Abstract
Background
The flow cytometry-based basophil activation test (BAT) is used for the diagnosis of allergic response. However, flow cytometry is time-consuming, requiring skilled personnel and cumbersome processing, which has limited its use in the clinic. Here, we introduce a novel microfluidic-based immunoaffinity BAT (miBAT) method.
Methods
The microfluidic device, coated with anti-CD203c, was designed to capture basophils directly from whole blood. The captured basophils are activated by anti-FcεRI antibody followed by optical detection of CD63 expression (degranulation marker). The device was first characterized using a basophil cell line followed by whole blood experiments. We evaluated the device with ex vivo stimulation of basophils in whole blood from healthy controls and patients with allergies and compared it with flow cytometry.
Results
The microfluidic device was capable of capturing basophils directly from whole blood followed by in vitro activation and quantification of CD63 expression. CD63 expression was significantly higher (P = 0.0002) in on-chip activated basophils compared with nonactivated cells. The difference in CD63 expression on anti-FcεRI-activated captured basophils in microfluidic chip was significantly higher (P = 0.03) in patients with allergies compared with healthy controls, and the results were comparable with flow cytometry analysis (P = 0.04). Furthermore, there was no significant difference of CD63% expression in anti-FcεRI-activated captured basophils in microfluidic chip compared with flow cytometry.
Conclusions
We report on the miBAT. This device is capable of isolating basophils directly from whole blood for on-chip activation and detection. The new miBAT method awaits validation in larger patient populations to assess performance in diagnosis and monitoring of patients with allergies at the point of care.
Single stream inertial focusing in a straight microchannel