Host Cell Protein Analysis

Host  Cell Proteins (HCPs) are a heterogeneous, complex group of proteins that  derived from the host cell. HCPs differ significantly in molecular mass,  isoelectric point, hydrophobic properties, and structures. Figure 1 shows a  harvested product pool of bacterial and mammalian fermentation by  two-dimensional difference gel electrophoresis (2D-DIGE). Identification and  quantification HCPs is very important in the development of biopharmaceutical  products to ensure product purity and patient safety. Protease of HCPs can  influence the protein pattern in culture and influence the purification  process. In addition, HCPs can undergo post-translational modifications that  make quantification and characterization more difficult. Moreover, high HCP  variance results in different biochemical properties which could cause an immune  response or allergic reactions even anaphylactic shock.

2D-DIGE silver stain of a harvested product    Figure  1 2D-DIGE silver stain of a harvested product

Although  the host cells contain thousands of HCPs and that could potentially contaminate  the final product and the host cells can be various organisms, such as,  bacteria, yeast and cell lines derived from mammalian or insect species, there  is no explicitly guideline that describes the selection of appropriate methods  for characterization and quantification of HCPs and other impurities for biopharmaceuticals.  Moreover, the regulatory agencies examine each protein drug on a case-by-case  basis to determine appropriate maximum acceptable levels of HCPs and patient  risk.

The  selection of appropriate test methods for identification and/or quantification  of HCP have been a concern during process development and manufacturing. The  methods used for HCP analysis can be classified as either immune-specific  methods, such as, western blotting and ELISA, or non-specific methods, such as,  electrophoresis and MS. In general, a qualitative or semi-quantitative method  would be sufficient for sample purification and for lot release. However,  during process of drug development and characterization, the ideal method not  only should have the capability to recognize all HCPs species, but also should  be quantitative, high-throughput, time-saving and labor-saving.

A  range of methods for the detection and characterization of HCPs are presented  in Creative Proteomics (Table 1). Among these, ELISA, electrophoresis, and MS are  the most common techniques and are sometimes they are used in combination for  detailed characterization.

Table  1, Overview methods of HCPs methods

Overview methods of HCPs methods

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