Monoamine oxidase A&B detection -

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Fluoro MAO
Fluorescent Monoamine Oxidase A&B Detection Kit

Key Benefits:

Non Radioactive
Can monitor multiple time points to follow kinetics.
One-step, no wash assay.
Adaptable for High Throughput format
Sensitive
Applications - Standard Fluorescence Plate Reader

Assay Principle

Monoamine oxidase (MAO) is a flavin-containing enzyme that catalyses the oxidation of a variety of amine-containing neurotransmitters such as serotonin, norepinephrine, epinephrine and dopamine to yield the corresponding aldehydes (1). MAO exists in two isoforms, namely MAO-A and MAO-B, which are the products of two distinct genes (2).

MAO-A and B exhibit different specificities to substrates and inhibitor selectivities. Extensive studies have been preformed to characterize their properties (3-7). MAO-A acts preferentially on serotonin and norepinephrine, and is inhibited by clorgyline. MAO-B acts preferentially on 2-phenylethylamine and benzylamine and is inhibited by deprenyl and pargyline.

Localized in the outer mitochondrial membrane, these enzymes are found throughout the body. Often only one form of the enzyme is present in a specific organ and/or within a specific cell type (8-9). In addition to their role in regulating neurotransmitters, these enzymes are also involved in processing biogenic amines (10) including tyramine (11).

The Fluoro MAO-A/B detection kit utilizes a non - fluorescent substrate, 10-Acetyl-3, 7-dihydroxyphenoxazine (ADHP) to detect H₂0₂ released from the conversion of a substrate to its aldehyde via MAO-A/B. Furthermore, H₂0₂ oxidizes ADHP in a 1:1 stoichiometry to produce a fluorescent product resorufin. This oxidation is catalyzed by Peroxidase.

Reaction:

Substrate + O2 + H₂0₂ + MAO-A/B ----------> aldehyde + NH3 + H₂0₂

H₂0₂ + Detection reagent (non-fluorescent) + Peroxidase ---------> Resorufin (fluorescent)

Excitation 530-570. Emission 590-600nm

The fluorescent monoamine oxidase detection kit can be used to monitor MAO activity and screen of MAO inhibitors.

Kit contents (for 500 assays)
Description	Part#	Storage after opening Kit or Reconstitution of Reagents.
Bottle: 5X Reaction Buffer pH 7.4	3020	4-8⁰C
1 vial: Detection reagent	4009	Aliquot in single use vials: Below –20⁰C
1 vial: Horseradish Peroxidase	6007	4-8⁰C
1 vial: MAO-B substrate Benzylamine	7006	Aliquot in vials: Below –20⁰C
1 vial: MAO-A/B substrate: Tyramine	7005	Aliquot in single use vials: Below –20⁰C
1 vial Pargyline: Monoamine Oxidase B inhibitor ^(12-14)	7003	Aliquot in vials: Below –20⁰C
1 vial Clorgyline: Monoamine Oxidase A inhibitor^(12-14)	7002	Aliquot in single use vials: Below –20⁰C

The following kits are available:
Catalog	Size	Price (Eur)
FLMAO 100-3	500 Tests	€245

Figure1. Recombinant MAO-B was serially diluted in 1X Reaction Buffer. 100 mL of diluted MAO-B per well was mixed with 100mL of Reaction Cocktail. The reaction was incubated at Room Temperature in the dark for 1 hour. Fluorescence was measured with a microplate reader using excitation at 530mM and fluorescence emission at 590mM.