<p>Detection of biomolecules (e.g., proteins and microRNAs) is essential to uncover fundamental mechanisms of life such as cell division, morphogenesis, and development. Current methods for biomolecules detection are primarily dependent on the absorption, production, or emission of photons (e.g., fluorescence, luminescence, and UV spectra) as output signals. Despite their prevalent usage for daily detection in life sciences, those methods are often limited by environmental noises. In addition, adapting these detection methods for high throughput analysis (i.e. simultaneous detection of multiple biomolecules) is difficult because of their limited multiplexing capabilities owing to spectral overlap. Furthermore, the detections are challenged by the low abundance of biomolecules due to the small size of biological samples (single cell, cell spheroid, or 3D engineered miniature tissue construct). For example, while ELISA is widely used for high sensitivity protein detection, it is difficult to use ELISA for low abundance serum analysis of single cells. Therefore, there is an urgent need to explore novel methods and principles for ultra-sensitive biomolecule detection.<br>
Here we report a new biomolecule detection method that can detect ultra-low concentration biomolecules. In this method, aptamer (single-stranded oligonucleotide or peptide) modified microparticles with attached complementary sequence (CS)-conjugated nanoparticles (NPs) are used as sensing materials. When target biomolecules (specific to the aptamer on the microparticles) are present near the microparticles, a small amount of biomolecule will induce the conformational disassociation of one complementary sequence - aptamer complex and release one nanoparticle from the microparticle. The higher the biomolecular concentration, the more the released nanoparticles. A resistive pulse sensor with a nanopore (2000nm by 2000nm) is then used to detect the released nanoparticles based on Coulter counting principle. The nanoparticles' size could be recalculated by the amplitude of the RPS signals. From the number of the released nanoparticles, biomolecular concentration can be calculated. We will first conjugate complementary sequence (CS) to 500nm Silica NPs (SNPs) method via click chemistry. After the CS-SNPs are made, CS-conjugated nanoparticles were attached to aptamer-modified micropillars via intermolecular CS-aptamer hybridization. Next, we used Adenosine as the target biomolecules to prove the method. Adenosine with various concentrations (from 1 nM to 10mM) was mixed separately with aptamer-modified microparticles for 1h. The counting results from the nanopore resistive sensor showed that the released nanoparticles exponentially increased as the biomolecular concentration increased. At the lowest concentrations we tested (1 nM and 10 nM), we measured 22,500/μL and 34,800/μL released nanoparticles respectively. The results suggested it is highly possible that the method is highly sensitive and can be used to measure biomolecular with smaller concentration (e.g. pM level) and a larger range of adenosine concentrations with fewer errors. The new method has the potentials to advance the field of biomolecule detection and benefit various biomedical and biological researches.&nbsp;<br>
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Sensitive Biomolecular Detection via Nanoparticle Counting

Paper Proceedings: https://asmedigitalcollection.asme.org/IMECE/IMECE2021/volume/85543

ASME 2021 International Mechanical Engineering Congress and Exposition (IMECE2021)

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