Rapid enumeration of CD4 + T lymphocytes using an integrated microfluidic system based on Chemiluminescence image detection at point-of-care testing.

An integrated microfluidic system has been developed for rapid enumeration of CD4 + T lymphocytes at point-of-care (POC) settings. A concise microfluidic chip, which consists of three separate chambers, respectively, for reaction, detection and waste storage, is developed to automate CD4 detection. To simplify CD4 + T lymphocyte enumeration, a single polycarbonate bead immobilized with CD4 antibody is adopted by the microfluidic chip to capture the CD4 antigen in the lysed testing sample. Desired performance is achieved by actuating the single bead for efficient mixing, as well as transferring it between different reaction chambers to reduce non-specific reaction. A controllable external magnetic field is applied to drive the single bead with a built-in ferrous core for different purposes. Chemiluminescence reaction is implemented in an independent chamber to reduce non-specific binding of enzyme. A simple flow control strategy is adopted to conveniently release the waste reagent into the waste storage chamber by just opening the vent hole without actively pumping. A sensitive CCD camera is used to collect the reaction signal by taking picture from the single bead, and then the signal intensity is further analyzed for CD4 + T lymphocyte enumeration. Experimental results show that rapid, convenient, accurate and low-cost CD4 + T lymphocyte enumeration can be obtained with the developed microfluidic system at POC test.

Breaking through the Poisson Distribution: A compact high-efficiency droplet microfluidic system for single-bead encapsulation and digital immunoassay detection.

Droplet encapsulation of a single cell or bead is widely used in digital detection, single-cell sequencing, and drug screening. However, the encapsulation of particles is totally random restricted by the Poisson distribution. The theoretical possibility of single-particle encapsulation is usually only approximately 10%. In ultra-high multiplexed digital detection or other applications that needing to measure large numbers of particles, the number of the partitions required to be counted is extremely high, further result in great increase of statistical number of invalid droplets and the redundancy of detection data. Here, a bead ordered arrangement droplet (BOAD) system is proposed to break through the Poisson distribution. BOAD system tactfully combines sheath flow, Dean vortex, and compression flow channel to achieve orderly arrangement of particles for the first time, and could achieve the fastest orderly arrangement of particles in the shortest structure. The efficiency of single-bead encapsulation is improved to as high as 86%. Further application to encapsulate encoding beads and IL-10-targeted magnetic beads demonstrates the potential for bead-based ultra-high multiplexed digital detection. Thus, use of the BOAD system is very promising for many applications needing high single-particle encapsulation ratio in limited partitions, such as multiplexed digital bio-detection, single-cell analysis, drug screening, and single exosome detection.

Analysis of the Mixing of Filler and Base Materials in Arc-Welded Single-Bead Surface Welds Using an EDXS Method.

This article deals with an analysis of mixing and determines the admixing rate of a base S355 steel plate in single-bead surface welds by measuring the chemical composition using a plane-scan energy dispersive X-ray spectroscopy (EDXS) on metallographic cross-sections. The results show that obtaining a larger number of EDXS measurements does not necessarily lead to obtaining a more accurate admixing rate. Due to the ever-present segregations that are generally near the base material, the disadvantage of this method is the subjective influence of the SEM operator on the estimated admixing rate. To obtain relevant results, a sufficiently wide area of well-mixed melt, including segregations, must be analyzed. This study showed that by using a sufficiently large number of appropriately selected sites with a sufficiently large surface area, it is possible to estimate the admixing rate from the chemical composition with an accuracy of ≥96% for the geometrically determined admixing rate D = 30%. From several equations, the best result showed an equation which is the arithmetic mean of the two different arithmetic means and in which the artificial influencing factor of the segregations of the base material is taken into account. With this equation, the same value of admixing rate, D = 30%, was obtained using the comparative geometric method.

Determining the Degree of Admixing Rate of the Base Material and the Melting Efficiency in Single-Bead Surface Welds Using Different Methods, Including New Approaches.

The precise determination of the admixing rate of the base material for certain welding parameters is very important because of the possible negative consequences. As such, it is the basis for corrections in welding technology. In the article, experimental and theoretical determinations of the admixing rate in single-bead surface welds that were arc welded onto S355 steel with different alloyed-steel-coated electrodes are discussed. The admixing rate was experimentally estimated from the ratio of the surface areas of metallographic cross-sections, from the ratios of the height and from chemical analyses of different regions of the surface weld, while it was theoretically estimated from the characteristics of the welding process and material constants. One of the key characteristics of the welding process is the melting efficiency, which can be estimated by means of different equations and from knowledge of the heat balance of the welding process. Both the average melting efficiency of the surface welding on the medium-thick S355 steel plate and the average admixing rate of the S355 steel into the surface welds have the same value, i.e., approximately 30%. New equations for estimating the melting efficiency of the arc welding with a coated electrode were developed on the basis of the results.

Manipulation of mass transport rates using bead-in-a-tube method.

In ultralow Pu analyses, the gold standard is thermal ionization mass spectrometry (TIMS), which requires pure sources to achieve its performance. This purity is achieved through step-wise purifications. In this work single, anion-exchange beads were trapped in the tubing to allow for dynamic solution cycling over the surface of the beads to improve the rates of metal complex uptake. Rates of Pu sorption on single ∼900 µm SIR-1200 and ∼620 µm Reillex-HPQ beads were determined for single beads trapped in a tube with syringe pump driven dynamic solution cycling over the bead, improving sorption and desorption rates. A static control was used as a comparison. Using 238Pu to enable facile activity-based measurements, rates were determined by measuring the residual Pu after contact with beads using liquid scintillation analysis (LSA) for fixed periods of time. Syringe pump driven dynamic solution cycling results in ∼5 and ∼15-fold improvements in the sorption rates for SIR-1200 and Reillex-HPQ. Impacts on desorption were also examined.

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.