Subculture (biology)
Encyclopedia
In Biology
a subculture is a new cell
or microbiological culture
made by transferring some or all cell
s from a previous culture to fresh growth medium
. This action is called subculturing or passaging the cells. Subculture is used to prolong the life and/or expand the number of cells or microorganisms in the culture.
Typically subculture is from a culture of a certain volume into fresh growth medium of equal volume, this allows long-term maintenance of the cell line. Subculture into a larger volume of growth medium is used when wanting to increase the number of cells for, for example, use in an industrial process or scientific experiment.
tissue cells somaclonal variation may arise over long periods in culture. Similarly in mammal
ian cell lines chromosomal aberrations have a tendency to increase over time. For microorganisms there is a tendency to adapt to culture conditions, which is rarely precisely like the microorganisms natural environment, which can alter their biology.
, the percentage of the growth surface covered by cells. The cells will often have a preferred range of confluencies for optimal growth, for example a mammalian cell line like HeLa
or Raw 264.7 generally prefer confluencies over 10% but under 100%, and subculture will normally try to keep the cells in this range. For subculture cells may be detached by one of several methods including trypsin
treatment to break down the proteins responsible for surface adherence, chelating sodium ions with EDTA
which disrupts some protein adherence mechanisms, or mechanical methods like repeated washing or use of a cell scraper. The detached cells are then resuspended in fresh growth medium and allowed to settle back onto their growth surface.
Biology
Biology is a natural science concerned with the study of life and living organisms, including their structure, function, growth, origin, evolution, distribution, and taxonomy. Biology is a vast subject containing many subdivisions, topics, and disciplines...
a subculture is a new cell
Cell culture
Cell culture is the complex process by which cells are grown under controlled conditions. In practice, the term "cell culture" has come to refer to the culturing of cells derived from singlecellular eukaryotes, especially animal cells. However, there are also cultures of plants, fungi and microbes,...
or microbiological culture
Microbiological culture
A microbiological culture, or microbial culture, is a method of multiplying microbial organisms by letting them reproduce in predetermined culture media under controlled laboratory conditions. Microbial cultures are used to determine the type of organism, its abundance in the sample being tested,...
made by transferring some or all cell
Cell (biology)
The cell is the basic structural and functional unit of all known living organisms. It is the smallest unit of life that is classified as a living thing, and is often called the building block of life. The Alberts text discusses how the "cellular building blocks" move to shape developing embryos....
s from a previous culture to fresh growth medium
Growth medium
A growth medium or culture medium is a liquid or gel designed to support the growth of microorganisms or cells, or small plants like the moss Physcomitrella patens.There are different types of media for growing different types of cells....
. This action is called subculturing or passaging the cells. Subculture is used to prolong the life and/or expand the number of cells or microorganisms in the culture.
Role
Cell lines and microorganisms cannot be held in culture indefinitely due to the gradual rise in toxic metabolites, use of nutrients and increase in cell number due to growth. Subculture is therefore used to produce a new culture with a lower density of cells than the originating culture, fresh nutrients and no toxic metabolites allowing continued growth of the cells without risk of cell death. Subculture is important for both proliferating (e.g. a microorganism like E. coli) and non-proliferating (e.g. terminally differentiated white blood cells) cells.Typically subculture is from a culture of a certain volume into fresh growth medium of equal volume, this allows long-term maintenance of the cell line. Subculture into a larger volume of growth medium is used when wanting to increase the number of cells for, for example, use in an industrial process or scientific experiment.
Passage number
It is often important to record the approximate number of divisions cells have had in culture by recording the number of passages or subcultures. In the case of plantPlant
Plants are living organisms belonging to the kingdom Plantae. Precise definitions of the kingdom vary, but as the term is used here, plants include familiar organisms such as trees, flowers, herbs, bushes, grasses, vines, ferns, mosses, and green algae. The group is also called green plants or...
tissue cells somaclonal variation may arise over long periods in culture. Similarly in mammal
Mammal
Mammals are members of a class of air-breathing vertebrate animals characterised by the possession of endothermy, hair, three middle ear bones, and mammary glands functional in mothers with young...
ian cell lines chromosomal aberrations have a tendency to increase over time. For microorganisms there is a tendency to adapt to culture conditions, which is rarely precisely like the microorganisms natural environment, which can alter their biology.
Protocols for passaging
The protocol for subculturing cells depends heavily on the properties of the cells involved.Non-adherent cells
Many cell types, in particular many microorganisms, grow in solution and not attached to a surface. These cell types can be subcultured by simply taking a small volume of the parent culture and diluting it in fresh growth medium. Cell density in these cultures is normally measured in cells per millilitre for large eukaryotic cells, or as optical density for 600 nm light for smaller cells like bacteria. The cells will often have a preferred range of densities for optimal growth and subculture will normally try to keep the cells in this range.Adherent cells
Adherent cells, for example many mammalian cell lines, grow attached to a surface such as the bottom of the culture flask. These cell types have to be detached from the surface before they can be subcultured. For adherent cells cell density is normally measured in terms of confluencyConfluency
In cell culture biology, confluency is the term commonly used as a measure of the number of the cells in a cell culture dish or a flask, and refers to the coverage of the dish or the flask by the cells...
, the percentage of the growth surface covered by cells. The cells will often have a preferred range of confluencies for optimal growth, for example a mammalian cell line like HeLa
HeLa
A HeLa cell is a cell type in an immortal cell line used in scientific research. It is the oldest and most commonly used human cell line. The line was derived from cervical cancer cells taken on February 8, 1951 from Henrietta Lacks, a patient who eventually died of her cancer on October 4, 1951...
or Raw 264.7 generally prefer confluencies over 10% but under 100%, and subculture will normally try to keep the cells in this range. For subculture cells may be detached by one of several methods including trypsin
Trypsin
Trypsin is a serine protease found in the digestive system of many vertebrates, where it hydrolyses proteins. Trypsin is produced in the pancreas as the inactive proenzyme trypsinogen. Trypsin cleaves peptide chains mainly at the carboxyl side of the amino acids lysine or arginine, except when...
treatment to break down the proteins responsible for surface adherence, chelating sodium ions with EDTA
EDTA
Ethylenediaminetetraacetic acid, widely abbreviated as EDTA , is a polyamino carboxylic acid and a colourless, water-soluble solid. Its conjugate base is named ethylenediaminetetraacetate. It is widely used to dissolve limescale. Its usefulness arises because of its role as a hexadentate ligand...
which disrupts some protein adherence mechanisms, or mechanical methods like repeated washing or use of a cell scraper. The detached cells are then resuspended in fresh growth medium and allowed to settle back onto their growth surface.