Quantification of complement system activation by measuring C5b-9 cell surface deposition using a cell-ELISA technique

J Immunol Methods. 2014 Dec 15:415:57-62. doi: 10.1016/j.jim.2014.09.002. Epub 2014 Sep 28.

Abstract

The complement system is an important aspect of immune defense against microbial invasion. Eukaryotic cells express various complement regulatory proteins to protect them from uncontrolled complement activation. However, some eukaryotic cells possess constitutive complement system activation that does not require specific triggering factors, which is known to have unexpected effects on cell proliferation and survival. This area of research is still preliminary and a standard method to measure complement system activation in eukaryotic cells has yet to be identified. Here, we present a quantitative in vitro method to measure complement system activation in eukaryotic cells by detecting C5b-9, the membrane attack complex, on cell surfaces. The results obtained using this assay correlated with C3b deposition measured using flow cytometry and C5b-9 deposition detected using an immunofluorescence assay. Furthermore, we showed that various cancer cell lines displayed different levels of complement system activation by using this assay.

Keywords: Complement system; ELISA; Membrane attack complex.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antibodies / pharmacology
  • Antigens, CD / genetics
  • Antigens, CD / immunology
  • Cell Adhesion Molecules, Neuronal / antagonists & inhibitors
  • Cell Adhesion Molecules, Neuronal / genetics
  • Cell Adhesion Molecules, Neuronal / immunology
  • Cell Line, Tumor
  • Cell Membrane / chemistry*
  • Cell Membrane / immunology
  • Complement Activation*
  • Complement C3b / pharmacology
  • Complement Membrane Attack Complex / analysis*
  • Endoglin
  • Enzyme-Linked Immunosorbent Assay / methods*
  • Fetal Proteins / antagonists & inhibitors
  • Fetal Proteins / genetics
  • Fetal Proteins / immunology
  • Flow Cytometry
  • Gene Expression
  • Human Umbilical Vein Endothelial Cells / cytology
  • Human Umbilical Vein Endothelial Cells / drug effects
  • Human Umbilical Vein Endothelial Cells / immunology
  • Humans
  • Mesenchymal Stem Cells / cytology
  • Mesenchymal Stem Cells / drug effects
  • Mesenchymal Stem Cells / immunology
  • Organ Specificity
  • Receptors, Cell Surface / antagonists & inhibitors
  • Receptors, Cell Surface / genetics
  • Receptors, Cell Surface / immunology

Substances

  • ALCAM protein, human
  • Antibodies
  • Antigens, CD
  • Cell Adhesion Molecules, Neuronal
  • Complement Membrane Attack Complex
  • ENG protein, human
  • Endoglin
  • Fetal Proteins
  • Receptors, Cell Surface
  • SC5b-9 protein complex
  • Complement C3b