Measurement of Soluble and Membrane‐Bound Interleukin 1 Using a Fibroblast Bioassay

Charles A. Dinarello1, Kathrin Muegge2, Scott K. Durum2

1 University of Colorado Health Sciences Center, Denver, Colorado, 2 National Cancer Institute, Frederick, Maryland
Publication Name:  Current Protocols in Immunology
Unit Number:  Unit 6.2
DOI:  10.1002/0471142735.im0602s37
Online Posting Date:  May, 2001
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Interleukin 1 (IL‐1) is both a major mediator of inflammation and an important signal for activation and differentiation of lymphoid cells. IL‐1 is produced by a large variety of cell types, but in most immunologic systems, activated macrophages are its major cellular source. This unit describes a bioassay for testing biological fluids based on the ability of IL‐1 to induce IL‐8 or IL‐6 in fibroblasts. Also listed are sources of commercially available kits (ELISAs, immunoassays) to measure IL‐1, and antibodies useful in setting up these assays.

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Table of Contents

  • Basic Protocol 1: IL‐1 Induction of IL‐8 by Human Fibroblasts
  • Support Protocol 1: Preparation of Fixed Membrane IL‐1α Samples
  • Commentary
  • Tables
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Basic Protocol 1: IL‐1 Induction of IL‐8 by Human Fibroblasts

  • Fibroblasts (e.g., MRC‐5; ATCC #CCL‐171; see above)
  • Complete RPMI‐5 ( appendix 2A)
  • Trypsin solution (unit 6.9)
  • Hanks' balanced salt solution (HBSS; appendix 2A) without Ca2+ and Mg2+
  • Sample to be tested for IL‐1 activity
  • 150‐cm2 tissue culture flask
  • 24‐well flat‐bottom tissue culture plate
  • Additional reagents and equipment for counting cells ( appendix 3A), trypan blue exclusion ( appendix 3B), and quantitating IL‐8 by ELISA (unit 6.12)

Support Protocol 1: Preparation of Fixed Membrane IL‐1α Samples

  • Cells to be tested for membrane IL‐1α (pro‐IL‐1α)
  • Hanks' balanced salt solution (HBSS; appendix 2A) with and without 1% (w/v) paraformaldehyde, room temperature
  • Serum‐free complete RPMI ( appendix 2A)
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Literature Cited

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Key References
   Dinarello, C.A. and Savage, N. 1989. Interleukin‐1 and its receptor. Crit. Rev. Immunol. 9:1‐20.
   Muegge, K. and Durum, S.K. 1989. The role of interleukin‐1 in the immune response. In Interleukin‐1, Inflammation and Disease (R. Bomford and B. Henderson, eds.) pp. 107‐122. Elsevier Science Publishing, New York.
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