Harvesting Nanoparticles: Routine measurement of biomarkers, that were previously impossible due to low abundance or instability

Lance A. Liotta MD PhD

Disease-associated blood biomarkers exist in exceedingly low concentrations within complex mixtures of high-abundance proteins such as albumin and immunoglobulins. For this reason, existing methods for biomarker discovery and measurement may not have the sensitivity to detect important biomarkers of early stage disease. A further roadblock is biomarker instability. Biomarkers in the blood may be highly labile and subjected to degradation during blood collection, transportation, and storage. Biomarker instability is a significant source of bias for validation, and can prohibit practical routine clinical use. The nanotechnology developed under this proposal overcomes all of these serious roadblocks that prevent promising biomarkers from being discovered, or hinder biomarker utility for patient benefit. We have created N-isopropylacrylamide (NIPAm) hydrogel porous sieving nanoparticles containing an internal affinity bait.

The nanoparticles perform three independent functions within minutes, in one step, in solution (serum, plasma, or urine): a) molecular size sieving, b) affinity capture of all (99.9%) solution phase low abundance target analyte molecules, and c) complete (99.9%)protection of harvested proteins from enzymatic degradation.

The captured analytes can be readily eluted for analysis. The technology is a panel of dry lyophilized, sub-micron sized harvesting particles that carry specific high affinity chemical baits for known classes of biomarkers. Following introduction of the blood or body fluid, the respective particle populations will remove all of their target molecules, in one step, in solution, from the entire volume of the sample and concentrate the sequestered analytes inside the particles. Analytes will then be eluted from the particles in a small volume to yield a much higher concentration and purification compared to the starting sample. Depending on the starting volume, this technology can concentrate a biomarker many hundred fold, and fully prevent biomarker degradation, within minutes.

Luchini, A., et al., Smart hydrogel particles: biomarker harvesting: one-step affinity purification, size exclusion, and protection against degradation. Nano Lett, 2008. 8(1): p. 350-61.

Luchini, A., et al., Nanoparticle Technology: Addressing the fundamental roadblocks to protein biomarker discovery. Journal of Materials Chemistry, 2009. DOI: 10.1039/b822264a.

Longo, C., et al., Core-shell hydrogel particles harvest, concentrate and preserve labile low abundance biomarkers. PLoS ONE, 2009. 4(3): p. e4763.

Fredolini, C., et al., Concentration and Preservation of Very Low Abundance Biomarkers in Urine, such as Human Growth Hormone (hGH), by Cibacron Blue F3G-A Loaded Hydrogel Particles. Nano Research, 2008. 1: p. 502-518.

Keywords: Nanotechnology, core-shell hydrogel nanoparticles, Proteomics, Biomarkers, Oncology, Immunoassay, Troponin, Clinical Chemistry

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