Infliximab (Remicade®) ELISA (mAb-based) Assay Kit

$1,190.00

The Eagle Biosicences Infliximab (mAb-based) ELISA Assay Kit is an enzyme immunoassay for the specific and precise measurement of free Infliximab in serum and plasma samples. This assay utilizes the specific monoclonal antibodies as the coating material on the microtiter plate so this assay specifically measures only Infliximab and no other TNF-α therapeutic.  The Eagle Biosciences Infliximab (mAb-based) ELISA Assay Kit is for research use only and not to be used in diagnostic procedures.

Infliximab (Remicade®) ELISA (mAb-based) Assay Kit

For Research Use Only

Size: 1×96 wells
Sensitivity: 5 ng/mL
Dynamic Range: 6 – 200 ng/mL
Incubation Time: 2 hours
Sample Type: Serum, Plasma
Sample Size: 10 µL

Additional Information

Assay Background


The drug Infliximab (trade names Remicade®, Remsima®, and Inflectra®) is a chimeric monoclonal antibody and used to treat auto-immune disorders. Infliximab reduces the amount of active human tumor necrosis factor alpha (hTNF-α) in the patient by binding to it and preventing it from signaling the receptors for TNF-α on the surface of various cell types. TNF-α is one of the key cytokines that triggers and sustains the inflammatory reactions. Infliximab is used for the treatment of psoriasis, Crohn’s disease, ankylosing spondylitis, psoriatic arthritis, rheumatoid arthritis, ulcerative colitis.

The specificity of this test system is achieved by using a monoclonal antibody named “Peri-1 mAb” for the coating of the microtiter plate. This antibody is specific for Infliximab only (regardless whether Remicade®, Remsima®, or Inflectra®) and does not cross react with other TNF-α catchers.

In this context, identification of biomarkers for (non-)response and risk factors for adverse drug reactions relating to serum concentrations and therapeutic drug monitoring of Infliximab would be very helpful.

Assay Principle


This Eagle Biosciences ELISA Assay Kit is based on Infliximab-specific mouse monoclonal antibody (catcher Ab, ImmunoGuide clone Peri-1). Diluted standards and samples are incubated in the microtiter plate coated with IG-Peri-1 mAb. After incubation, the wells are washed. A horseradish peroxidase (HRP)-conjugated anti-human IgG monoclonal antibody is added and binds to the Fc part of Infliximab. Following incubation, wells are washed and the bound enzymatic activity is detected by addition of chromogen-substrate. The color developed is proportional to the amount of Infliximab in the sample or standard. Results of samples can be determined by using the standard curve. Binding of Infliximab to the solid phase, pre-coated with Peri-1, is inhibited by recombinant human TNF-α or Antibody to Infliximab (ATI) in a concentration dependent manner. Therefore, the ImmunoGuide Infliximab ELISA (mAb-based) measures the free form of Infliximab.

Assay Procedure


  1. Pipette 100 µL of Assay Buffer into each of the wells to be used.
  2. Pipette 50 µL of each 1:10 Diluted Standard, and 1:100 Diluted Samples into the respective wells of the microtiter plate.
  3. Cover the plate with adhesive seal. Shake plate carefully. Incubate 60 min at room temperature (RT, 20-25°C).
  4. Remove adhesive seal. Aspirate or decant the incubation solution. Wash the plate 3 X 300 μL of Diluted Wash Buffer per well. Remove excess solution by tapping the inverted plate on a paper towel.
  5. Pipette 100 μL of Enzyme Conjugate (HRP-anti human IgG mAb) into each well.
  6. Cover plate with adhesive seal. Shake plate carefully. Incubate 30 min at RT.
  7. Remove adhesive seal. Aspirate or decant the incubation solution. Wash the plate 3 X 300 μL of Diluted Wash Buffer per well. Remove excess solution by tapping the inverted plate on a paper towel.
  8. Pipette 100 µL of Ready-to-Use TMB Substrate Solution into each well.
  9. Incubate 10 min at RT. Avoid exposure to direct sunlight.
  10. Stop the substrate reaction by adding 100 µL of Stop Solution into each well. Briefly mix contents by gently shaking the plate. Color changes from blue to yellow.
  11. Measure optical density (OD) with a photometer at 450 nm (Reference at OD620 nm is optional) within 15 min after pipetting the Stop Solution.

