Well-Orientation Strategy for Direct Immobilization of Antibodies: Development of the Immunosensor Using the Boronic Acid-Modified Magnetic Graphene Nanoribbons for Ultrasensitive Detection of Lymphoma Cancer Cells

摘要

This work presents an effective strategy for the well-oriented immobilization of antibodies in which boronic acid is directly attached to the surface and with no need of the long and flexible spacer. A magnetic graphene nanoribbon-boronic-acid-based immunosensor was developed and tested for the impedimetric detection of lymphoma cancer cells, a blood cancer biomarker. Magnetic graphene nanoribbons (MGNRs) were modified with boronic acid (BA) to create a supporting matrix that is utilized by immobilizing anti-CD20 antibodies with good orientation. The prepared biosensing layer (MGNR/BA/Ab) with well-oriented antibodies was premixed into whole blood samples to interact with lymphoma cancer cell receptors. In the presence of target cell receptors, an immunocomplex was formed between anti-CD20 antibodies and lymphoma cancer cell receptors. Then, the biosensing layer was magnetically collected on a screen-printed carbon electrode (SPCE) and placed in a homemade electrochemical cell configuration to measure impedimetric signals. The fabrication steps of the immunosensor were characterized by various techniques, such as resonance light scattering, fluorescence, electrochemical impedance spectroscopy, and cyclic voltammetry. The assay is highly sensitive: the calculated limit of detection of lymphoma cancer cells was as low as 38 cells/mL, and the detection was linear from 100 to 1 000 000 cells/mL. The specificity of the immunosensor is also very high, and there is no interference effect with several potential interferents, such as the breast cancer (MCF-7), human embryonic kidney (HEK293), and leukemia (HL-60 and KCL-22) cell lines. The performance of the immunosensor for lymphoma cancer cells in clinical blood samples is consistent with that of commercial flow cytometric assays.