33335 - Soluble C5b-9 Complement

Test Code: 33335

TA TMA Transplant-Associated Thrombotic Microangiopathy

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Clinical and Procedure
Clinical Utility

The Soluble Terminal Complement Complex enzyme immunoassay (EIA) measures the amount of the SC5b-9 complex present in human plasma. The Terminal Complement Complex (TCC, SC5b-9) is generated by the assembly of C5 through C9 as a consequence of activation of the complement system by either the classical, lectin or alternative pathway. The membrane attack complex (MAC), a form of TCC, is a stable complex that mediates the irreversible target cell membrane damage associated with complement activation. Complexes formed in the absence of a target membrane bind to naturally occurring regulatory serum proteins, e.g. the S protein forming non-cytolytic complexes in plasma.1,2  

About Sc5b-9 Complement 

The SC5b-9 EIA test quantitates SC5b-9 in plasma. The drug eculizumab, a terminal complement inhibitor, has been shown to inhibit hemolysis in patients with paroxysmal nocturnal hemoglobinuria (PNH) and to inhibit complement-mediated thrombotic microangiopathy in patients with atypical HUS (aHUS).3,4 Eculizumab is a humanized monoclonal antibody that binds to the complement protein C5 with high affinity, thereby inhibiting its cleavage to C5a and C5b and preventing the generation of the cytolytic terminal complement complex, C5b-9. Elevated plasma levels of SC5b-9 may be an indicator of an insufficient level of eculizumab to maintain blocking the formation of the terminal attack complex.5,6 Atypical HUS patients have been shown to display increased levels of C3a, C5a and C5b-9 in plasma prior to treatment.7,8 Elevated plasma levels of the biomarkers C3a C5a may reflect ongoing activation even though full inhibition of the global activity (CH50 and sC5b-9) has been demonstrated.9,10  

Elevated concentrations of C5b-9 are associated with the development of transplant-associated thrombotic microangiopathy (TA-TMA), a complication of hematopoietic stem cell transplant. 11-13 Patients with higher sC5b-9 concentrations at baseline may require the use of higher doses of eculizumab to treat TA-TMA14 especially in children. Because of this association, measurement of sC5b-9 before transplant as part of a diagnostic evaluation and then repeat measurements during therapy have been proposed as tools to follow-up patients.15 Importantly, while the elevation of sC5b-9 has shown very high sensitivity for TA-TMA, it has shown only a modest specificity, ranging from 40% to 50%, and the increased sC5b-9 may be found in other transplant complications as well as several other conditions where complement activation may occur: immune-complex disease, infection, atypical hemolytic uremic syndrome, C3 glomerulopathies, etc. 

Procedure

Microtiter plates are coated with monoclonal antibody specific to the C9 ring of the soluble C5b-9 (sC5b-9) complex. Controls, standards, and patient samples are exposed to the plate. After washing the plate, a horseradish peroxidase-conjugated anti-sC5b-9 complex antibody is added followed by a substrate to initiate color change. 

Turnaround Time

Same day (within 24 of hours of receipt of specimen), Monday through Saturday.

Specimen Information
Specimen Type Test Code CPT Code NY Approved Volume Assay Range Special Instructions
plasma 33335 86160 Yes

Optimal 1 mL; minimum 0.3 ml
  • Samples should be centrifuged (preferably immediately) within 2 hours, after collection at 2000g for 10 minutes.
  • Plasma should be immediately aliquoted into two aliquots and frozen 7 days at -20° C or below
  • K2 EDTA plasma is the required sample type.
Shipping

Ship Monday through Friday. Friday shipments must be labeled for Saturday delivery.

Disclaimer

Specimens are approved for testing in New York only when indicated in the Specimen Information field above. The CPT codes provided are based on Eurofins Viracor’s interpretation of the American Medical Association's Current Procedural Terminology (CPT) codes and are provided for informational purposes only. CPT coding is the sole responsibility of the billing party. Questions regarding coding should be addressed to your local Medicare carrier. Eurofins Viracor assumes no responsibility for billing errors due to reliance on the CPT codes illustrated in this material.