Typical Standard Curve


Publications

Citations


  1. Lee YS, Baek SH, Kim MJ, Lee YM, Lee Y, Choe YH. Efficacy of Early Infliximab Treatment for Pediatric Crohn’s Disease: A Three-year Follow-up. Pediatr Gastroenterol Hepatol Nutr. 2012; 15(4):243-9.
  2. Bortlik M, Machkova N, Duricova D, Malickova K, Hrdlicka L, Lukas M, Kohout P, Shonova O, Lukas M. Pregnancy and newborn outcome of mothers with inflammatory bowel diseases exposed to anti-TNF-α therapy during pregnancy: three-center study. Scand J Gastroenterol. 2013; 48(8):951-8.
  3. Grosen A, Julsgaard M, Kelsen J, Christensen LA. Infliximab concentrations in the milk of nursing mothers with inflammatory bowel disease. J Crohns Colitis. 2014; 8(2):175-6.
  4. Krajcovicova A, Hlavaty T, Zelinkova Z, Letkovsky J, Huorka M. Delayed hypersensitivity reaction after initial dose of infliximab: a case report. Eur J Gastroenterol Hepatol. 2014; 26(4):485-7.
  5. Ducourau E, Mulleman D, Paintaud G, Chu Miow Lin D, Lauféron F, Antibodies toward infliximab are associated with low infliximab concentration at treatment initiation and poor infliximab maintenance in rheumatic disease. Arthritis Research & Therapy 2011; 13:R105.
  6. Farkas K, Rutka M, Bálint A, Nagy F, Bor R, Milassin Á, Szepes Z, Molnár T. Efficacy of the new infliximab biosimilar CT-P13 induction therapy in Crohn’s disease and ulcerative colitis – experiences from a single center. Expert Opin Biol Ther. 2015; 15(9):1257-62.
  7. Lee YM, Kang B, Lee Y, Kim MJ, Choe YH. Infliximab “Top-Down” Strategy is Superior to “Step-Up” in Maintaining Long-Term Remission in the Treatment of Pediatric Crohn Disease. J Pediatr Gastroenterol Nutr. 2015;60(6):737-43.
  8. Gecse KB, Végh Z, Lakatos PL. Optimizing biological therapy in Crohn’s disease. Expert Rev Gastroenterol Hepatol. 2015 Oct 16:1-9.
  9. Choe JY, Prodanovic N, Niebrzydowski J, Staykov I, Dokoupilova E, Baranauskaite A, Yatsyshyn R, Mekic M, Porawska W, Ciferska H, Jedrychowicz-Rosiak K, Zielinska A, Choi J, Rho YH, Smolen JS. A randomised, double-blind, phase III study comparing SB2, an infliximab biosimilar, to the infliximab reference product Remicade in patients with moderate to severe rheumatoid arthritis despite methotrexate therapy. Ann Rheum Dis. 2015. pii: annrheumdis-2015-207764.
  10. Thomas SS, Borazan N, Barroso N, Duan L, Taroumian S, Kretzmann B, Bardales R, Elashoff D, Vangala S, Furst DE. Comparative Immunogenicity of TNF Inhibitors: Impact on Clinical Efficacy and Tolerability in the Management of Autoimmune Diseases. A Systematic Review and Meta-Analysis. BioDrugs. 2015; 29(4):241-58.
  11. Plasencia C, Jurado T, Villalba A, Peitedado D, Casla MT, Nuño L, Bonilla MG, Martínez-Feito A, Martín-Mola E, Pascual-Salcedo D, Balsa A. Effect of Infliximab Dose Increase in Rheumatoid Arthritis at Different Trough Concentrations: A Cohort Study in Clinical Practice Conditions. Front Med (Lausanne). 2015 Oct 8;2:71. doi: 10.3389/fmed.2015.00071.
  12. Minar P, Saeed SA, Afreen M, Kim MO, Denson LA. Practical Use of Infliximab Concentration Monitoring in Pediatric Crohn’s Disease. J Pediatr Gastroenterol Nutr. 2015 Nov 5.doi: 10.1097/MPG.0000000000001029.
  13. Plasencia C, Jurado T, Villalba A, Peitedado D, Casla MT, Nuño L, Bonilla MG, Martínez-Feito A, Martín-Mola E, Pascual-Salcedo D, Balsa A. Effect of Infliximab Dose Increase in Rheumatoid Arthritis at Different Trough Concentrations: A Cohort Study in Clinical Practice Conditions. Front Med (Lausanne). 2015 Oct 8;2:71. doi: 10.3389/fmed.2015.00071.
  14. Van Stappen T, Billiet T, Vande Casteele N, Compernolle G, Brouwers E, Vermeire S, Gils A. An Optimized Anti-infliximab Bridging Enzyme-linked Immunosorbent Assay for Harmonization of Anti-infliximab Antibody Titers in Patients with Inflammatory Bowel Diseases. Inflamm Bowel Dis. 2015; 21(9):2172-7.
  15. Elberdín L, Outeda M, Salvador P, Paradela S, Fernández-Torres RM, Iglesias R, Fonseca E, Martín I. Infliximab drug and antibody levels in patients with dermatological conditions. Int J Clin Pharm. 2015; 37(2):320-6.
  16. Van Stappen T, Brouwers E, Tops S, Geukens N, Vermeire S, Declerck PJ, Gils A Generation of a Highly Specific Monoclonal Anti-Infliximab Antibody for Harmonization of TNF-Coated Infliximab Assays. Ther Drug Monit. 2015; 37(4):479-85.
  17. Steenholdt C, Bendtzen K, Brynskov J, Thomsen OØ, Ainsworth MA Clinical implications of measuring drug and anti-drug antibodies by different assays when optimizing infliximab treatment failure in Crohn’s disease: post hoc analysis of a randomized controlled trial. Am J Gastroenterol. 2014; 109(7): 1055-64.
  18. Marits P, Landucci L, Sundin U, Davidsdottir L, Nilsson J, Befrits R, Wikström AC, Eberhardson M Trough s-infliximab and antibodies towards infliximab in a cohort of 79 IBD patients with maintenance infliximab treatment. J Crohns Colitis. 2014; 8(8):881-9.
  19. Steenholdt C, Ainsworth MA, Tovey M, Klausen TW, Thomsen OO, Brynskov J, Bendtzen K Comparison of techniques for monitoring infliximab and antibodies against infliximab in Crohn’s disease. Ther Drug Monit. 2013; 35(4):530-8.
  20. Wang SL, Ohrmund L, Hauenstein S, Salbato J, Reddy R, Monk P, Lockton S, Ling N, Singh S Development and validation of a homogeneous mobility shift assay for the measurement of infliximab and antibodies-to-infliximab levels in patient serum. J Immunol Methods. 2012; 382(1-2):177-88.>

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