Haptoglobin Electrochemical Diagnostic Method for Subclinical Mastitis Detection in Bovine Milk

This work proposes an outstanding screening magneto-bioassay for the early identification of bovine subclinical mastitis. Haptoglobin (Hp) was used as a promising biomarker for the diagnosis of subclinical mastitis. To this end, novel chitosan-modified magnetic particles, coated with haemoglobin-modified polyaniline (MNPs@Chi/PANI-Hb), have been integrated in a sensitive electrochemical Hp bioassay. Haemoglobin was used as bioreceptor due to its high affinity against Hp. The appropriateness of their synthesis and their modifications were demonstrated by XRD, FT-IR spectroscopy, thermogravimetry and N-2 adsorption-desorption analyses. After the optimization of the experimental parameters the main analytical features were obtained showing excellent performance. The electrochemical biosensor in milk matrix presented a dynamic range of 0.001 to 0.32 & mu;g mL(-1) with a detection limit of 0.031 & mu;g mL(-1), a value much lower than the normal Hp values in bovine milk, making it highly suitable for such determinations. Finally, real milk samples, obtained from local dairy farmers, were analysed by the electrochemical bioassay and compared against a commercial ELISA kit for Hp detection. Milk samples were correctly classified as "acceptable" or "unacceptable" milk considering the Hp concentration obtained with the proposed bioassay, confirming its excellent predictive capacity for subclinical mastitis diagnosis.