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Bio-Impedance Sensing Device (BISD) for Detection of Human CD4+ Cells

N.N. Mishra, S. Retterrer, T.J. Zieziulewicz, M. Isaacson, D. Szarowski, D.E. Mousseau, D.A. Lawrence and J.N. Turner
New York City Department of Health, US

Keywords: biosensor, microelectrode, impedance, human blood, CD4+ cell

The aim of this work is to develop a small on-chip bio-impedance sensing device (BISD) to rapidly detect and quantify cells with a specific phenotype in a heterogenous population of cells (e.g., human CD4+ cells in blood for prognosis of HIV-infected individuals) using a minimal sample volume and minimal preparation. The transducers, gold/titanium microelectrodes (100x100m and 80x80m), have been fabricated on glass substrates. The microelectrode surface is non-covalently modified sequentially with protein-G, human albumin, monoclonal anti-human CD4 antibody and mouse IgG. The anti-human CD4 antibody binds CD4+ cells present in human blood. The basic function of the microelectrodes was characterized using electrochemical cyclic voltammetry before and after protein-G deposition. The binding of biomolecules, protein-G and antibodies, as well as cells was detected by precisely measuring the electrical current, as a function of frequency (1-8 kHz), that flowed between the microelectrode and the much larger reference electrode. This current was measured using a specially designed very low-noise amplifier based on instrument grade operational amplifiers. This measurement was plotted as impedance using the constant amplitude voltage applied between the two electrodes. A series of AC impedance and output voltage measurements have been made with different size gold microelectrodes with adsorbed protein layers as well as CD4+ cells. When human peripheral blood mononuclear cells (PBMC) containing CD4+ cells were incubated on the biosensor a high increase in impedance was observed, this was due to the presence of CD4+ cells, which decrease the current flow. Incubation of the captured cells with FITC-labeled anti-CD4 demonstrated that all cells were CD4+ demonstrating the selectivity of the BISD system. The BISD system is a promising tool for the detection of CD4+ cells in HIV-infected individuals and for the detection of antigen:antibody or receptor:ligand interactions

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