The design and application of an automated luminometer for chemiluminescence

An automated luminometer was designed and constructed to facilitate the investigation of reaction conditions and quantitation of the chemiluminescence observed during the oxidation of NADH by horseradish peroxidase in the presence of eosin. The luminometer design incorporates a computer controlled reagent induction system, pulse counting detector and real time display of luminescence profiles. Each of the major reaction conditions was studied individually to determine the most favorable conditions for the analytical application of this reaction. In addition, this chemiluminescent reaction was studied to determine if the luminescing species is recycled. If this were the case it would provide the first essential step necessary for the future design of a chemiluminescent probe that could be reusable or be used for continuous monitoring. During the course of this investigation it was observed that the luminescing species, eosin, was being bleached. A number of experiments were undertaken to determine the nature of the bleaching process. Those steps which suppressed the bleaching process, unfortunately, also suppressed the emission process. Despite the fact that this reaction does not appear to recycle the luminescing species, it does provide a chemiluminescent method for the determination of NADH over the range 5x10⁻⁶ M to 5x10⁻⁴ M. This chemiluminescent reaction has also been coupled to several enzyme systems, which reduce NAD⁺ to NADH, in order to obtain a chemiluminescent signal proportional to the substrate concentration.

Name thesis:degree_name

Ph. D.

Level thesis:degree_level

doctoral

Discipline thesis:degree_discipline

Chemistry

Department dc:contributor.department

Chemistry

Grantor dc:publisher

Virginia Polytechnic Institute and State University

Year dc:date.issued

1984

Author dc:creator

• Chapple, Ian

Chair dc:contributor.committeechair

• Dessy, Raymond E.

Committee members dc:contributor.committeemember

• McNair, Harold M.
• Mason, John G.
• Wolf, J.F.
• McGrath, James E.

Statement dc:rights

• In Copyright

Licence dc:rights.uri

• http://rightsstatements.org/vocab/InC/1.0/

Language dc:language.iso

en_US

Handle dc:identifier.uri

http://hdl.handle.net/10919/74692

OAI identifier oai:identifier

oai:vtechworks.lib.vt.edu:10919/74692