Alternative complement pathway hemolytic assays reveal incomplete complement blockade in patients treated with eculizumab

Clin Immunol. 2017 Oct:183:1-7. doi: 10.1016/j.clim.2017.06.007. Epub 2017 Jun 21.


Eculizumab is a monoclonal anti-C5 antibody used in the treatment of atypical hemolytic uremic syndrome (aHUS). We monitored complement inhibition in 16 eculizumab-treated patients suffering from HUS or transplant rejection (not aHUS patients). Blood samples were obtained one to four weeks after the last eculizumab injection. We observed that eculizumab efficiently blocked the terminal pathway (TP) through classical pathway (CP) activation measured by kinetic hemolytic assay (HA) (<10%) but incompletely blocked the TP through alternative pathway (AP) activation measured by rabbit (APH50>23%) or chicken erythrocytes HA (AP100>15%). Conversely, functional ELISA revealed a complete blockade of TP through AP activation in all patients (<10%). C5a and sC5b9 levels were not correlated with residual APH50 or AP100. Similar results were obtained after in vitro addition of increasing amounts of eculizumab to a control serum (in vitro APH50>60% and AP100>20%). We also showed that ELISA was less sensitive than HA.

Keywords: Anti-C5; Assay sensitivity; Complement alternative pathway; Complement classical pathway; Drug monitoring; ELISA; Hemolytic assay.

MeSH terms

  • Adolescent
  • Adult
  • Aged
  • Antibodies, Monoclonal, Humanized / therapeutic use*
  • Atypical Hemolytic Uremic Syndrome / drug therapy*
  • Atypical Hemolytic Uremic Syndrome / immunology
  • Child
  • Child, Preschool
  • Complement Activation / immunology
  • Complement C5a / immunology
  • Complement Inactivating Agents / therapeutic use*
  • Complement Membrane Attack Complex / immunology
  • Complement Pathway, Alternative / immunology*
  • Complement Pathway, Classical / immunology*
  • Enzyme-Linked Immunosorbent Assay
  • Female
  • Graft Rejection / drug therapy*
  • Humans
  • Infant
  • Kidney Transplantation
  • Male
  • Middle Aged
  • Treatment Outcome
  • Young Adult


  • Antibodies, Monoclonal, Humanized
  • Complement Inactivating Agents
  • Complement Membrane Attack Complex
  • Complement C5a
  • eculizumab