Sensors & Transducers (Apr 2019)
A Lateral Flow Immunoassay Kit for the Detection of Tropical House Dust Mite Blomia Tropicalis
Abstract
IgE-mediated allergic reactions such as asthma, atopic dermatitis and allergic rhinitis are triggered by allergens such as those from the house dust mite Blomia tropicalis (Bt). Detection of allergens is crucial in the management and prevention of allergic diseases. The study aimed to provide a proof of concept for a sandwich-format lateral flow immunoassay (LFIA) kit for the detection of Bt-specific proteins using gold nanoparticle-labeled polyclonal antibodies. Anti-Bt polyclonal antibodies produced from rabbits were purified using Protein A chromatography, and were labeled with gold nanoparticles (AuNP) synthesized using the citrate reduction method. These were then dispensed on glass fiber pad that serves as the conjugate pad. Test and control lines were established through the immobilization on nitrocellulose membrane pad of the unlabeled antibodies and anti-rabbit IgG respectively. The LFIA kit was constructed from the assembly of cellulose fiber pad as the sample pad, conjugate pad, and the nitrocellulose membrane pad. The anti-Bt polyclonal antibody concentration was 0.99 mg/mL upon purification and pre-concentration. Gold nanoparticles with 21.2 ± 1.0 nm diameter as observed under transmission electron microscope, were used for antibody conjugation at an optimal pH of 8.5 (borate buffer) and an optimal ratio of 10 µL 50 µg/mL antibody:100 µL AuNP. Optimal conditions such as color intensity and fastest migration time were observed with the treatment of 0.05 % Tween20 and 10 % sucrose in the conjugate pads; 5 % bovine serum albumin (BSA) and 0.05 % Tween20 in the sample pads, and 1 % BSA in the test pads. The limit of detection of the Bt LFIA is 0.054 µg/mL. This study provides a prototype for a low-cost, rapid, and equipment-free detection of Blomia tropicalis-specific proteins.
