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Core-shell hydrogel particles harvest, concentrate and preserve labile low abundance biomarkers.

Longo C, Patanarut A, George T, Bishop B, Zhou W, Fredolini C, Ross MM, Espina V, Pellacani G, Petricoin EF, Liotta LA, Luchini A - PLoS ONE (2009)

Bottom Line: Particle sequestered PDGF was fully protected from exogenously added tryptic degradation.Beyond PDGF, the sequestration and protection from degradation for a series of additional very low abundance and very labile cytokines were verified.We envision the application of harvesting core-shell nanoparticles to whole blood for concentration and immediate preservation of low abundance and labile analytes at the time of venipuncture.

View Article: PubMed Central - PubMed

Affiliation: Department of Dermatology, University of Modena and Reggio Emilia, Modena, Italy.

ABSTRACT

Background: The blood proteome is thought to represent a rich source of biomarkers for early stage disease detection. Nevertheless, three major challenges have hindered biomarker discovery: a) candidate biomarkers exist at extremely low concentrations in blood; b) high abundance resident proteins such as albumin mask the rare biomarkers; c) biomarkers are rapidly degraded by endogenous and exogenous proteinases.

Methodology and principal findings: Hydrogel nanoparticles created with a N-isopropylacrylamide based core (365 nm)-shell (167 nm) and functionalized with a charged based bait (acrylic acid) were studied as a technology for addressing all these biomarker discovery problems, in one step, in solution. These harvesting core-shell nanoparticles are designed to simultaneously conduct size exclusion and affinity chromatography in solution. Platelet derived growth factor (PDGF), a clinically relevant, highly labile, and very low abundance biomarker, was chosen as a model. PDGF, spiked in human serum, was completely sequestered from its carrier protein albumin, concentrated, and fully preserved, within minutes by the particles. Particle sequestered PDGF was fully protected from exogenously added tryptic degradation. When the nanoparticles were added to a 1 mL dilute solution of PDGF at non detectable levels (less than 20 picograms per mL) the concentration of the PDGF released from the polymeric matrix of the particles increased within the detection range of ELISA and mass spectrometry. Beyond PDGF, the sequestration and protection from degradation for a series of additional very low abundance and very labile cytokines were verified.

Conclusions and significance: We envision the application of harvesting core-shell nanoparticles to whole blood for concentration and immediate preservation of low abundance and labile analytes at the time of venipuncture.

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SDS PAGE analysis showing chemokines uptake by particles.Core-shell particles were incubated with the following chemokines, mucosae-associated epithelial chemokine (MEC/CCL28), stromal cell-derived factor-1 beta, (SDF-1β/CXCL12b), and eotaxin-2 (CCL24), in presence of bovine serum albumin (BSA). Solutions of the chemokines and BSA are shown in lanes 1, 4, and 7. After incubation with the particles, no chemokine was left in the supernatant (S, lane 2, 5, and 8) and all the chemokine was captured by particles (P, lanes 3, 6, and 9). BSA was totally excluded by particles.
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pone-0004763-g008: SDS PAGE analysis showing chemokines uptake by particles.Core-shell particles were incubated with the following chemokines, mucosae-associated epithelial chemokine (MEC/CCL28), stromal cell-derived factor-1 beta, (SDF-1β/CXCL12b), and eotaxin-2 (CCL24), in presence of bovine serum albumin (BSA). Solutions of the chemokines and BSA are shown in lanes 1, 4, and 7. After incubation with the particles, no chemokine was left in the supernatant (S, lane 2, 5, and 8) and all the chemokine was captured by particles (P, lanes 3, 6, and 9). BSA was totally excluded by particles.

Mentions: In Fig. 8 SDS PAGE analysis is shown on core shell acrylic acid functionalized particles incubated with other relevant models for serological biomarker, namely mucosae-associated epithelial chemokine (MEC/CCL28, 12,300 Da), stromal cell-derived factor-1 beta, (SDF-1β/CXCL12b, 8,500 Da), and eotaxin-2 (CCL24, 8,800 Da) mixed with BSA. Chemokines were totally removed from solution, captured and concentrated by particles, whereas BSA was completely excluded.


