경쟁적 ELISA를 이용한 도파민과 세로토닌의 검출 = Detection of Dopamine and Serotonin by Competitive Enzyme-Linked Immunosorbent Assay

다국어 초록 (Multilingual Abstract)

Dopamine (DA) and serotonin (5-Hydroxytryptamine, 5-HT) are neurotransmitters and hormones that exist in small amounts but have important role in the body. Serum and 24-hour urine are used as specimens, and are usually examined by HPLC-MS. In this study, we tried to detect DA and 5-HT by competitive ELISA using antigen-antibody (Ab) reaction. After immobilizing $5{\mu}g/mL$ BSA conjugate on a 96-well surface, hormone and primary Ab, which are respectively diluted to different concentrations, were treated. Then, HRP-conjugated secondary Ab and TMB were added to measure absorbance. The regression equation and $R^2$ value were calculated based on absorbance, and sensitivity of Ab to hormone as well as the correlation between hormone concentration and absorbance were determined. In DA ELISA, $R^2$, the correlation between the concentration of hormone and absorbance, was the highest by 0.91 when anti-dopamine Ab was diluted 6,000 times and 7,000 times. In 5-HT ELISA, $R^2$ was bigger than 0.90 in every concentration except 3,000 times and 6,000 times. Both DA and 5-HT were not effectively detected at low concentrations (less than $1.0{\times}10^{-7}M$); and because reference value of serum DA is lower than this, HPLC-MS was required to detect serum DA. However, competitive ELISA may be effective in detecting 24-hour urine DA, serum, and 24-hour 5-HT. Further studies are needed to detect hormones more accurately at lower concentrations.

참고문헌 (Reference)

1 Raoof JB, "Voltammetric determination of ascorbic acid and dopamine in the same sample at the surface of a carbon paste electrode modified with polypyrrole/ferrocyanide films" 50 (50): 4694-4698, 2005

2 Pussard E, "Validation of HPLC-amperometric detection to measure serotonin in plasma, platelets, whole blood, and urine" 42 (42): 1086-1091, 1996

3 Berger M, "The expanded biology of serotonin" 60 : 355-366, 2009

4 Huisman H, "Studies on the immune response and preparation of antibodies against a large panel of conjugated neurotransmitters and biogenic amines: Specific polyclonal antibody response and tolerance" 112 (112): 829-841, 2010

5 Safavi A, "Simultaneous determination of dopamine, ascorbic acid, and uric acid using carbon ionic liquid electrode" 359 (359): 224-229, 2006

6 Ramesh P, "Selective determination of dopamine using unmodified, exfoliated graphite electrodes" 561 (561): 173-180, 2004

7 Belin GK, "Rapid analysis of serotonin and propranolol using miniaturized CE with deep-UV fluorescence detector" 30 (30): 2565-2571, 2009

8 The National Collaborating Centre for Chronic Conditions, "Parkinson's Disease" Royal College of Physicians 59-100, 2006

9 González-Flores D, "Ingestion of Japanese plums (Prunus salicina Lindl. cv. Crimson Globe) increases the urinary 6-sulfatoxymelatonin and total antioxidant capacity levels in young, middle-aged and elderly humans: Nutritional and functional characterization of their content" 50 (50): 229-236, 2011

10 이인희, "In Vitro에서 히스타민 측정 시 ELISA법과 HPLC-MS 분석법의 비교" 대한임상검사과학회 47 (47): 306-312, 2015

11 Neumeister A, "Implications of genetic research on the role of the serotonin in depression: emphasis on the serotonin type 1A receptor and the serotonin transporter" 174 (174): 512-524, 2004

12 Siddiqi HA, "Henry's Clinical diagnosis and management by laboratory methods" Elsevier 2017

13 Gardner D, "Greenspan’s Basic & clinical endocrinology" The McGraw-Hill Companies 2011

14 Kim JS, "Competitive enzyme-linked immunosorbent assay for the determination of catecholamine, dopamine in serum" 619 (619): 87-93, 2008

15 Goldberg L, "Cardiovascular and renal actions of dopamine:Potential clinical applications" 24 (24): 1-29, 1972

16 Bernheimer H, "Brain dopamine and the syndromes of Parkinson and Huntington clinical, morphological and neurochemical correlations" 20 (20): 415-455, 1973

17 Carrera V, "A simple and rapid HPLC-MS method for the simultaneous determination of epinephrine, norepinephrine, dopamine and 5-hydroxytryptamine:Application to the secretion of bovine chromaffin cell cultures" 847 (847): 88-94, 2007