A Rapid, Simple, and Low-Cost CD4 Cell Count Sensor Based on Blocking Immunochromatographic Strip System

Abstract

The counting of CD4 + T lymphocytes (CD4 cells) is a critical test for evaluating the immune function of HIV-infected peoples and tumor patients. A rapid, simple, accurate, and low-cost CD4 cell counting method as a diagnostic tool is increasingly required in the clinic. We designed and developed a novel fluorescent immunochromatographic strips (ICS) system based on the blocking principle for counting CD4 cells. The strategy of this system is to count CD4 cells indirectly, by measuring the free CD4 antibodies that were not bound by CD4 cells. The fluorescent antibodies bound to CD4 cells were blocked at the filter pads, resulting in a decrease in fluorescence of free CD4 antibodies measured. The number of CD4 cells was inversely related to the fluorescence intensity. The CD4 count-ICSs exhibited a quasilinear response (R 2 = 0.96) to logarithmic CD4 cell concentrations in PBMC samples in the range of 50−1000 cells/μL. In addition, the CD4 count-ICSs reliably quantified CD4 cells in whole blood samples, where the assay exhibited a linear correlation (R 2 = 0.976) readout for CD4 cell concentrations ranging from 100 to 800 cells/μL. To validate the clinical applicability of this method, 54 blood samples were measured: the detection results showed a high correlation (R 2 > 0.97) with the flow cytometry (FCM) analysis. The fluorescent ICSs can be used to count CD4 cells in blood samples, which have a high coincidence rate with FCM analysis; therefore, the CD4 count ICS system is an excellent candidate method for CD4 cell counting in resource-limited settings.

Biosensors for Detecting Lymphocytes and Immunoglobulins

“…Xiao et al counted CD4 + cells using fluorescent immuno-chromatographic strips (ICSs) that measured the free CD4 + antibodies not bound by CD4 + cells [104]. In whole-blood samples, ICSs exhibited a linear range of 100-800 cells/µL and LOD of 44 cells/µL.…”

Paper-Based Cytometer for the Detection and Enumeration of White Blood Cells According to Their Immunophenotype

“…Recently, LFTs have been developed to indirectly enumerate certain WBC subsets through either lysis of WBCs and subsequent measurement of the amount of surface marker antigen 38−40 or by surface marker-antibody depletion. 41 Analogous paper-based devices designed to enumerate WBCs operate by "WBC trapping" or retaining WBCs based on their size in devices that only incorporate vertical flow and require off-chip labeling. 42−44 While this design may be adequate for some applications (e.g., total WBC count), the restriction to vertical flow and lack of assay integration greatly limits their potential applications and complicates user operation.…”

“…While these membranes allow for the transport of soluble blood analytes, they are incapable of blood cell transport and instead rely on the filtration or lysis of blood cells and wicking of purified plasma or lysate. Recently, LFTs have been developed to indirectly enumerate certain WBC subsets through either lysis of WBCs and subsequent measurement of the amount of surface marker antigen − or by surface marker-antibody depletion . Analogous paper-based devices designed to enumerate WBCs operate by “WBC trapping” or retaining WBCs based on their size in devices that only incorporate vertical flow and require off-chip labeling. − While this design may be adequate for some applications ( e.g.…”

A Novel Paper-Based Cytometer for the Detection and Enumeration of White Blood Cells According to their Immunophenotype

“…Recently, LFTs have been developed to indirectly enumerate certain WBC subsets through either lysis of WBCs and subsequent measurement of the amount of surface marker antigen [37][38][39] or by surface marker-antibody depletion. 40 Analogous paper-based devices designed to enumerate WBCs operate by "WBC trapping" or retaining WBCs based on their size in devices that only incorporate vertical flow and require off-chip labeling. [41][42][43] While this design may be adequate for some applications (e.g., total WBC count), the restriction to vertical flow and lack of assay integration greatly limits their potential applications and complicates user operation.…”