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n enzyme-linked immunosorbent assay, a horseradish peroxidase-catalyzed fluorogenic reaction, and chemiluminescence (CL) analysis have been combined to develop a sandwich ELISA for Staphylococcal enterotoxin B (SEB) using monoclonal antibodies for different epitopes of SEB. The enzyme catalyzed reaction of 3-(4-hydroxyphenyl propionate) with the urea complex of hydrogen peroxide produced a fluorescent dimer which was detected by chemiluminescence analysis. The CL response to SEB is linear in the range from 6.0 to 564 pg mL ( r = 0.9993), and the detection limit is 3.3 pg mL (S/N = 3). Intra- and interassay coefficients of variation are <7.0% at three concentrations (24, 96 and 384 pg mL). The method was applied to the analysis of SEB in serum, lake water and milk samples. The results compared well with those obtained by conventional ELISAs. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

Abstract: A novel chemiluminescence immunoassay was developed, in which two monoclonal antibodies recognised different epitopes of staphylococcal enterotoxin B (SEB) using HRP-functionalised mesoporous silica nanoparticle (MSN) as label. The enzyme-functionalised MSN was fabricated by co-immobilizing HRP and anti-SEB onto the surface of MSN simultaneously using glutaraldehyde as the linkage. Because the large surface area of MSN carriers increased the amount of HRP and the proportion between the enzyme and antibody molecules on the surface of MSN were optimised, the conjugates provided a much higher signal. This is an improvement over the traditional sandwich immunoassay which often has one or two enzyme molecules per antibody. The analysis showed a linear response within the range of 0.01–1ngmL−1 (r =0.9967), and the detection limit (DL) was 4pgmL−1 (3σ). This approach was successfully demonstrated as a simple, cost-effective, sensitive method to detect SEB in milk, water and serum samples. [Copyright &y& Elsevier]