Which one of the two common reporter systems is more suitable for chemiluminescent enzyme immunoassay: alkaline phosphatase or horseradish peroxidase?

Abstract

Alkaline phosphatase and horseradish peroxidase are the most commonly used reporter systems in chemiluminescent enzyme immunoassay (CLEIA). Which one, therefore, would be better when establishing a CLEIA method for a new target substance? There was no standard answer. In this study, both reporters were compared systematically including luminescence kinetics, conjugation methods, optimal condition and detection performance, using two common drugs, SD-methoxy-pyrimidine and enrofloxacin, as determination objects. The results revealed that there was much difference between the luminescence kinetics of the two systems. However, there was little difference between these systems when detecting the same substance, including in optimal conditions and determination of performance. Both reporters were suitable for establishing chemiluminescent enzyme immunoassays. Therefore, the choice of alkaline phosphatase or horseradish peroxidase as the reporter system in chemiluminescent enzyme immunoassays depends on availability. Conversely, these two report systems could be applied in simultaneous analysis of multicomponents due to their different optical behaviors and similar performances. But attention should be paid to conjugation method and coating buffer, which affected the luminescent intensity of different determination targets.

Advances in immunoassay‐based strategies for mycotoxin detection in food: From single‐mode immunosensors to dual‐mode immunosensors

“…The most common are enzyme‐based CL sensors, which enzymatically label antigens or antibodies for immune responses (Chen et al., 2017). Among them, HRP, as the natural enzyme, can catalyze the oxidation of the luminol–H 2 O 2 system and detect the target by linking its concentration to the CL intensity of luminol (Yu et al., 2016). In addition, nanozymes and metal NPs including Ag NPs and Au NPs have been found potential roles in the construction of CL immunosensors, which can act as catalysts or energy acceptors to increase the CL signal through surface plasmon resonance (SPR) and enhance the detection sensitivity.…”

Mycotoxins in Pistachios (Pistacia vera L.): Methods for Determination, Occurrence, Decontamination

“…CLEIA (combines chemiluminescence (CL) and enzyme immunoassay) detection techniques are currently the most sensitive in immunoassay research. CLEIA is becoming increasingly popular for the detection of trace compounds due to its great qualities of high specificity, lower limit of detection, good linearity range and environmental friendliness [111]. The main two label enzymes used in CLEIA are horseradish peroxidase (HRP) and alkaline phosphatase (ALP).…”

“…Due to the low cost and the ease of access, horseradish peroxidase is considered the most used. While the luminescence efficiency of the horseradish peroxidase system can be increased by using a suitable enhancer [111], it is quite poor when compared to the ALP system [111]. In any case, CL substrates, such as the luminol/peroxide/enhancer system for horseradish peroxidase (HRP) or dioxetane-based substrates for alkaline phosphatase, can efficiently detect enzyme labels.…”