Malaysian
Journal of Analytical Sciences Vol 24 No 2 (2020): 179 - 187
DEVELOPMENT OF AN ENZYME-LINKED
IMMUNOSORBENT ASSAY (ELISA) FOR DOMOIC ACID DETECTION IN SELECTED SHELLFISH
SEAFOOD USING α-AMINO-3-HYDROXY-5-METHYL-4-ISOXAZOLEPROPIONIC ACID (AMPA)
RECEPTOR
(Pembangunan
Imunojerapan Berpaut Enzim Berpesaingan Tak Langsung (ELISA) bagi Pengesanan Domoik Asid
dalam Makanan Laut Bercengkerang Terpilih menggunakan Reseptor
α-Amino-3-Hidroksi-5-Metil-4-Isoksazolepropionik Asid (AMPA))
Hafiza Mohamed Zuki*, Lai Yun Ru, Noor Sheryna Jusoh,
Azrilawani Ahmad, Marinah Mohd Ariffin
Faculty of Science
and Marine Environment,
Universiti
Malaysia Terengganu,
21030 Kuala
Nerus, Terengganu, Malaysia
*Corresponding author: hafiza@umt.edu.my
Received: 20 November 2019;
Accepted: 12 March 2020
Abstract
In this study, a selective and sensitive
enzyme-linked immunosorbent assay (ELISA) method using ionotropic glutamate
receptor of amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) was
successfully developed in order to determine the presence of domoic acid (DA)
in shellfish seafood. The optimum concentration of AMPA receptor and goat
anti-mouse IgG alkaline phosphate (AP) were obtained at 1.3 ppm and 1.7 ppm,
respectively. Absorbance response of the optimised assay towards DA recorded
limit of detection (LOD) and limit of quantification (LOQ) at 0.72 ppm and 1.2
ppm, respectively. Cross-reaction with glutamic acid (GA) showed that the assay
was less sensitive than DA, with cross-reactivity values of 12.96%. The
competency of the immunoassay towards real samples analysis was studied using mussel
and cockle. Detection of non-spiked samples recorded the presence of DA at 0.96
ppm for mussel and 0.56 ppm for cockle. Meanwhile, recovery of the spike samples
with 3 ppm DA was found at 2.69 ppm for mussel and 2.60 ppm for cockle, with
recovery percentages of 86.7% and 89.7%, respectively.
Keywords: enzyme-linked
immunosorbent assay, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid,
domoic acid, shellfish seafood
Abstrak
Dalam kajian
ini, satu kaedah imunojerapan berpaut enzim berpesaingan tak langsung (ELISA)
yang selektif dan sensitif dengan menggunakan reseptor glutamat ionotropik asid
α-amino-3-hidroksi-5-metil-4-isoksazolepropionik (AMPA) telah berjaya dibangunkan
untuk menentukan kehadiran asid domoik (DA) dalam makanan laut bercengkerang.
Kepekatan optimum reseptor AMPA dan anti-tikus kambing IgG alkali fosfat (AP) masing-masing
adalah didapati pada 1.3 ppm dan 1.7 ppm. Tindak balas penyerapan terhadap
ujian optimum DA mencatatkan had pengesanan (LOD) dan had kuantifikasi (LOQ) masing-masing
diperolehi sebanyak 0.72 ppm dan 1.2 ppm. Tindak balas silang dengan asid
glutamat (GA) menunjukkan bahawa ujian GA adalah kurang sensitif daripada DA,
dengan nilai tindak balas-silang 12.96%. Kecekapan imunoasai bagi analisis
sampel sebenar telah dikaji menggunakan kupang dan kerang. Pengesanan sampel
yang tidak dipaku mencatatkan kehadiran DA pada 0.96 ppm untuk kupang dan 0.56
ppm untuk kerang. Sementara itu, perolehan semula sampel yang dipaku dengan 3
ppm DA diperolehi pada 2.69 ppm bagi kupang dan 2.6 ppm bagi kerang, dengan
peratusan pemulihan masing-masing sebanyak 86.7% dan 89.7%.
Kata kunci: ujian imunojerapan berpaut enzim
berpesaingan tak langsung, asid α-mino-3-hidroksi-5-metil-4-isoksazolepropionik,
asid domoik, makanan laut bercengkerang
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