a high concentration of O 2 that could be used to produce a sufficient amount of 1 O 2 . [34][35][36] Then, the PSCL was thought to be caused by a type I mechanism, where an electron was transferred from a luminol molecule to the 3 S*. As a result, a luminol radical and a dye-anion were produced.…”

Enhancers of the non‐peroxidative metal porphyrin chemiluminescence reaction

“…All solutions were prepared in this water unless otherwise specified. A photon-counting luminometer that accepts 12 mm diameter by 75 mm long plastic test tubes was used (8). For each measurement 1OpL of 0 ng/mL or 100 ng/mL porphyrin compound solution were added to 1 mL of luminol solution in a test tube.…”

“…Backgrounds were influenced by ozone in the air and fluctuated according to the use of machinery in the room. Background was lowered by removing oxygen from air that came into contact with the luminol solution during the chemiluminescence measurement (8).…”

“…The enhancement ratio was calculated as described above. A Mn-meso-tetra(4-carboxypheny1)porphine protein conjugate that contained 32 Mn-porphyrins and one bovine serum albumin per antibody fragment was prepared as described elsewhere (8).…”