Vero HCP ELISA Kit (One-step ELISA) HYI113
This kit is based on the solid-phase enzyme-linked immunosorbent assay (ELISA) with a double-antibody sandwich technique to detect residual host cell proteins (HCPs) from vero cells. A sheep polyclonal antibody specific to vero HCPs was employed in the assay to capture any remaining HCPs in the sample. The antibody coverage is assessed by the current mainstream method. Both the Calibration Standard (or test sample) and the HRP (Horseradish Peroxidase) labeled anti-Vero HCP antibody were simultaneously added to the microtiter plate, which coated with the affinity purified capture antibody and followed by incubation and washing. Then TMB (3,3',5,5'-tetramethylbenzidine) substrate was added into reaction, HRP catalyzed the oxidation of TMB by H2O2 to produce a blue product (maximum absorption peak at 655 nm). Then the stop solution was added to terminate the enzymatic reaction, resulting in a yellow colored product (maximum absorption peak at 450nm). The absorbance values at 450nm wavelength were positively correlated with the HCPs concentration in the Calibration Standard and the sample. The concentration of HCPs in the sample can be calculated using the dose-response curve.
Applications
This kit is suitable for the quantitation of Vero cell derived host cell proteins (HCPs) in various bioproducts, produced by sonication cell lysis. The kit is for RESEARCH USE ONLY and not intended for clinical use.
Figure 1. Schematic diagram
Components
|
Reagent |
Quantity |
Notes |
|
Vero HCP Calibration Standard |
2 bottles |
Lyophilized powder. Dissolve it with the reconstitution solution (500 μL), and let it stand for about 5 minutes until transparent. Please refer to the details on the label of the tube. |
|
Anti-Vero HCP Microtiter Strips |
8 well ×12 strips |
Strips pre-coated with sheep anti-Vero HCP affinity antibody in a vacuumed bag with desiccant. Seal and store immediately after use. |
|
Reconstitution Solution |
1×1.5 mL |
Only used to dissolve Vero HCP Calibration Standard. |
|
Diluent |
2×25 mL |
For dilution of Calibration Standard, Anti-Vero:HRP (100×) and samples. |
|
Wash Buffer Concentrate (10×) |
2×25 mL |
Easy to be crystallized at low temperature, please incubate at 37℃ in water bath before use. Dilute 10 times with freshly prepared ultra-pure water to obtain 1×Wash Buffer Solution. |
|
Anti-Vero:HRP (100×) |
1×120 μL |
Affinity purified sheep antibody conjugated to HRP in a protein matrix with preservative. Dilute 100 times in diluent before use. |
|
TMB Substrate |
1×12 mL |
Sealed and keep away from light. Equilibrate to room temperature (RT) for 20 minutes before use. |
|
Stop Solution |
1×6 mL |
1 M hydrochloric acid. Avoid direct contact with eyes, skin, and clothing. |
|
Sealing Film |
3 pieces |
Cover the strips with it during incubation to prevent contamination and liquid evaporation. |
Note: Room temperature refers to 25±3℃.
Storage Conditions
Store the kit at 2-8℃. Use within the expiration date labeled upon the kit package. The opened components should be stored as follows.
Table 2. Recommended storage conditions for opened components
|
Component |
Stability |
|
Anti-Vero HCP Microtiter Strips |
Store in the bag with desiccant at 2-8℃ for up to 30 days. |
|
Reconstituted Vero HCP Calibration Standard |
Store at -20℃ , freeze and thaw for no more than 3 times. |
Materials Required But Not Supplied in the Kit
1.Sterile centrifuge tubes for dilution
2.Absorbent paper for plate drying
3.Pipette Tips
4.Multi-channel reagent reservoirs (50 mL)
Related equipment
1.Microplate reader capable of measuring absorbance at 450 nm, with the correction wavelength set at 620 nm to 650 nm.
2.Single or multi-channel pipettes
3.Microplate thermoshaker
4.Incubator (optional)
5.Plate washer (optional)