P.pastoris HCP ELISA Kit (One-step ELISA) HYI114
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 P.pastoris cells. A sheep polyclonal antibody specific to P.pastoris 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 with anti-P.pastoris HCP antibody were simultaneously added to the microtiter plate 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 colored 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 450 nm). The absorbance values at 450 nm wavelength was positively correlated with the HCPs concentration in the Calibration Standard and the samples. The concentration of HCPs in the samples can be calculated using a dose-response curve.
Applications
This kit is intended for use in determining the presence of host cell protein (HCP) contamination in products manufactured with Pichia pastoris (P.pastoris) strains such as GS115 and X33 and so on.
The kit is for RESEARCH USE ONLY and is not intended for clinical use.
Figure 1. Schematic diagram
Components
|
Reagent |
Quantity |
Notes |
|
P.pastoris 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-P.pastoris HCP Microtiter Strips |
8 well ×12 strips |
Strips pre-coated with sheep anti-P.pastoris HCP antibody in a vacuumed bag with desiccant. Seal and store immediately after use. |
|
Reconstitution Solution |
1×1.5 mL |
Only used to dissolve P.pastoris HCP Calibration Standard. |
|
Diluent |
2×25 mL |
For dilution of Calibration Standard, Anti-P.pastoris:HRP(100×) and samples. |
|
Wash Buffer Concentrate (10×) |
2×25 mL |
Dilute 10 times with freshly prepared ultra-pure water to obtain 1×Wash Buffer Solution. |
|
Anti-P.pastoris :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℃.