CHO HCP ELISA Kit

A One-step immunosorbent ELISA method is used in this assay. Samples containing CHOK1 HCP simultaneously react with HRP-labeled goat anti-CHOK1 antibody and anti-CHOK1 antibody coated on the ELISA plate, finally forming a sandwich complex of solid-phase antibody-HCP-labeled antibody. Unbound antigen-antibody can be removed by washing the ELISA plate. The TMB substrate is added to the well for sufficient reaction. The color development is stopped after adding the stop solution, and the OD or absorbance value of the reaction solution at 450/650nm is read with a microplate reader. The OD value or absorbance value is proportional to the HCP content in the solution. From this, the HCP concentration in the solution can be calculated according to the standard curve.

Application

This kit is used to quantitatively detect the content of CHOK1 host cell protein residues in samples.

Components

Equipment Required

Storage and stability

1.Transport at -25~-15°C.

2.Storage conditions are as shown in Table 1; components 1-2 are stored ≤–20°C,5-6 are stored RT,3、4、7、8 are stored at 2-8℃; the validity period is 12 months.

Product parameters

1. Sensitivity: 1ng/mL

2. Detection range:3- 100ng/mL

3. Precision: Intra-assay CV≤ 10%, inter- assay CV≤ 15%

4. HCP coverage: >80%

5. Specificity: This kit is universal as it specifically reacts with CHOK1 HCP independent of purification process.

Reagent preparation

1. PBST 0.05%

Take 15 ml of 20×PBST 0.05%, diluted in ddH₂O, and made up to 300 ml.

2. 1.0% BSA

Take 1g of BSA from the bottle and dilute in 100 ml of PBST 0.05%, mix well until completely dissolved, and store at 2-8°C. The prepared dilution buffer is valid for 7 days. It is recommended to prepare as needed.

3. Detection solution 2μg/mL

Take 48μL of 0.5 mg/mL Anti-CHO HCP-HRP and dilute in 11,952μL of 1% BSA to obtain a final concentration of 2μg/mL detection solution.

4. QC and Preparation of CHOK1 HCP Standards

 Table: Preparation of QC and Standards 

Assay Procedure

1. Prepare the reagents as indicated in “Reagent Preparation” above.

2. Take 50μL of standards, samples and QCs (refer to Table 3) into each well, then add 100μL of Detection solution (2μg/mL); Cover plate with sealer, and place the ELISA plate on the thermomixer. Incubate at 500rpm, 25±3℃ for 2 hours.

3. Invert the microplate in the sink and discard the coating solution. Pipette 300μL of PBST 0.05% into each well to wash the ELISA plate and discard the solution, and repeat the washing 3 times. Invert the plate on a clean paper towel and pat dry.

4. Add 100μL ofTMB substrate (see Table 1) to each well, seal the ELISA plate, and incubate in the dark at 25±3℃ for 15 min.

5. Pipette 100μL of stop solution into each well.

6. Measure absorbance at wavelength of 450/650nm with microplate reader.

7. Analyze data by SoftMax or equivalent software. Plot standard curve by using a four-parameter logistic regression model.

Standard Curve Example

NOTE: If the concentration of HCP in the sample exceeds the upper limit of the standard curve, it needs to be properly diluted with dilution buffer before testing.

NOTES

The stop solution is 2M sulfuric acid, please handle with care to avoid splashing!