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Nano Research

, Volume 1, Issue 6, pp 502–518 | Cite as

Concentration and preservation of very low abundance biomarkers in urine, such as human growth hormone (hGH), by Cibacron Blue F3G-A loaded hydrogel particles

  • Claudia Fredolini
  • Francesco Meani
  • K. Alex Reeder
  • Sally Rucker
  • Alexis Patanarut
  • Palma J. Botterell
  • Barney Bishop
  • Caterina Longo
  • Virginia Espina
  • Emanuel F. PetricoinIII
  • Lance A. Liotta
  • Alessandra LuchiniEmail author
Open Access
Research Article

Abstract

Urine is a potential source of diagnostic biomarkers for detection of diseases, and is a very attractive means of non-invasive biospecimen collection. Nonetheless, proteomic measurement in urine is very challenging because diagnostic biomarkers exist in very low concentration (usually below the sensitivity of common immunoassays) and may be subject to rapid degradation. Hydrogel nanoparticles functionalized with Cibacron Blue F3G-A (CB) have been applied to address these challenges for urine biomarker measurement. We chose one of the most difficult low abundance, but medically relevant, hormones in the urine: human growth hormone (hGH). The normal range of hGH in serum is 1 to 10 ng/mL but the urine concentration is suspected to be a thousand times less, well below the detection limit (50 pg/mL) of sensitive clinical hGH immunoassays. We demonstrate that CB particles can capture, preserve and concentrate hGH in urine at physiological salt and urea concentrations, so that hGH can be measured in the linear range of a clinical immunometric assay. Recombinant and cadaveric hGH were captured from synthetic and human urine, concentrated and measured with an Immulite chemiluminescent immunoassay. Values of hGH less than 0.05 ng/mL (the Immulite detection limit) were concentrated to 2 ng/mL, with a urine volume of 1 mL. Dose response studies using 10 mL of urine demonstrated that the concentration of hGH in the particle eluate was linearly dependent on the concentration of hGH in the starting solution, and that all hGH was removed from solution. Thus if the starting urine volume is 100 mL, the detection limit will be 0.1 pg/mL. Urine from a healthy donor whose serum hGH concentration was 1.34 ng/mL was studied in order detect endogenous hGH. Starting from a volume of 33 mL, the particle eluate had an hGH concentration of 58 pg/mL, giving an estimated initial concentration of hGH in urine of 0.175 pg/mL. The nanotechnology described here appears to have the desired precision, accuracy and sensitivity to support large scale clinical studies of urine hGH levels.

Keywords

Urine proteomics Cibacron Blue F3G-A human growth hormone hydrogel nanoparticles N-isopropylacrylamide 

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Copyright information

© Tsinghua Press and Springer-Verlag GmbH 2008

Authors and Affiliations

  • Claudia Fredolini
    • 1
    • 2
    • 5
  • Francesco Meani
    • 3
    • 5
  • K. Alex Reeder
    • 5
  • Sally Rucker
    • 5
  • Alexis Patanarut
    • 6
  • Palma J. Botterell
    • 6
  • Barney Bishop
    • 6
  • Caterina Longo
    • 4
    • 5
  • Virginia Espina
    • 5
  • Emanuel F. PetricoinIII
    • 5
  • Lance A. Liotta
    • 5
  • Alessandra Luchini
    • 5
    Email author
  1. 1.Department of UrologyS. Giovanni Bosco HospitalTorinoItaly
  2. 2.Department of Medicine and Experimental OncologyUniversity of TurinTurinItaly
  3. 3.Gynecology and Obstetrics DepartmentUniversity of BresciaBresciaItaly
  4. 4.Department of Dermatology and VenereologyUniversity of Modena and Reggio EmiliaModenaItaly
  5. 5.Center for the Applied Proteomics and Molecular MedicineGeorge Mason UniversityManassasUSA
  6. 6.Chemistry and Biochemistry DepartmentGeorge Mason UniversityManassasUSA

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