Abstract
This study demonstrates the rapid and label-free detection of Interleukin-6 (IL-6) using carbon nanotube micro-arrays with aptamer as the molecular recognition element. Single wall carbon nanotubes micro-arrays biosensors were manufactured using photo-lithography, metal deposition, and etching techniques. Nanotube biosensors were functionalized with 1-Pyrenebutanoic Acid Succinimidyl Ester (PASE) conjugated IL-6 aptamers. Real time response of the sensor conductance was monitored with increasing concentration of IL-6 (1 pg/mL to 10 ng/mL), exposure to the sensing surface in buffer solution, and clinically relevant spiked blood samples. Non-specific Bovine Serum Albumin (BSA), PBS samples, and anti-IgG functionalized devices gave similar signatures in the real time conductance versus time experiments with no significant change in sensor signal. Exposure of the aptamer functionalized nanotube surface to IL-6 decreased the conductance with increasing concentration of IL-6. Experiments based on field effect transistor arrays suggested shift in drain current versus gate voltage for 1 pg and 1 ng of IL-6 exposure. Non-specific BSA did not produce any appreciable shift in the Ids versus Vg suggesting specific interactions of IL-6 on PASE conjugated aptamer surface gave rise to the change in electrical signal. Both Z axis and phase image in an Atomic Force Microscope (AFM) suggested unambiguous molecular interaction of the IL-6 on the nanotube-aptamer surface at 1 pg/mL concentration. The concentration of 1 pg falls below the diagnostic gray zone for cancer (2.3 pg-4 ng/mL), which is an indicator of early stage cancer. Thus, nanotube micro-arrays could potentially be developed for creating multiplexed assays involving cancer biomarker proteins and possibly circulating tumor cells all in a single assay using PASE functionalization protocol.
Highlights
Single wall carbon nanotubes are ideal candidates for label-free sensing; every atom is on the surface, their electronic properties are very sensitive to the surrounding charge environment [1]
To solve this issue aptamers have been used for biosensing of immunoglobin E (IgE) using carbon nanotube (CNT)-FETs and showed improvement in sensor performance compared to an antibody coated nanotube channel under similar conditions [17]
This paper demonstrates the detection of ultra-low levels of IL-6 protein (BioLegend, Cat.#: 575704)
Summary
Single wall carbon nanotubes are ideal candidates for label-free sensing; every atom is on the surface, their electronic properties are very sensitive to the surrounding charge environment [1]. The smaller arrays that have been developed for detection of cancer cells in buffy coats in the past [10] and more recently demonstrated larger arrays to capture spiked breast cancer cells in blood using the same PASE functionalization protocol [12,13] provide a unique opportunity to create multiplexed assays for the detection of proteins and capture of circulating tumor cells simultaneously all in one digital format This may enable higher information content on the disease progression. The effect on the mobile charge on the material is not realized if the target molecule is placed Debye length away from the surplus charge [46], which can be the case in the ionic environment that the CNT-FET biosensors operate in for biological applications with reference to the antibody size To solve this issue aptamers have been used for biosensing of immunoglobin E (IgE) using CNT-FETs and showed improvement in sensor performance compared to an antibody coated nanotube channel under similar conditions [17]. This study demonstrated the ability to detect IL-6 at concentrations as low as 1 pg/mL by using nanotube micro-arrays
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