The ARTμS: a novel microfluidic CD4+ T-cell enumeration system for monitoring antiretroviral therapy in HIV patients

Abstract

We report on a novel and cost-effective microfluidic platform that integrates immunomagnetic separation and cell enumeration via DNA-induced bead aggregation. Using a two-stage immunocapture microdevice, 10 μL of whole blood was processed to isolate CD4+ T-cells. The first stage involved the immuno-subtraction of monocytes by anti-CD14 magnetic beads, followed by CD4+ T-cell capture with anti-CD4 magnetic beads. The super hydrophilic surface generated during polydimethylsiloxane (PDMS) plasma treatment allowed for accurate metering of the CD4+ T-cell lysate, which then interacted with silica-coated magnetic beads under chaotropic conditions to form aggregates. Images of the resulting aggregates were captured and processed to reveal the mass of DNA, which was used to back-calculate the CD4+ T-cell number. Studies with clinical samples revealed that the analysis of blood within 24 hours of phlebotomy yielded the best results. Under these conditions, an accurate cell count was achieved (R(2) = 0.98) when compared to cell enumeration via flow cytometry, and over a functional dynamic range from 106-2337 cells per μL.

Analysis of Leukocyte Behaviors on Microfluidic Chips

“…Because CD4 + T cells and neutrophils both express genes which belong to stroke‐related expression profile, the result of the following reverse transcription‐polymerase chain reaction (RT‐PCR) experiment can be directly adopted into stroke diagnosis and even stroke subtype determination with accuracy. Besides traditional immobilization, Landers and co‐workers developed a novel microfluidic system for immunomagnetic separation. On their device, monocytes and CD4 + T cells in blood samples were captured and isolated by anti‐CD14 magnetic beads and anti‐CD4 + magnetic beads, respectively.…”

“…A typical application is the enumeration of CD4 + T cells in whole blood samples through anti‐CD4‐coated microchannels for human immunodeficiency virus (HIV) diagnosis . For example, Landers and co‐workers developed a microfluidic platform that integrated immunomagnetic separation and cell enumeration by bead aggregation. Clinical sample studies showed that an accurate cell count result was achieved ( R 2 = 0.98) compared to flow cytometry.…”

Biochips—New Platforms for Cell‐Based Immunological Assays

“…A 7.7% lower granulocyte ratio accompanied by a 6.8% higher lymphocyte ratio and a 0.7% higher monocyte ratio were observed in this system, when comparing with the outcome from a conventional hematology analyzer. Liu et al developed a microfluidic CD4+ T‐cell count system based on immunomagnetic separation combined with cell enumeration via quantitation of cellular DNA by bead aggregation . The counting results showed a high correlation with the gold standard flow cytometry ( R 2 = 0.98).…”

Fundamentals of affinity cell separations

“…Cell separations have transformed over the years to become both an enabling science as well as an analysis/detection method in its own right . Cell separations have been used to make an impact on many biomedical problems including AIDS diagnosis , circulating tumor cell detection , diagnosis of other cancers , measurement of antigen expression, stroke diagnosis , and many others. Benefitting from the microfluidics boom of the last two decades, cell separation methods have taken on new modalities such as centrifugal devices , flow‐altering/cell rolling systems , and multi‐zone, multi‐parameter affinity separations .…”

“…Benefitting from the microfluidics boom of the last two decades, cell separation methods have taken on new modalities such as centrifugal devices , flow‐altering/cell rolling systems , and multi‐zone, multi‐parameter affinity separations . Cell separations can take advantage of physical effects such as size or deformability , external forces such as magnetic fields , acoustic fields , and molecular interaction via affinity capture . Cell affinity separations can be driven by a variety of external forces, including gravity, pressure, and electrophoretic/electrokinetic flow.…”