Lensless imaging for point-of-care testing

Abstract

We show a platform that merges a microfluidic chip with lensless imaging for CD4(+) T-lymphocyte counting at resource-limited settings. To capture CD4(+) T lymphocytes, anti-CD4 antibody was immobilized on a microfluidic chip. The captured cells were detected by a charge coupled device (CCD) sensor using lensless shadow imaging techniques. Gray scale shadow images of captured cells on the chip (24 mm x 4 mm x 50 mum) were enumerated in three seconds using an automatic cell counting software. The device achieved 70.2 +/- 6.5% capture efficiency, 88.8 +/- 5.4% capture specificity for CD4(+) T-lymphocytes, 96 +/- 1.6% CCD efficiency, and 83.5 +/- 2.4% overall platform performance (n = 9 devices). This integrated platform has potential for point-of-care testing (POCT) to rapidly capture, image and count specific cell types from unprocessed whole blood.

Fig. 1

Schematic view of the mechanical cell filtration method and shear stress at the fluidic channel floor based on channel geometry and flow rate. (a) Cell selection method in a microfluidic channel and design parameters. The subscript 1 and 2 is used for monocytes and T-lymphocytes, respectively. Cell sizes are based on microvilli and ruffles are estimated from SEM images. The moment ratio between monocytes and T-lymphocytes is derived from: P_xA₁d₁/P_xA₂d₂ = (d₁/d₂)³ = 2.37, for 12 μm and 9 μm cell diameters, respectively. The area, A, and diameter, d represent cross sectional area and diameter of each cell. P_x represents pressure at a distance x from the channel entrance. It has a uniform magnitude as a function of y. The height, H, is the fluidic channel height. Fluid velocity and shear stress as a function of x, and shear stress at wall are described by U_x, τ_x, and τ_w, respectively. (b) Calculated shear stress as a function of channel height for 5 μl/min and 20 μl/min volumetric flow rate. Shear stress range of 0.3 ~ 1.2 N/m² was chosen to maximize cell attachment and 2 N/m² to achieve mechanical filtration. The design parameters were 4 mm (W) × 50 μm (H) and flow rate, 5 μl/min and 20 μl/min.

Fig. 2

A schematic view of the CCD imaging platform: (a) CCD imaging platform to detect the captured cells. When light is incident on the captured cells, the cell membrane diffracts and transmits light. A shadow of the captured CD4⁺ T-lymphocytes generated by diffraction can be imaged by the CCD in one second. Image is obtained with the lens-less CCD imaging platform. (b) Picture of microfluidic chip and CCD imaging platform. Field of view of the CCD sensor is 35 mm × 25 mm. The entire microfluidic device can be imaged without alignment by simply placing the microfluidic channel on the sensor. (c) Image taken with the lens-less CCD imaging platform and magnified view at the microfluidic channel centre is shown. The magnified picture shows an image obtained by diffraction. Scale bar, 100 μm.

Fig. 3

Efficiency of the microfluidic chip and the CCD imaging platform.