Methylene-blue-encapsulated liposome for immobilization-free electrochemical immunoassay of interleukin-6 from nervous headache

Interleukin-6 (IL-6) protects neurons by inhibiting the expression of factors related to neuronal injury in nervous headache patients. The development of rapid and sensitive IL-6 detection methods will be very advantageous for easing the pain of such patients. In this work, an immobilization-free immunodetection method is explored for the voltammetric screening of IL-6 in serum samples from nervous headache patients. Initially, methylene-blue-encapsulated liposomes (MBLS) labeled with anti-IL-6 detection antibodies are confined in an anti-IL-6 capture antibody-coated microplate through a sandwich-type immunoreaction, and subsequently subjected to lysis treatment. After that, the lytic solution is transferred into a detection cell including a Nafion-modified working electrode. Methylene blue molecules with positive charge are captured on the negatively charged Nafion membrane, thus generating a voltammetric signal. The voltammetric peak currents are relative to the amount of IL-6 in the solution. Under optimized experimental situations, MBLS-based split-type electrochemical sensing protocols have acceptable voltammetric currents for IL-6 from 0.01 to 100 pg mL⁻¹, and allow screening at a concentration of IL-6 as low as 9.1 fg mL⁻¹. The batch-to-batch coefficients of variation were ≤11.95%. Good anti-interference capability was achieved against other biomolecules. Seven human serum specimens and two diluted serum samples including IL-6 obtained from nervous headache patients were determined by MBLS-based electrochemical immunoassay, and achieved well-matched results in comparison with those of the IL-6 ELISA protocol.