Core-shell hydrogel particles harvest, concentrate and preserve labile low abundance biomarkers.

Longo C, Patanarut A, George T, Bishop B, Zhou W, Fredolini C, Ross MM, Espina V, Pellacani G, Petricoin EF, Liotta LA, Luchini A - PLoS ONE (2009)

SDS PAGE analysis showing chemokines uptake by particles.Core-shell particles were incubated with the following chemokines, mucosae-associated epithelial chemokine (MEC/CCL28), stromal cell-derived factor-1 beta, (SDF-1β/CXCL12b), and eotaxin-2 (CCL24), in presence of bovine serum albumin (BSA). Solutions of the chemokines and BSA are shown in lanes 1, 4, and 7. After incubation with the particles, no chemokine was left in the supernatant (S, lane 2, 5, and 8) and all the chemokine was captured by particles (P, lanes 3, 6, and 9). BSA was totally excluded by particles.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2651577&req=5

pone-0004763-g008: SDS PAGE analysis showing chemokines uptake by particles.Core-shell particles were incubated with the following chemokines, mucosae-associated epithelial chemokine (MEC/CCL28), stromal cell-derived factor-1 beta, (SDF-1β/CXCL12b), and eotaxin-2 (CCL24), in presence of bovine serum albumin (BSA). Solutions of the chemokines and BSA are shown in lanes 1, 4, and 7. After incubation with the particles, no chemokine was left in the supernatant (S, lane 2, 5, and 8) and all the chemokine was captured by particles (P, lanes 3, 6, and 9). BSA was totally excluded by particles.
Mentions: In Fig. 8 SDS PAGE analysis is shown on core shell acrylic acid functionalized particles incubated with other relevant models for serological biomarker, namely mucosae-associated epithelial chemokine (MEC/CCL28, 12,300 Da), stromal cell-derived factor-1 beta, (SDF-1β/CXCL12b, 8,500 Da), and eotaxin-2 (CCL24, 8,800 Da) mixed with BSA. Chemokines were totally removed from solution, captured and concentrated by particles, whereas BSA was completely excluded.

Bottom Line: Particle sequestered PDGF was fully protected from exogenously added tryptic degradation.Beyond PDGF, the sequestration and protection from degradation for a series of additional very low abundance and very labile cytokines were verified.We envision the application of harvesting core-shell nanoparticles to whole blood for concentration and immediate preservation of low abundance and labile analytes at the time of venipuncture.

View Article: PubMed Central - PubMed

Affiliation: Department of Dermatology, University of Modena and Reggio Emilia, Modena, Italy.

ABSTRACT

Background: The blood proteome is thought to represent a rich source of biomarkers for early stage disease detection. Nevertheless, three major challenges have hindered biomarker discovery: a) candidate biomarkers exist at extremely low concentrations in blood; b) high abundance resident proteins such as albumin mask the rare biomarkers; c) biomarkers are rapidly degraded by endogenous and exogenous proteinases.

Methodology and principal findings: Hydrogel nanoparticles created with a N-isopropylacrylamide based core (365 nm)-shell (167 nm) and functionalized with a charged based bait (acrylic acid) were studied as a technology for addressing all these biomarker discovery problems, in one step, in solution. These harvesting core-shell nanoparticles are designed to simultaneously conduct size exclusion and affinity chromatography in solution. Platelet derived growth factor (PDGF), a clinically relevant, highly labile, and very low abundance biomarker, was chosen as a model. PDGF, spiked in human serum, was completely sequestered from its carrier protein albumin, concentrated, and fully preserved, within minutes by the particles. Particle sequestered PDGF was fully protected from exogenously added tryptic degradation. When the nanoparticles were added to a 1 mL dilute solution of PDGF at non detectable levels (less than 20 picograms per mL) the concentration of the PDGF released from the polymeric matrix of the particles increased within the detection range of ELISA and mass spectrometry. Beyond PDGF, the sequestration and protection from degradation for a series of additional very low abundance and very labile cytokines were verified.

Conclusions and significance: We envision the application of harvesting core-shell nanoparticles to whole blood for concentration and immediate preservation of low abundance and labile analytes at the time of venipuncture.

Show MeSH