A simple and low-cost paper chip-based smartphone sensor for enumeration of T lymphocyte subsets

Lymphocyte count reflects the cellular immune function status of the body, and T lymphocyte subsets are of great value in the adjuvant diagnosis, efficacy monitoring, and prognostic assessment of clinical diseases. There is an urgent need to develop a simple and economical cell-counting platform for T lymphocyte subsets to meet clinical needs of cancer prognosis. Based on the vertical flow assay (VFA) and a polystyrene (PS) microsphere-labeled antibody specifically binding to target cells, we established a simple T lymphocyte counting assay (PS-VFA). Labeled cells induced a signal, which was detected on the filter membrane. We used 3D printing to create a cell phone holder and a test kits. The cell phone holder eliminates ambient light interference and provides the appropriate focus, and a smartphone was used for image acquisition and data analysis, revealing the number of T lymphocytes to be tested via RGB signal analysis. The entire detection process takes about 15 min, and the detection device is extremely cheap, about $20, and can be reused. CD3, CD4, and CD8 cells count exhibited a quasilinear response to logarithmic cell concentrations in a single nucleated cell solution, and detection limits ranging from 100 to 1400 cells/μL for CD3(R² = 0.96), 100 to 1400 cells/μL for CD4(R² = 0.97), and 25 to 800 cells/μL for CD8(R² = 0.96), respectively. And the CV values of this method are all less than 10 %. To validate the clinical applicability of this method,16 patients with cancer and 34 healthy individuals samples were measured, the detection results showed a high correlation with the flow cytometry (FCM) analysis. These results revealed that PS-VFA showed good performance compared with flow cytometry and precision evaluation. This analysis approach is a promising alternative for the costly standard flow cytometry-based tools for T lymphocyte subsets in tumor patients in resource-limited settings.