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Analysis of Inflammatory Mediators in Newborn Dried Blood Spot Samples by Chip-Based Immunoaffinity Capillary Electrophoresis

  • Terry M. Phillips
  • Edward F. Wellner
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1972)

Abstract

A chip-based immunoaffinity capillary electrophoresis (ICE) system has been developed for measuring inflammatory mediators in dried blood samples routinely taken from newborn babies. A defined area of each dried blood spot was removed from the sample card and its contents eluted. The recovered eluates were injected into the chip and the analytes of interest isolated by the immunoaffinity disk within the chip. The captured analytes were labeled in-situ with a red light-emitting laser dye and electro-eluted into the chip separation channel. Electrophoretic separation of all of the analytes was achieved within 2.0 min with quantification of each peak being performed by online LIF detection and integration of each peak area. The degree of accuracy and precision achieved by the chip-based system is comparable to conventional immunoassays and the system is robust enough to be applied to the analysis of clinical samples.

Key words

Capillary electrophoresis Dried blood spots Microchip Immunoaffinity Cytokines Chemokines Inflammation Newborns 

Notes

Acknowledgement

This work was supported by the intra-mural research programme at NIH.

References

  1. 1.
    Medzhitov R (2008) Origin and physiological roles of inflammation. Nature 454:428–435CrossRefGoogle Scholar
  2. 2.
    Mussap M (2012) Laboratory medicine in neonatal sepsis and inflammation. J Matern Fetal Neonatal Med 25(Suppl 4):32–34PubMedGoogle Scholar
  3. 3.
    Brandes M, Legler DF, Spoerri B et al (2000) Activation-dependent modulation of B lymphocyte migration to chemokines. Int Immunol 12:1285–1292CrossRefGoogle Scholar
  4. 4.
    Collington SJ, Hallgren J, Pease JE et al (2010) The role of the CCL2/CCR2 axis in mouse mast cell migration in vitro and in vivo. J Immunol 184:6114–6123CrossRefGoogle Scholar
  5. 5.
    Castro PR, Marques SM, Viana CT et al (2014) Deletion of the chemokine receptor CCR2 attenuates foreign body reaction to implants in mice. Microvasc Res 95:37–45CrossRefGoogle Scholar
  6. 6.
    Turner MD, Nedjai B, Hurst T et al (2014) Cytokines and chemokines: at the crossroads of cell signalling and inflammatory disease. Biochim Biophys Acta 1843:2563–2582CrossRefGoogle Scholar
  7. 7.
    Bernardini G, Santoni A (2014) The pathophysiological role of chemokines in the regulation of NK cell tissue homing. Crit Rev Oncog 19:77–90CrossRefGoogle Scholar
  8. 8.
    Patadia M, Dixon J, Conley D et al (2010) Evaluation of the presence of B-cell attractant chemokines in chronic rhinosinusitis. Am J Rhinol Allergy 24:11–16CrossRefGoogle Scholar
  9. 9.
    Pedersen L, Andersen-Ranberg K, Hollergaard M, Nybo M (2017) Quantification of multiple elements in dried blood spot samples. Clin Biochem 50:703–709CrossRefGoogle Scholar
  10. 10.
    Villar LM, de Oliveira JC, Cruz HM, Yoshida CF, Lampe E, Lewis-Ximenez LL (2011) Assessment of dried blood spot samples as a simple method for detection of hepatitis B virus markers. J Med Virol 83:1522–1529CrossRefGoogle Scholar
  11. 11.
    Kwan A, Puck JM (2015) History and current status of newborn screening for severe combined immunodeficiency. Semin Perinatol 39:194–205CrossRefGoogle Scholar
  12. 12.
    Dénes J, Szabó E, Robinette SL, Szatmári I, Szőnyi L, Kreuder JG, Rauterberg EW, Takáts Z (2012) Metabonomics of newborn screening dried blood spot samples: a novel approach in the screening and diagnostics of inborn errors of metabolism. Anal Chem 84:10113–10120CrossRefGoogle Scholar
