Tropomyosin analysis in foods using an electrochemical immunosensing approach

Allergies to seafood have a relevant impact on public health since the unknown intake of products thereof can cause serious health problems. Tropomyosin (TPM) has been identified as a major shellfish allergen. Crustaceans, especially shrimps, are of particular concern because of the presence of TPM in their muscle tissue. Although attempts have been made to reduce its allergenicity by boiling, this does not guarantee the total absence of TPM [1]. So, the detection of trace levels of TPM in food products can prevent health problems. For this purpose, electrochemical immunosensors are adequate analytical tools since they provide highly selective, sensitive, fast, and cheap analysis and are suitable for in situ applications.

In this work, a simple voltammetric immunosensor for the determination of TPM in commercial food products was developed. A sandwich-type immunoassay was performed on screen-printed carbon electrodes. Monoclonal and polyclonal antibodies were used to capture and detect TPM. To obtain the analytical signal an alkaline phosphatase-labelled secondary antibody and 3-indoxyl phosphate / silver nitrate (enzymatic substrate) were employed; the enzymatically deposited silver was analysed by linear sweep voltammetry. Using a factorial design, several experimental parameters were optimized: capture antibody 20 µg mL^-1; secondary antibody dilution 1:2,000; alkaline phosphatase-labelled detection antibody dilution 1:40,000; and an assay time of ~3 h. A linear concentration range was established between 2.5 and 20 ng mL^-1 (i_p (µA) = 0,787 × [TPM] (ng mL^-1) + 5,45, n = 5, r = 0,990, V_x0 = 8.8 %) and the LOD was 1.7 ng mL^-1. These performance characteristics will allow the quantification of trace levels of the target allergen in food products.

[1] Laly, S.J, Sankar, T.V., Panda, S.K., Effect of pressure cooking alone and in combination with other treatments on shrimp allergic protein, tropomyosin. Journal of Food Science and Technology (2021). https://doi.org/10.1007/s13197-021-05124-2