References
  1. Podack, ER, Kolb, WP, and Müller-Eberhard, HJ. The C5b-6 complex: formation, isolation, and inhibition of its activity by lipoprotein and the S-protein of human serum. J Immunol. 1978; 120: 1841-8. 
  2. Podack, ER and Müller-Eberhard, HJ. Isolation of human S-protein, of an inhibitor of the membrane attack complex of complement. J Biol Chem. 1979; 254:9808-14. 
  3. Hillmen P, Hall C, Marsh JC, et al. Effect of eculizumab on hemolysis and transfusion requirements in patients with paroxysmal nocturnal hemoglobinuria. N Engl J Med. 2004; 350:552-9.
  4. Legendre, CM, Licht, C, Muus, P, et al. Terminal complement inhibitor eculizumab in atypical hemolytic-uremic syndrome. N Engl J Med 2013; 368:2169-81.
  5. Prüfer, F, Scheiring, J, Sautter, S, et al. Terminal complement complex (C5b-9) in children with recurrent hemolytic uremic syndrome. Semin Thromb Hemost 2006; 32:121-7. 
  6. Zimmerhackl, LB, Hofer, J, Cortina G, et al. Prophylactic eculizumab after renal transplantation in atypical hemolytic-uremic syndrome. N Engl J Med. 2010; 362:1746-8. 
  7. Mache CJ, Acham-Roschitz B, Fremeaux-Bacchi V, Kirschfink M, Zipfel PF, Roedi S, Vester U, Ring E. Complement inhibitor eculizumab in atypical hemolytic uremic syndrome Clin J Am Soc Nephrol 2009: 4: 1312-16. 
  8. Wehling C, Amon O, Hohenstein B, Pape L, Bommer M, Kirschfink M. Eculizumab drug monitoring in serum and urine opens new insights into therapy of complement-medicated nephropathies. Mol Immunol 2013; 56: 257. 
  9. Spero R, Cataland V, Holers M, Geyer S, Yang S, and Wu HM. Biomarkers of the alternative pathway and terminal complement activity at presentation confirms the clinical diagnosis of aHUS and differentiates aHUS from TTP. Blood 2014; 123:3733-8. 
  10. Noris M, Galbusera M, Gastoldi S, Macor P, Banteria F, Bresin E, Tripodo C, Bettoni A, Donadelli R, Valoti E, Tedesco F, Amore A, Coppo R, Ruggenenti P, Gotti E and Remuzzi G. Dynamics of complement activation in aHUS and how to monitor eculizumab therapy. Blood 2014 124:1715-1726. 
  11. Qi J, Wang J, Chen J, Su J, et al: Plasma levels of complement activation fragments C3b and sC5b-9 significantly increased in patients with thrombotic microangiopathy after allogeneic stem cell transplantation. Ann Hematol. 2017 Nov;96(11):1849-1855 
  12. Horvath O, Kallay K, Csuka D, et al: Early increase in complement terminal pathway activation marker sC5b-9 Is predictive for the development of thrombotic microangiopathy after stem cell transplantation. Biol Blood Marrow Transplant. 2018 May;24(5):989-996 
  13. Mezo B, Horvath O, Sinkovits G, Veszeli N, Krivan G, Prohaszka Z: Validation of early increase in complement activation marker sC5b-9 as a predictive biomarker for the development of thrombotic microangiopathy after stem cell transplantation. Front Med (Lausanne). 2020 Oct 6;7:569291 
  14. Jodele S, Dandoy CE, Lane A, et al: Complement blockade for TA-TMA: lessons learned from a large pediatric cohort treated with eculizumab. Blood. 2020 Mar 26;135(13):1049-1057 
  15. Young JA, Pallas CR, Knovich MA: Transplant-associated thrombotic microangiopathy: theoretical considerations and a practical approach to an unrefined diagnosis. Bone Marrow Transplant. 2021 Aug;56(8):1805-1817
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