  13. 13.
    Inaba Y, Schwartz CE, Bui QM, Li X, Skinner C, Field M, Wotton T, Hagerman RJ, Francis D, Amor DJ, Hopper JL, Loesch DZ, Bretherton L, Slater HR, Godler DE (2014) Early detection of fragile X syndrome: applications of a novel approach for improved quantitative methylation analysis in venous blood and newborn blood spots. Clin Chem 60:963–973CrossRefGoogle Scholar
  14. 14.
    Nelson KB, Grether JK, Dambrosia JM, Walsh E, Kohler S, Satyanarayana G, Nelson PG, Dickens BF, Phillips TM (2003) Neonatal cytokines and cerebral palsy in very preterm infants. Pediatr Res 53:600–607CrossRefGoogle Scholar
  15. 15.
    Page-Sharp M, Strunk T, Salman S, Hibbert J, Patole SK, Manning L, Batty KT (2017) Simultaneous determination of pentoxifylline, metabolites M1 (lisofylline), M4 and M5, and caffeine in plasma and dried blood spots for pharmacokinetic studies in preterm infants and neonates. J Pharm Biomed Anal 146:302–313CrossRefGoogle Scholar
  16. 16.
    Welling L, Boelen A, Derks TG, Schielen PC, de Vries M, Williams M, Wijburg FA, Bosch AM (2017) Nine years of newborn screening for classical galactosemia in the Netherlands: effectiveness of screening methods, and identification of patients with previously unreported phenotypes. Mol Genet Metab 120:223–228CrossRefGoogle Scholar
  17. 17.
    Muzembo BA, Mbendi NC, Nakayama SF (2017) Systematic review with meta-analysis: performance of dried blood spots for hepatitis C antibodies detection. Public Health 153:128–136CrossRefGoogle Scholar
  18. 18.
    Phillips TM, Wellner EF (2007) Analysis of inflammatory biomarkers from tissue biopsies by chip-based immunoaffinity CE. Electrophoresis 28:3041–3048CrossRefGoogle Scholar
  19. 19.
    Phillips TM, Wellner EF (2009) Chip-based immunoaffinity CE: application to the measurement of brain-derived neurotrophic factor in skin biopsies. Electrophoresis 30:2307–2312CrossRefGoogle Scholar
  20. 20.
    Kalish H, Phillips TM (2012) Assessment of chemokine profiles in human skin biopsies by an immunoaffinity capillary electrophoresis chip. Methods 56(2):198–203CrossRefGoogle Scholar
  21. 21.
    Guzman NA, Phillips TM (2011) Immunoaffinity capillary electrophoresis: a new versatile tool for determining protein biomarkers in inflammatory processes. Electrophoresis 32:1565–1578Google Scholar
  22. 22.
    O’Shannessy DJ, Quarles RH (1987) Labeling of the oligosaccharide moieties of immunoglobulins. J Immunol Methods 99:153–161CrossRefGoogle Scholar
  23. 23.
    Phillips TM (2004) Rapid analysis of inflammatory cytokines in cerebrospinal fluid using chip-based immunoaffinity electrophoresis. Electrophoresis 25:1652–1659CrossRefGoogle Scholar
  24. 24.
    Phillips TM, Wellner E (2006) Measurement of neuropeptides in clinical samples using chip-based immunoaffinity capillary electrophoresis. J Chromatogr A 1111:106–111CrossRefGoogle Scholar
  25. 25.
    Phillips TM, Wellner E (2013) Detection of cerebral spinal fluid-associated chemokines in birth traumatized premature babies by chip-based immunoaffinity CE. Electrophoresis 34:1530–1538CrossRefGoogle Scholar
  26. 26.
    Pierce Biotechnology Technical Tip #6 (2008) Extinction coefficients. http://www.piercenet.com/previews/guides/tech-tips/

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Terry M. Phillips
    • 1
    • 2
  • Edward F. Wellner
    • 3
  1. 1.Department of Pharmaceutics, School of PharmacyVirginia Commonwealth UniversityRichmondUSA
  2. 2.Department of PharmaceuticsVirginia Commonwealth UniversityWashingtonUSA
  3. 3.National Institute of Bioimaging and BioengineeringNational Institutes of HealthBethesdaUSA

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