Intended Use
This kit is intended for use in determining the presence of host cell protein (HCP) impurities in products manufactured by expression in the Chinese Hamster Ovary (CHO) cell line. This generic assay is a method to aid in optimal purification process development, process control, routine quality control, and product release testing. The kit is for Research and Manufacturing Use Only and is not intended for diagnostic use in humans or animals.
Principle of the Procedure
This assay is based on a sandwich ELISA to be performed in a microplate format. Sample potentially containing CHO HCP are incubated in microtiter plate wells which have been pre-coated with the affinity-purified CHO
Intended Use
This kit is intended for use in determining the presence of host cell protein (HCP) impurities in products manufactured by expression in the Chinese Hamster Ovary (CHO) cell line. This generic assay is a method to aid in optimal purification process development, process control, routine quality control, and product release testing. The kit is for Research and Manufacturing Use Only and is not intended for diagnostic use in humans or animals.
Principle of the Procedure
This assay is based on a sandwich ELISA to be performed in a microplate format. Sample potentially containing CHO HCP are incubated in microtiter plate wells which have been pre-coated with the affinity-purified CHO
Reagents and Materials Provided
Storage and Stability
- 1) Unopened kit should be stored at 2℃ to 8℃ for stability until the expiration date printed on the kit.
- 2) Opened reagents should be stored at 2℃ to 8℃ and are stable for 6 weeks. Opened ELISA microplate should be sealed in foil bag with desiccant.
Equipment required
- 1) Microplate reader spectrophotometer with dual wavelength capability at 450nm (detection wavelength) and 650nm (reference wavelength).
- 2) Thermomixer or equivalent (400-600 rpm, 37℃).
Reagent Preparation
Bring all reagents to room temperature (25℃±3℃) for 20 minutes before use. Remaining reagents should be store at 2℃ to 8℃ immediately after use.
- 1) Prepare washing buffer (1×) by diluting 50mL of the Wash concentrate (10×) with 450mL of deionized or distilled water.
- 2) Dilute the Standard (0.01mg/mL) with dilution buffer to 100ng/mL, 50ng/mL, 25ng/mL, 12ng/mL, 6ng/mL, 3ng/mL, 0.7ng/mL, 0ng/mL.
Standards (100ng/mL to 0ng/mL) are prepared according to the following dilution scheme:
- 3) Detector antibody (1×) : Dilute to work concentrate at 1:99 by dilution buffer before use.
- 4) Enzyme conjugate (1×) : Dilute to work concentrate at 1:99 by dilution buffer before use.
Assay Procedure
- 1) Take out the required strips form the foil of the ELISA microplate after reaching room temperature. Remaining plate strips not used in
the assay should be sealed in foil bag with desiccant and stored at 2℃ to 8℃.
- 2) Prepare samples and standards in advance.
- 3) Pipette 100μL of standards or samples into each well. We recommend that assays are carried out in duplicate in order to minimize spurious results. Cover and incubate on shaker at 500rpm for 60 minutes at 37℃.
- 4) Prepare washing buffer (1×) and detector antibody (1×) in advance.
- 5) Dump contents of the wells. Blot and gently but firmly tap over absorbent paper to remove most of the residual liquid. Fill wells generously
with 300μL/well of washing buffer (1×). Dump and tap again. Repeat for a total of 5 washes.
- 6)Pipette 100μL of the detector antibody (1×) into each well. Cover and incubate on shaker at 500rpm for 30 minutes at 37℃.
- 7) Prepare enzyme conjugate (1×) in advance.
- 8) Dump contents of the wells. Blot and gently but firmly tap over absorbent paper to remove
most of the residual liquid. Fill wells generously with 300μL/well of washing buffer (1×). Dump and tap again. Repeat for a total of 5 washes.
- 9) Pipette 100μL of the enzyme conjugate (1×) into each well. Cover and incubate on shaker at 500rpm for 30 minutes at 37℃.
- 10) Bring required substrate solution to room temperature (25℃±3℃) for 20 minutes before use and protect from light