Tocilizumab mAb-based ELISA Assay


The Tocilizumab mAb-based ELISA Assay is an enzyme immunoassay for the specific and precise measurement of free Tocilizumab 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 Tocilizumab and no other TNF-α therapeutic.  The Eagle Biosciences Tocilizumab ELISA (mAb-based) Assay Kit is for research use only and not to be used in diagnostic procedures.

Tocilizumab mAb-based ELISA Assay

The Tocilizumab mAb-based ELISA Assay is For Research Use Only

Size: 1 x 96 wells
Sensitivity: 2 ng/mL
Dynamic Range: 6 – 200 ng/mL
Incubation Time: 2.5 hours
Sample Type: Serum, Plasma
Sample Size: 10 µL
Alternative Names: Actemra

Assay Background

The drug Tocilizumab (trade name Actemra®) is a humanized monoclonal antibody (mAb) that specifically targets both soluble interleukin-6 receptor (sIL-6R) and membrane bound interleukin-6 receptor (mIL-6R) with high affinity, thereby preventing pro-inflammatory effects of IL-6. The specificity of this testsystem is achieved by using a monoclonal antibody (clon IG-7C84) for the coating of the microtiter plate and also as a conjugate.
This Eagle Biosciences Tocilizumab ELISA (mAb-based) is developed for the specific measurement of Tocilizumab in sera, plasma and other biological fluids by the advantage of using a site-directed IG-7C84 mouse monoclonal antibody (mAb) specific for Tocilizumab only. Binding of Tocilizumab to the solid phase, pre-coated with IG-7C84, is inhibited by recombinant human interleukin-6 receptor alpha (rh-IL6Ra) in a concentration dependent manner. Therefore, this Tocilizumab ELISA (mAb-Based) measures the free form of Tocilizumab. The choice of specifically measuring the free form allows investigators to analyze the concentration-effect relationship. Due to the use of a mAb as catcher and also as detection reagent this test kit has an unmatched specificity.

The kit is shipped at ambient temperature and should be stored at 2-8°C. Keep away from heat or direct sun light. The storage and stability of specimen and prepared reagents is stated in the corresponding chapters. The microtiter strips are stable up to the expiry date of the kit in the broken, but tightly closed bag when stored at 2–8°C.

Related Products

Tocilizumab (Actermra) ELISA Kit
Anti-Tocilizumab (Actermra) ELISA Assay
Omalizumab mAb Based ELISA

Additional Information

Assay Principle

This ELISA is based on a Tocilizumab-specific monoclonal antibody (catcher Ab, ImmunoGuide clone IG-7C84). Diluted standards and diluted samples are incubated in the microtiter plate coated with IG-7C84 mAb. After incubation, the wells are washed. A horseradish peroxidase (HRP)-conjugated anti-Tocilizumab antibody is added and binds to the Tocilizumab. Following incubation, the wells are washed and the bound enzymatic activity is detected by addition of chromogen-substrate. The colour developed is proportional to the amount of Tocilizumab in the sample or standard. Results of samples can be determined by using the standard curve. Binding of Tocilizumab to the solid phase, pre-coated with IG-7C84, is inhibited by recombinant human interleukin-6 receptor-α (rh-IL6Rα) in a concentration dependent manner. Therefore, the ImmunoGuide Tocilizumab ELISA (mAb-Based) measures the free form of Tocilizumab.

Assay Procedure

  1. Pipette 100 µL of Assay Buffer into each of the wells to be used.
  2. Pipette 75 µL of each 1:10 Diluted Standard, and 1:1000 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 Tocilizumab) into each well.
  6. Cover plate with adhesive seal. Shake plate carefully. Incubate 60 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.

Typical Standard Curve

Tocilizumab mAb-based ELISA Assay


Product Manual



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