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Tips for Maximizing Protein Expression in Mammalian Cells


Tips for Maximizing Protein Expression in Mammalian Cells

Producing mammalian-based proteins sometimes requires a specified mammalian host cell system. Therapeutic and antibody protein applications commonly rely on mammalian cell systems for production. Understanding how to maximize protein expression in mammalian cells can make a difference in yields and potency. We examine these efforts below.


Understand the Selection of Signal Peptides

The signal peptide is responsible for secreting the proteins from mammalian cells. This is a critical first step in expressing mammalian cells, and a few signal peptides are available. Depending on the protein of interest, the lab may make a suggestion on the designated signal peptide. Otherwise, common options available include the CD33 leader sequence, IL-2, and the in-house developed signal peptide.


Use Fusion Tags

Fusion tags are another avenue to explore when maximizing the expression of mammalian cells. Labs use professional discernment to determine the addition or position of a designated tag based on the intended expression applications. The most common fusion tags for mammalian expression systems are His, FLAG, and Fc.


His tags are polyhistidine tags, and the composition is a sequence between 6 and 10 histidine residues. The FLAG tag is the most common epitope tag for unnatural proteins. FLAG octapeptide is an artificially curated tag and is best for detection applications. Fc tags are fusion proteins known as chimeric fusion proteins. They are genetically linked to the protein of interest and best serve in vivo and in vitro applications.


Optimize Transfection Conditions

Optimizing transfection conditions helps regulate cell viability and creates a pathway for maximum expression values. The primary objective of the optimization is to find the most appropriate balance for the transfected cells and gain an understanding of the best range of conditions to achieve this.


The cells should remain healthy at 75 to 90 percent confluency during transfection. This percentage should exceed 95 percent during the physical harvest. Monitoring the passage values and the cell’s biological response is vital because any unreliability will require an adjustment.


Consider Stable Cell Lines

Stable cell lines offer several advantages for protein expression in mammalian cell systems. Through gene integration and selection, stable cell lines ensure the presence of the gene of interest in all daughter cells, resulting in consistent protein expression over time. Gene amplification techniques can further increase gene copy numbers, leading to higher protein expression levels in mammalian cells.


Additionally, stable integration can induce epigenetic modifications that enhance transcriptional activity and overall protein production. By clonally isolating high expressers, stable cell lines reduce clonal variability and provide a more uniform protein expression across the cell population. With the ability to adapt and optimize over time, stable cell lines offer a reliable platform for achieving high protein expression levels and minimizing variability in mammalian cell systems.


Bon Opus Biosciences offers complete mammalian cell protein expression services with His or Fc fusion tags and a six-to-eight-week turnaround time. Our team of industry experts works diligently and transparently through each step, ensuring the yields exceed your demands. Request your quote today!

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