A Single-Bead-Based, Fully Integrated Microfluidic System for High-Throughput CD4+T Lymphocyte Enumeration

A single-bead-based, fully integrated microfluidic system has been developed for high-throughput CD4+T lymphocyte enumeration at point-of-care testing. Instead of directly counting CD4+T lymphocytes, CD4+T lymphocyte enumeration is achieved by quantitatively detecting CD4 antigen from the lysed blood sample with a functionalized polycarbonate single bead based on chemiluminescence. To implement the sandwiched chemiluminescence immunoassay with reduced nonspecific binding, a streamlined microfluidic chip with multiple reaction chambers is developed to allow each reaction step to be completed in an independent chamber where reagent is pre-stored. With simple magnetic control, the single bead with an embedded ferrous core can be consecutively transported between each of two adjacent chambers for different reactions. Meanwhile, enhanced mixing can be achieved by moving the single bead back and forth inside one chamber with magnetic actuation. High-throughput detection can be performed when a linear actuation stage is adopted to introduce synchronous magnetic control to multiple single beads in parallel microfluidic chips. A sensitive charge-coupled device (CCD) camera is adopted for high-throughput chemiluminescence detection from multiple single beads. Experimental results show that with the fully integrated microfluidic system, easy-to-operate, accurate, low-cost, immediate, and high-throughput CD4+T lymphocyte enumeration can be successfully achieved at resource-poor settings.