The Protein Man's Blog | A Discussion of Protein Research

Reporters have contributed significantly to the field of biology and its progress. Genetic reporters are widely used in drug development, biomedical research, and biochemistry. In a nutshell, the reporter gene is fused with the gene of interest and cloned into an expression vector. The expression vector with the chimeric sequence is transferred into suitable cells. The expression of the gene in the cell is measured by the activity of the reporter protein. A textbook good reporter gene should have high sensitivity and can be measured quantitatively after expression.

Determination of protein concentration is necessary and widely used in protein biology, molecular biology, and other research applications. The concentration of protein samples have to be estimated before proceeding to isolation, purification, and analysis. Various protein estimation methods have been developed so far. Each method has its own advantages and disadvantages over others. Most of these methods depend on the levels of tryptophan, tyrosine and other aromatic amino acids.

Host cell proteins (HCPs) are one of the main impurities in bio-therapeutics that are carried over from the host expression system. Although HCPs are at low levels, they can trigger immunogenic reactions in the human system when administered along with bio-therapeutics. The similarity of physicochemical properties of HCPs and therapeutic proteins result in co-purification. Detection and elimination of these HCPs during bio-therapeutics purification are required. Development of analytical tools and methods for the detection of these HCPs during protein purification and quality control is a challenge. The final threshold level of HCPs varies for different bio-therapeutics, so manufacturers have
to analyze and report the HCP levels at all stages of downstream processing before the
product is released according to ICH-Q6B guidelines.

Maintaining the stability and solubility of target proteins before, during and after purification is one of the biggest challenges researchers often run into. Proteins are extremely sensitive to solution conditions (i.e., pH and conductivity) and temperature. They also tend to aggregate at high concentrations required for structural studies. This can be a serious problem since protein aggregation can cause artifacts and hamper the biological activity of the target protein.