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Electrophoresis. 2009 Nov;30(22):3947-54. doi: 10.1002/elps.200900311.

Receptor affinity CE for measuring bioactive inflammatory cytokines in human skin biopsies.

Author information

1
Laboratory of Bioengineering and Physical Science, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, MD 20892, USA. phillipt@mail.nih.gov

Abstract

A chip-based receptor affinity CE system has been employed to measure the concentrations of bioactive pro-inflammatory cytokines in biopsy materials obtained from human atopic skin lesions. The device employs a replaceable affinity disk to which recombinant cytokine receptors have been chemically immobilized. Homogenates obtained from micro-dissected human skin samples were injected into the system where the bioactive cytokines were captured in the receptor affinity port and labeled in situ with a laser dye. The captured cytokines were released and separated by CE, the resolved peaks being detected and measured by laser-induced fluorescence. When compared with conventional cell-based bioassays, the affinity receptor chip showed reasonable correlation with r(2) values of 0.998 for interferon gamma, 0.994 for IL-6 and 0.991 for tumor necrosis factor alpha. The complete process including cytokine capture, labeling, and analysis took approximately 12.5 min with intra- and inter-assay CVs below 5.3% and recoveries of 84.9-98.4% at the 100 pg/mL concentration in buffer solutions and 84.5-95% in normal human tissue extract. The system could indicate clear differences between the various clinical stages of atopic dermatitis in human patients and could run 4-6 samples per hour. This system, like previous chip-based systems designed in our laboratory, holds the potential for being modified to be a portable unit that could be used in clinics and other biomedical screening studies.

PMID:
19938183
DOI:
10.1002/elps.200900311
[Indexed for MEDLINE]

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