A cell line is a population of cells grown and maintained in a laboratory. Cell line expansion increases the number of these cells from a small initial sample to a large, uniform population. This process provides a consistent and reproducible source of biological material, widely utilized across biological research and biotechnology.
Understanding Cell Line Expansion
The objective of cell line expansion is to obtain enough cells for multiple experiments or as starting material for various applications. This process ensures studies are performed on genetically similar cells, promoting consistent results.
Different types of cell lines are used for expansion. Immortalized cell lines, such as HeLa or Chinese hamster ovary (CHO) cells, proliferate indefinitely due to genetic modifications or cancerous origin. They are frequently expanded because of their robust growth and stability. In contrast, primary cells are isolated directly from tissues and have a limited lifespan, undergoing only a finite number of divisions. Adherent cells, which attach and grow on a surface, are handled differently from suspension cells, which grow freely in liquid media, with each type requiring specific expansion methods.
The Process of Cell Line Growth
Growing cells in a laboratory requires precise control over several environmental factors to support their growth and proliferation. Cells are maintained in nutrient-rich media, which supply essential amino acids, vitamins, and salts necessary for their metabolism and division. The pH of the medium is carefully regulated, maintained between 7.2 and 7.4 for most mammalian cells, often using a carbon dioxide-bicarbonate buffering system.
Temperature is another parameter controlled, with 37°C being optimal for most mammalian cell lines, mimicking physiological conditions. Carbon dioxide (CO2) levels, around 5%, are maintained in incubators to regulate the medium’s pH. Sterile conditions are essential to prevent contamination, requiring specialized equipment like biological safety cabinets (laminar flow hoods) and incubators.
The expansion process begins with initial seeding, where a small number of cells are transferred into a new culture vessel containing fresh medium. As cells grow and divide, they consume nutrients and produce waste, eventually reaching a state called confluency, where they cover the available surface or reach a high density in suspension. At this point, cells are “subcultured” or “passaged” by detaching them (for adherent cells, often using enzymes like trypsin) and diluting them into new vessels with fresh media, allowing continued proliferation. This serial passaging ensures cells remain in an active growth phase and do not become overcrowded.
Maintaining Cell Line Quality
Maintaining cell line quality during expansion is essential for reliable scientific outcomes. Contamination is a persistent threat, with bacteria, fungi, and mycoplasma being common culprits. Mycoplasma, in particular, are problematic as they lack a cell wall, making them resistant to common antibiotics, and their presence may not be visibly obvious. Strict aseptic techniques, regular cleaning of workspaces and equipment, and routine testing for contaminants are employed to mitigate these risks.
Genetic stability is another consideration, as cells can undergo genetic changes or “genetic drift” during prolonged culture, especially with increasing passage numbers. Such alterations can lead to changes in cell morphology, growth rates, gene expression, and overall behavior, impacting experimental reproducibility. To minimize these issues, researchers monitor cell morphology, track passage numbers, and aim to use low-passage cells (fewer than 15 passages) for experiments.
Cell line identity verification is also performed to confirm cell identity, often using techniques like Short Tandem Repeat (STR) profiling for human cell lines. Cryopreservation, the process of freezing cells at ultra-low temperatures, is extensively used to create master and working cell banks. This practice preserves the genetic and phenotypic characteristics of cell lines, allowing researchers to return to early-passage, high-quality stock as needed.
Why Cell Line Expansion Matters
Cell line expansion is a foundational technique that supports advancements across science and medicine. In basic research, expanded cell lines serve as models to investigate fundamental cell biology, elucidate disease mechanisms, and understand gene function. They provide a controlled environment to study cellular processes without the complexities of a whole organism.
In drug discovery and development, expanded cell lines are used extensively for high-throughput screening of potential drug candidates and for toxicity testing. This allows researchers to rapidly assess how new compounds affect human or animal cells, accelerating the development of new therapies. In biotechnology and biomanufacturing, large quantities of expanded cells are used for the production of vaccines, therapeutic proteins, and antibodies. For instance, Chinese hamster ovary (CHO) cells are widely employed to produce recombinant proteins due to their robust growth and protein production capabilities.
Expanded cell lines also play a significant role in regenerative medicine and cell therapies. Stem cell research, for example, relies on expanding specific stem cell populations to develop treatments for tissue repair and regeneration. The ability to generate vast numbers of uniform cells from a single source enables both fundamental scientific inquiry and the development of innovative medical treatments.