Monoclonal Antibody-Based Sandwich ELISA for the Detection of Staphylococcal Enterotoxin A

Hua Kuang,Chuanlai Xu,Wenbing Wang,Wei Ma,Liguang Xu,Liqiang Liu,Libing Wang

doi:10.3390/ijerph10041598

Abstract

A sensitive and specific monoclonal antibody-based sandwich enzyme-linked immunosorbent assay (ELISA) was established and validated for the detection of staphylococcal enterotoxin A (SEA). After routine fusion and selection, 10 monoclonal antibodies showed high affinity for SEA. An optimal pair for sandwich ELISA was selected by pairwise interaction analysis. After optimization, the limit of detection (LOD) and linear dynamic range of the method were established, and were found to be 0.0282 ng/mL and 0.06–2 ng/mL, respectively. The recovery in pure milk ranged from 82.67% to 111.95% and the intra- and inter-assay coefficients of variation ranged from 3.16% to 6.05% and from 5.16% to 10.79%, respectively. Cross-reactivity with staphylococcal enterotoxin B (SEB), staphylococcal enterotoxin C (SEC), staphylococcal enterotoxin D (SED), and staphylococcal enterotoxin E (SEE) in this method were insignificant. These results indicate that the sandwich ELISA method developed in our study is effective for routine identification of SEA in food samples.

Highlights

Staphylococcus aureus, a well-known food-borne pathogen, causes staphylococcal food poisoning (SFP) by producing enterotoxins in food such as meats and dairy products [1,2]
Seven days after cell fusion, the monoclonal antibodies (mAbs) anti-staphylococcal enterotoxin A (SEA) in the supernatant fluid from the cell culture plates was detected by indirect enzyme-linked immunosorbent assay (ELISA)
10 hybridoma cell lines designated as mAb1 to mAb10 and showing high affinity for SEA were obtained

Summary

Introduction

Staphylococcus aureus, a well-known food-borne pathogen, causes staphylococcal food poisoning (SFP) by producing enterotoxins in food such as meats and dairy products [1,2]. Staphylococcal enterotoxins (SEs) are single-chain proteins with molecular weights ranging from 27 to 34 kDa [3]. The actual toxic dose of SEs is still not very clear, it was reported that 200 ng or less of SEA can produce illness in sensitive individuals [6]. Research on some SFP outbreaks found that SEA was detected at a concentration around 0.5 ng/mL [6,7]. Highly sensitive and reliable methods for analyzing SEs in food samples are critical for monitoring food products and traceback investigations in SFP outbreaks

Methods

Results

Conclusion