Ranibizumab Specific ELISA Kit

Catalog No. ABIN5684029

This Colorimetric ELISA kit is designed for the quantitative measurement of Human Ranibizumab Specific.

Quick Overview for Ranibizumab Specific ELISA Kit (ABIN5684029)

Target: Ranibizumab Specific

Reactivity: Human

Detection Method: Colorimetric

Method Type: Sandwich ELISA

Detection Range: 0 ng/mL - 300 ng/mL

Application: ELISA

Sample Type: Humor aquosus

Product Details

Minimum Detection Limit: 0 ng/mL

Purpose: Enzyme immunoassay for the specific and quantitative determination of free Ranibizumab in aqueous humour. Not for use in serum or plasma.

Analytical Method: Quantitative

Specificity: The solid phase (MTP) is coated by a highly specific monoclonal antibody directed against Ranibizumab. Therefore any cross reactivity to the other therapeutical monoclonal antibodies is excluded except for Bevacizumab.

Cross-Reactivity (Details): There is no cross reaction with any other proteins present in native human aqueous humour samples. A screening test was performed with different aqueous humour samples. All produced OD450/620 nm values (ranged from 0.016 to 0.023) less than the mean OD (0.134) of standard D (1 ng/mL). In addition, binding of Ranibizumab to the solid phase is inhibited by recombinant human VEGF.
Therefore, the ImmunoGuide Ranibizumab ELISA (mAb-Based) measures the biologically active free form of Ranibizumab, i.e. not pre-occupied by human VEGF antigen. No cross reaction was observed with the other therapeutic antibodies including Infliximab, Rituximab, Veolizumab, Trastuzumab, Nivolumab and Aflibercept at concentrations tested up to 50 μg/mL. All produced mean OD450/620 nm values (ranged from 0.052 to 0.068) less than standard D.
Because the 2A1 mAb, used for coating solid phase, is also reactive against Bevacizumab, it causes full cross reaction. But a quantification of Bevacizumab in aqueous homour by this testkit is possible only by using the drug-specific standards (i.e. Bevacizumab).

Sensitivity: 0.33 ng/mL

Characteristics: The Ranibizumab ELISA (mAb-based) measures the free form of Ranibizumab.

Components: Microtiter ELISA Plate
Ranibizumab Standards A-E, Concentrate (10X)
Assay Buffer
Dilution Buffer, Concentrate (5X)
Biotinylated alpha-hKappa mAb
Enzyme Conjugate
TMB Substrate Solution
Stop Solution
Wash Buffer, Concentrate (20x)
Adhesive Seal

Material not included: 1. Micropipettes (< 3 % CV) and tips to deliver 5-1000 μL.
2. Bidistilled or deionised water and calibrated glasswares (e.g. flasks or cylinders).
3. Wash bottle, automated or semi-automated microtiter plate washing system.
4. Microtiter plate reader capable of reading absorbance at 450 nm (reference wavelength at 600-650 nm is optional).
5. Absorbent paper towels, standard laboratory glass or plastic vials, and a timer.

Application Details

Application Notes: This kit has been developed for the measurement of drug levels in research and diagnostic uses. It is suitable for Therapeutical Drug Monitoring (TDM) purposes.

Comment:

The Ranibizumab ELISA (mAb-based) is suitable also for being used by an automated ELISA processor.

Sample Volume: 5 μL

Plate: Pre-coated

Protocol: This ELISA is based on a Ranibizumab-specific monoclonal antibody (catcher Ab, clone 2A1). Diluted standards and samples are incubated in the microtiter plate coated with IG-2A1 mAb. After incubation, the wells are washed. A biotinylated anti-human kappa light chain monoclonal antibody (clone 7G8 Z) is added and binds to the kappa light chain part of Ranibizumab. Following incubation, wells are washed, and the horseradish peroxidase (HRP)-conjugated streptavidin is added and binds to the biotinylated 7G8(Z) mAb. Following incubation, wells are washed and the bound enzymatic activity is detected by addition of chromogen-substrate. The colour developed is proportional to the amount of Ranibizumab in the sample or standard. Results of samples can be determined by using the standard curve. Preincubation of Ranibizumab with recombinant human VEGF inhibited the reaction. Therefore, the Ranibizumab ELISA (mAb-based) measures the free form of Ranibizumab.

Assay Precision: Intra-assay CV: <10%.
Inter-assay CV: <10%

Restrictions: For Research Use only

Handling

Storage: RT,4 °C

Storage Comment: 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.

Expiry Date: 24 months

Target Details

Target: Ranibizumab Specific

Background: The drug Ranibizumab (trade name Lucentis®) is a recombinant humanized IgG1 kappa isotype monoclonal antibody fragment (Fab) designed for intraocular use. Ranibizumab binds to and inhibits the biologic activity of human vascular endothelial growth factor A (VEGF-A). Ranibizumab, which lacks an Fc region, has a molecular weight of approximately 48 kilodaltons. Ranibizumab concentration in aqueous humour peakes the first day after injection of 0.5 mg Ranibizumab into intravitreal site of human eye (range, 36.9-66.1 μg/mL). Nonlinear regression analysis determines an initial peak concentration (cmax) of 56.1 μg/mL and an elimination half-life (t1/2) of 7.2 days. In animal studies, following intravitreal injection, Ranibizumab was cleared from the vitreous with a half-life of approximately 3 days. In animal studies, systemic exposure of Ranibizumab was more than 2000-fold lower than in the vitreous. According to the data indicated in the prescribring information the pre-treatment incidence of immunoreactivity to Ranibizumab is 0 % -5 % across treatment groups. After monthly dosing with Ranibizumab for 6 to 24 months, antibodies to Ranibizumab are detected in approximately 1 %-9 % of patients. It might be beneficial to identify biomarkers for (non-) response and risk factors for adverse drug reactions that might be related to drug levels and maintaining the effective concentration in order to potentially avoid some side effects with a reliable method.