Method for determining diamine oxidase
Disclosed is a method for determining histaminase (DAO; EC 1.4.3.6) activity in a sample. Said method comprises the following steps: an aqueous solution is supplied which contains a predefined amount of a diamine; the aqueous solution is mixed and incubated with the sample for a given period of time in conditions in which the diamine can be reacted with a DAO possibly present in the sample; the diamine is derivatized; the amount of derivatized diamine is determined; the predefined amount of diamine is compared to the amount of derivatized diamine; and the activity of histaminase possibly present is determined.
1. A method for determining diamine oxidase (DAO) activity in a sample comprising:
providing a known amount of a diamine admixed with the sample;
incubating the diamine under conditions at which the diamine would react with any present DAO in the sample;
derivatizing diamine present after the incubation;
determining an amount of any derivatized diamine; and
comparing the known amount of diamine with the amount of derivatized diamine to determine the amount of diamine that was degraded; and
determining the DAO activity based on the amount of degraded diamine.
2. The method of claim 1 , wherein the diamine is in an aqueous solution or lyophilized.
3. The method of claim 1 , further comprising:
incubating with the diamine and the diamine oxidase a compound potentially inhibiting the activity of diamine oxidase, for a fixed period under conditions at which the diamine can be reacted by a DAO possibly present in the sample;
derivatizing diamine present after the incubation;
determining an amount of derivatized diamine, if any;
comparing the known amount of diamine with the amount of derivatized diamine to determine the activity of the diamine oxidase in the presence of the potentially inhibiting compound; and
comparing the activity of the diamine oxidase with the potentially inhibiting compound with the activity of the diamine oxidase without the potentially inhibiting compound, wherein the potentially inhibiting compound is determined to be a diamine oxidase inhibitor if is lowers the activity of the diamine oxidase by at least 20%.
4. The method of claim 1 , wherein the diamine is histamine, putrescine, spermidine, and/or cadaverine.
5. The method of claim 1 , wherein the aqueous solution comprises a Tris (Tris(hydroxymethyl)-aminomethane) buffer solution having a pH of 7 to 9.5.
6. The method of claim 1 , wherein the aqueous solution contains a diamine in an amount of from 5 to 400 μM.
7. The method of claim 1 , wherein the incubation of the aqueous solution with the sample takes place at a temperature of from 10 to 50° C.
8. The method of claim 1 , wherein the incubation of the aqueous solution with the sample is effected for 30 minutes to 36 hours.
9. The method of claim 1 , wherein incubation of an aqueous solution with the sample is stopped before derivatizing with amino guanidine.
10. The method of claim 1 , wherein derivatizing the diamine comprises acylating.
11. The method of claim 10 , wherein the diamine is acylated with NHS biotin.
12. The method of claim 1 , wherein the amount of derivatized diamine is determined by chromatography, electrophoresis, or immunoassay.
13. The method of claim 12 , wherein the amount of derivatized diamine is determined by high-performance liquid chromatography (HPLC), capillary electrophoresis (CE), enzyme-linked immunosorbent assay (ELISA), or radioimmunoassay (RIA).
14. The method of claim 2 , wherein peroxidase is admixed with the aqueous solution and/or the sample.
15. The method of claim 14 , wherein the peroxidase is catalase.
16. The method of claim 14 , wherein the peroxidase is admixed in an amount of from 0.1 to 10 unit(s) of peroxidase per 100 μl sample.
17. The method of claim 16 , wherein the peroxidase is admixed in an amount of 1 unit of peroxidase per 100 μl sample.