Proteins & Peptides Residual Host Cell Protein (HCP) Analysis

Residual host cell protein (HCP) is a major class of process-related impurities that regulatory agencies require manufacturers to demonstrate have been eliminated during the development of protein/peptide biologics to ensure drug purity, manufacturing process consistency, and patient safety. CD Formulation uses a range of highly sensitive methods to identify and quantify residual HCP under global regulatory requirements to ensure that products meet specification limits for these process residues and support process validation and GMP batch release.

Residual HCP & Why Detect Residual HCP?

Host cell protein (HCP) is residual protein impurities expressed with the desired recombinant biotherapeutic product. During the expression of recombinant protein drugs, host cell systems (e.g., Chinese Hamster Ovary (CHO) cells) express many endogenous proteins, such as nucleotides and proteins. These endogenous endogenous proteins are called HCP. The presence of HCPs in the final drug product can lead to decreased stability, immunogenicity, and toxicity to patients, thus posing a significant safety risk. According to regulatory guidance ICH Q6B, residual HCP must be detected and quantified to ensure that the manufactured biopharmaceutical meets quality standards and safety requirements.

Regulatory agencies point out that HCP testing is a tool to verify process consistency, and manufacturers should detect most HCP in early processing and must demonstrate the removal of HCP during purification and reduce them to acceptable levels. The US FDA recommends a range of 1-100 ppm.

Fig. 1 Objectives of HCP assays for monitoring HCPs in biologics. (Pilely K, et al., 2022)

Explore Our Proteins & Peptides HCP Analysis Services

Monitoring HCPs is a critical quality attribute (CQA) in the production of biopharmaceuticals such as recombinant proteins. As part of our quality control testing program, we provide our global customers unparalleled residual HCP testing services to demonstrate that the HCP levels of your protein/peptide biopharmaceuticals do not exceed regulatory guidelines and are effectively purified during the manufacturing process.

Our HCP analysis team provides a range of GLP/GCP/GCMP compliant methods such as ELISA, LC-MS, and gel electrophoresis to identify and quantify total HCP levels, allowing for monitoring downstream processes and support batch release and comparability studies in accordance with regulatory requirements such as ICH Q6B / FDA.

Our HCP analysis team has expertise in developing residual protein assays for any cell type (HEK293, E. coli, and CHO cells), providing customized HCP analysis solutions based on your needs.

Our services include but are not limited to:

• HCP analytical method development and optimization.
• HCP analytical method validation.
• HCP detection and qualitative analysis.
• HCP accurate quantitative analysis.
• Process validation support.
• Batch release testing support.
• Biosimilar studies support.
• Comparability studies support.

How Do We Detect Residual HCP?

At CD Formulation, we provide powerful quantitative analysis using real-time enzyme-linked immunosorbent assay (ELISA), high-performance liquid chromatography-mass spectrometry (LC-MS), gel electrophoresis, and western blot to support process validation, monitor batch-to-batch variability, and support GMP batch release.

ELISA for HCP Analysis

ELISA is the industry standard for quantitative HCP analysis in biopharmaceutical products and process development and is currently the most widely used HCP detection method. Our HCP-ELISA analysis services support batch release testing of bulk drug substances as well as analysis of downstream process performance.

ELISA has a wide HCP coverage, providing total HCP measurements (ng/mg). However, due to the lack of flexibility, the lack of any detailed information on the characterization of individual host cell protein HCP, and the dependence on antibodies, it is difficult to fully detect all HCPs present in the sample.

LC-MS for HCP Analysis

For regulatory filings, LC-MS has been an orthogonal method to support and complement ligand binding assays (LBA). This method can identify and quantify individual HCP as well as total HCP content.

Our HCP analysis team uses LC-MS-based methods to identify and quantify individual HCP present in different stages and final products. In addition, it is also used in conjunction with commercial ELISA kits to increase coverage so that you can detect more host cell proteins present in your product.

Fig. 2 Workflow of LC-MS-based host cell protein (HCP) analysis. (CD Formulation)

Gel Electrophoresis for HCP Analysis

Gel electrophoresis analysis involves the separation of HCP into individual components by gel electrophoresis followed by western blot analysis. This method is used to qualitatively analyze HCP components, which is a powerful tool for separating and characterizing HCP.

Our HCP analysis team utilizes different two-dimensional electrophoresis (2DE) methods, including traditional 2D SDS-PAGE with silver and SYPRO Ruby fluorescent dye staining as well as two-dimensional difference gel electrophoresis (2D-DIGE), provide the most comprehensive HCP antibody coverage analysis to support the characterization of complex mixtures such as HCP.

In our analytical laboratory, 2D gel electrophoresis is routinely used for coverage determination and analysis to ensure the identification of any HCP in downstream processes and final products.

Validation of Quantitative Methods for Residual HCP

ICH guidelines state that all quantitative assays must be validated. Based on ICH guidelines, we validate HCP assays using standard matrices, including:

• Precision
• Accuracy
• Limit of Detection (LOD)
• Limit of Quantification (LOQ)
• Linearity
• Specificity

Why Choose Us for Proteins & Peptides Residual HCP Analysis？

• We perform HCP analysis using a variety of analytical methods including ELISA, LC-MS, and gel electrophoresis to support your protein/peptide biologics development program.
• Our team of experts has many years of experience in protein and peptide analysis, especially in the detection and quantification of HCP.
• We are able to identify and quantify HCP at very low concentrations.
• Our laboratory facilities and analytical methods adhere to strict regulatory guidelines, ensuring that our results are reliable and meet industry standards.
• We offer flexible experimental design options and customized solutions to meet the needs of any of your projects.
• We adhere to strict quality standards and regulatory guidelines to ensure that all testing is conducted in accordance with industry best practices.

Publication

Residual HCP testing is a critical component to ensure safety, efficacy, and consistency in the development and manufacture of therapeutic proteins and peptides. Please feel free to contact us if you are interested in our services. Learn how CD Formulation can help you conduct HCP analysis and verify the safety of your products in relation to regulatory requirements.

How It Works

STEP 1

Submit a service request describing your specific research or development need.

STEP 2

We'll email you to provide your quote and confirm order details if applicable.

STEP 3

Execute the project with real-time communication, and deliver the final report promptly.