Introduction to Cell Culture

Cell culture refers to the removal of cells from an animal or plant and their subsequent growth in a favorable artificial environment. The cells may be removed from the tissue directly and disaggregated by enzymatic or mechanical means before cultivation, or they may be derived from a cell line or cell strain that has already been established.

While cell culture can be challenging due to the need for sterile conditions and precise control over environmental factors, it is a fundamental skill in many areas of biological research and biotechnology. There are three main types of cell culture: primary culture, where cells are directly isolated from tissues; secondary culture, which involves subculturing primary cells; and cell lines, which are either naturally immortalized or genetically modified to proliferate indefinitely. Basic techniques of cell culture include aseptic handling to prevent contamination, preparing and maintaining culture media, subculturing (or passaging) cells to prevent overgrowth, and cryopreservation for long-term storage. Mastering these techniques is essential for ensuring the health and viability of cultured cells and obtaining reliable experimental results.

Primary culture refers to the stage of the culture after the cells are isolated from the tissue and proliferated under the appropriate conditions until they occupy all the available substrate (i.e., reach confluence). At this stage, the cells must be subcultured (i.e., passaged) by transferring them to a new vessel with fresh growth medium to enable more room for continued growth. After the first subculture, the primary culture becomes known as a cell line or subclone. Cell lines derived from primary cultures have a limited life span (i.e., they are finite; see below), and as they are passaged, cells with the highest growth capacity predominate, resulting in a degree of genotypic and phenotypic uniformity in the population. If a subpopulation of a cell line is positively selected from the culture by cloning or some other method, this cell line becomes a cell strain. A cell strain often acquires additional genetic changes subsequent to the initiation of the parent line.

Normal cells usually divide only a limited number of times before losing their ability to proliferate, which is a genetically determined event known as senescence; these cell lines are known as finite. However, some cell lines become immortal through a process called transformation, which can occur spontaneously or can be chemically or virally induced. When a finite cell line undergoes transformation and acquires the ability to divide indefinitely, it becomes a continuous cell line.

While choosing from finite cell lines may give you more options to express the correct functions, continuous cell lines are often easier to clone and maintain.

Read about key starting considerations for cell culture

Culture conditions vary widely for each cell type, but the artificial environment in which the cells are cultured invariably consists of a suitable vessel containing the following:

• A substrate or medium that supplies the essential nutrients (amino acids, carbohydrates, vitamins, minerals)
• Growth factors
• Hormones
• Gases (O₂, CO₂)
• A regulated physico-chemical environment (pH, osmotic pressure, temperature)

Most cells are anchorage-dependent and must be cultured while attached to a solid or semi-solid substrate (adherent or monolayer culture), while others can be grown floating in the culture medium (suspension culture).

Cell lines in continuous culture are likely to suffer undesirable outcomes such as genetic drift, senescence, and microbial contamination, and even the best-run laboratories can experience equipment failure. An established cell line is a valuable resource, and its replacement is expensive and time consuming. Therefore, it is vitally important that they are frozen down and preserved for long-term storage. A properly maintained frozen cell stock is an important part of cell culture.

Read more about cell culture environment
Read more about cell freezing protocols
Read more about cell morphology and culture types

Cell culture is one of the major tools used in cellular and molecular biology, providing excellent model systems for studying the normal physiology and biochemistry of cells (e.g., metabolic studies, aging), the effects of drugs and toxic compounds on the cells, and mutagenesis and carcinogenesis. It is also used in drug screening and development, and large-scale manufacturing of biological compounds (e.g., vaccines, therapeutic proteins). The major advantage of using cell culture for any of these applications is the consistency and reproducibility of results that can be obtained from using a batch of clonal cells.

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