Analysis for TNF-α using solid-phase affinity capture with radiolabel and MALDI-MS detection

Gregory B. Hurst, Michelle V. Buchanan, Linda J. Foote, Stephen Kennel

Research output: Contribution to journalArticle

28 Citations (Scopus)

Abstract

Screening of mutant mice for subtle phenotypes requires sensitive, high- throughput analyses of sentinel proteins in functional pathways. The cytokine TNF-α is upregulated during inflammatory reactions associated with antoimmune diseases. We have developed a method to monitor the concentration of TNF-α under physiological conditions. TNF-α is captured, purified, and concentrated using monoclonal antibody-coated microbeads. The capture is efficient (>80%) and can be used in the concentration range <100 pg/mL to >50 ng/mL, as determined by detection of 125I-labeled TNF-α. The bead capture of TNF-α can be combined with direct detection by MALDI-MS for sample concentrations of > 10 ng/mL. TNF-α can be captured and detected from diluted mouse serum, with minimal interferences observed in the MALDI spectrum. This method is adaptable to high-throughput sample handling with microfluidic devices and automated mass spectrometric analysis.

Original languageEnglish (US)
Pages (from-to)4727-4733
Number of pages7
JournalAnalytical Chemistry
Volume71
Issue number20
DOIs
StatePublished - Oct 15 1999
Externally publishedYes

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Throughput
Microfluidics
Screening
Monoclonal Antibodies
Cytokines
Proteins
cyhalothrin

All Science Journal Classification (ASJC) codes

  • Analytical Chemistry

Cite this

Analysis for TNF-α using solid-phase affinity capture with radiolabel and MALDI-MS detection. / Hurst, Gregory B.; Buchanan, Michelle V.; Foote, Linda J.; Kennel, Stephen.

In: Analytical Chemistry, Vol. 71, No. 20, 15.10.1999, p. 4727-4733.

Research output: Contribution to journalArticle

Hurst, Gregory B. ; Buchanan, Michelle V. ; Foote, Linda J. ; Kennel, Stephen. / Analysis for TNF-α using solid-phase affinity capture with radiolabel and MALDI-MS detection. In: Analytical Chemistry. 1999 ; Vol. 71, No. 20. pp. 4727-4733.
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