Method of time-of-flight mass spectrometry with laser desorption/ionization (MALDI-TOF) for studying the protein profile of helminths of commercial fish

The purpose of the research is to develop a method of sample preparation using reagents and materials of Russian production to study the protein profile of sanitary-significant helminths of commercial fish using the method of time-offlight mass spectrometry with laser desorption/ionization (MALDI-TOF).

Materials and methods. 98 specimens of helminths from three fish species were selected for the study: adult cestodes Triaenophorus nodulosus from the intestines of a pike (Esox lucius), plerocercoids cestodes Diphyllobothrium ditremum from the wall of the stomach and intestines of European vendace (Coregonus albula), nematode larvae Eustrongylides excisus from muscle tissue and liver of Common perch (Perca fluviatilis). Ichthyological material was obtained using passive fishing gear (nets) in the period from January to August 2025 and delivered to the laboratory in frozen form. In the laboratory, fish were dissected and muscle tissue and internal organs were examined under binocular microscope Motic SMZ-171T. For species identification of parasites, the Identifier of Parasites of Freshwater Fish Fauna of the USSR Gusev (1985), Аvdeev (1987) was used. Photo documentation was performed using the microscope Zeiss AxioImager Z.1. Sample preparation (extraction) of helminths was carried out according to the patent N RU 2768162 C1 Method for preparing adult nematode specimens for identification using MALDI TOFF mass spectrometry method (2022). Mass spectrometry of helminths was performed on a mass spectrometer MALDI (Microflex, Bruker Germany).

Results and discussion. Currently, in the Russian Federation, there is a need to develop, verify, and implement into practical activities of testing laboratories the working protocols of the MALDI-TOF MS method for diagnosing infections of fish used for food purposes, and to create a unified reference library of the spectra of helminth proteins found in Russia. Also, a standard operating protocol for diagnostic testing of helminths using time-of-flight mass spectrometry has not been developed. We conducted research to refine a method for sample preparation (extraction) of helminths using Russianmade consumables and reagents. As a result of our research, we developed a sample preparation protocol and obtained 41 mass spectra.

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