Microfluidic heavy metal immunoassay based on absorbance measurement

A simple and rapid flow-based multioperation immunoassay for heavy metals using a microfluidic device was developed. The antigen-immobilized microparticles in a sub-channel were introduced as the solid phase into a main channel structures through a channel flow mechanism and packed into a detection area enclosed by dam-like structures in the microfluidic device. A mixture of a heavy metal and a gold nanoparticle-labeled antibody was made to flow toward the corresponding metal through the main channel and make brief contact with the solid phase. A small portion of the free antibody was captured and accumulated on the packed solid phase. The measured absorbance of the gold label was proportional to the free antibody portion and, thus, to the metal concentration. Each of the monoclonal antibodies specific for cadmium-EDTA, chromium-EDTA, or lead-DTPA was applied to the single-channel microfluidic device. Under optimized conditions of flow rate, volume, and antibody concentration, the theoretical (antibody K(d)-limited) detection levels of the three heavy metal species were achieved within only 7 min. The dynamic range for cadmium, chromium, and lead was 0.57-60.06 ppb, 0.03-0.97 ppb, and 0.04-5.28 ppb, respectively. An integrated microchannel device for simultaneous multiflow was also successfully developed and evaluated. The multiplex cadmium immunoassay of four samples was completed within 8 min for a dynamic range of 0.42-37.48 ppb. Present microfluidic heavy metal immunoassays satisfied the Japanese environmental standard for cadmium, chromium and, lead, which provided in the soil contamination countermeasures act.