Chemiluminescent
Labels Released from Long Spacer
Arm-Functionalized Magnetic Particles: A Novel Strategy for Ultrasensitive
and Highly Selective Detection of Pathogen Infections
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Abstract
Previously, the unique advantages
provided by chemiluminescence
(CL) and magnetic particles (MPs) have resulted in the development
of many useful nucleic acid detection methods. CL is highly sensitive,
but when applied to MPs, its intensity is limited by the inner filter-like
effect arising from excess dark MPs. Herein, we describe a modified
strategy whereby CL labels are released from MPs to eliminate this
negative effect. This approach relies on (1) the magnetic capture
of target molecules on long spacer arm-functionalized magnetic particles
(LSA-MPs), (2) the conjugation of streptavidin-alkaline phosphatase
(SA-AP) to biotinylated amplicons of target pathogens, (3) the release
of CL labels (specifically, AP tags), and (4) the detection of the
released labels. CL labels were released from LSA-MPs through LSA
ultrasonication or DNA enzymolysis, which proved to be the superior
method. In contrast to conventional MPs, LSA-MPs exhibited significantly
improved CL detection, because of the introduction of LSA, which was
made of water-soluble carboxymethylated β-1,3-glucan. Detection
of hepatitis B virus with this technique revealed a low detection
limit of 50 fM, high selectivity, and excellent reproducibility. Thus,
this approach may hold great potential for early stage clinical diagnosis
of infectious diseases