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Receptor affinity CE for measuring bioactive inflammatory cytokines in human skin biopsies
Author(s) -
Phillips Terry M.,
Kalish Heather,
Wellner Edward
Publication year - 2009
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.200900311
Subject(s) - receptor , cytokine , human skin , atopic dermatitis , chemistry , tumor necrosis factor alpha , recombinant dna , bioassay , microbiology and biotechnology , chromatography , medicine , immunology , biology , biochemistry , gene , genetics
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.