Immunochemical assay with monoclonal antibodies for detection of staphylococcal enterotoxin H

Natalia Vasilyevna Rudenko ^b,c, Anna Petrovna Karatovskaya ^b,c,Anatolyi Nikolaevich Noskov ^d, Anna Olegovna Shepelyakovskaya ^a,b,Margarita Pavlovna Shchannikova ^a,b,c,

Irina Vladimirovna Loskutova ^a,b,c, Olga Anatolievna Artyemieva ^a,Daria Alexandrovna Nikanova ^a, Elena Alexandrovna Gladyr ^a,Fedor Alexandrovich Brovko ^a,b,c,*

^a Ernst All-Russian Research Institute of Animal Husbandry, Dubrovitsy, Moscow Region 142132, Russia

^b Pushchino Branch, ShemyakineOvchinnikov Institute of Bioorganic Chemistry, 6 Prospekt Nauki, Pushchino, Moscow Region 142290, Russia

^c Pushchino State Institute of Natural Science, 3 Prospekt Nauki, Pushchino, Moscow Region 142290, Russia

^d Gamaleya Federal Research Center of Epidemiology and Microbiology, 18 Gamaleya Street, Moscow 123098, Russia

Staphylococcal enterotoxins cause food poisoning of various degrees of severity. For milk and meat products, there is a high probability of contamination with staphylococcal enterotoxin H (SEH). In this regard specific and sensitive methods are required to be developed for its detection and monitoring. In this work, the gene seh was expressed and a preparation of recombinant toxin was obtained. Using hybridoma technology, a panel of high-affinity monoclonal antibodies (mAbs) to SEH was produced. The antibodies were characterized and shown to have no cross-reactivity towards the main staphylococcal enterotoxins (A, B, C1, D, E, G and I). Based on these mAbs, a method for specific and quantitative detection of SEH was developed in the format of sandwich enzyme immunoassay (linear range, 0.2–3 ng/ml). All the mAbs produced revealed SEH by immunoblotting. Immunochemical analysis of the culture fluids of staphylococcal isolates obtained from the milk of mastitis-infected cows by immunoblotting and sandwich enzyme immunoassay demonstrated the conformity of these methods. Using the developed method, the toxin was revealed in blood serum and liquid food products practically to 100%. From non-liquid foods, it was shown to be extracted to a maximum with a buffer of pH 4.0–4.5.

Keywords: Staphylococcus aureus, Staphylococcal enterotoxins, Staphylococcal enterotoxin H, Monoclonal antibodies, Sandwich enzyme immunoassay