The idea of cell cultures dates back to the end of the nineteenth century. It was not a practical laboratory technique until the development of antibiotics. Cell cultures have replaced embryonated eggs as the preferred type of growth medium for many viruses. Cell culture consists of cells grown in culture media in the laboratory. These cultures can be propagated and handled like bacterial cultures; they are more convenient to work with than whole animals or embryonated eggs.
Major problem with cell cultures:
- The process requires trained technicians with experience in working on a full time basis.
- State health laboratories and hospital laboratories do not isolate and identify viruses in clinical work.
- Tissue or serum for analysis is sent to central laboratories to identify virus
There are three types of tissue culture; organ culture, explant culture and cell culture.
Organ cultures are mainly done for highly specialized parasites of certain organs e.g. tracheal ring culture is done for isolation of coronavirus.
Explant culture is rarely done.
Cell culture is mostly used for identification and cultivation of viruses.
Cell culture is the process by which cells are grown under controlled conditions.
Cells are grown in vitro on glass or a treated plastic surface in a suitable growth medium.
At first growth medium, usually balanced salt solution containing 13 amino acids, sugar, proteins, salts, calf serum, buffer, antibiotics and phenol red are taken and the host tissue or cell is inoculated.
On incubation the cell divide and spread out on the glass surface to form a confluent monolayer.
Types of cell culture
1. Primary cell culture:
These are normal cells derived from animal or human cells.
They are able to grow only for limited time and cannot be maintained in serial culture.
They are used for the primary isolation of viruses and production of vaccine.
Examples: Monkey kidney cell culture, Human amnion cell culture
2. Diploid cell culture (Semi-continuous cell lines):
They are diploid and contain the same number of chromosomes as the parent cells.
They can be sub-cultured up to 50 times by serial transfer following senescence and the cell strain is lost.
They are used for the isolation of some fastidious viruses and production of viral vaccines.
Examples: Human embryonic lung strain, Rhesus embryo cell strain
3. Heteroploid cultures (Continuous cell lines):
They are derived from cancer cells.
They can be serially cultured indefinitely so named as continuous cell lines
They can be maintained either by serial subculture or by storing in deep freeze at -70°c.
Due to derivation from cancer cells they are not useful for vaccine production.
Examples: HeLa (Human Carcinoma of cervix cell line), HEP-2 (Humman Epithelioma of larynx cell line), Vero (Vervet monkey) kidney cell lines, BHK-21 (Baby Hamster Kidney cell line).
Susceptible Cell Lines
Herpes Simplex Vero Hep-2, human diploid (HEK and HEL),human amnion
VZV human diploid (HEL, HEK)
CMV human diploid fibroblasts
Adenovirus Hep2, HEK,
Poliovirus MK, BGM, LLC-MK2, human diploid, Vero, Hep-2,Rhadomyosarcoma
Coxsackie B MK, BGM, LLC-MK2, vero, hep-2
Echo MK, BGM, LLC-MK2, human diploid, Rd
Influenza A MK, LLC-MK2, MDCK
Influenza B MK, LLC-MK2, MDCK
Parainfluenza MK, LLC-MK2
Mumps MK, LLC-MK2, HEK, Vero
RSV Hep-2, Vero
Rhinovirus human diploid (HEK, HEL)
Measles MK, HEK
Rubella Vero, RK13
Advantages of cell culture
Relative ease, broad spectrum, cheaper and sensitivity
Disadvantage of cell culture
The process requires trained technicians with experience in working on a full time basis.
State health laboratories and hospital laboratories do not isolate and identify viruses in clinical work.
Tissue or serum for analysis is sent to central laboratories to identify virus.
Cultivation of plant viruses and bacteriophages
Cultivation of plant viruses
There are some methods of Cultivation of plant viruses such as plant tissue cultures, cultures of separated cells, or cultures of protoplasts, etc. viruses can be grown in whole plants.
Leaves are mechanically inoculated by rubbing with a mixture of viruses and an abrasive. When the cell wall is broken by the abrasive, the viruses directly contact the plasma membrane and infect the exposed host cells. A localized necrotic lesion often develops due to the rapid death of cells in the infected area. Some plant viruses can be transmitted only if a diseased part is grafted onto a healthy plant.
Cultivation of bacteriophages
Bacteriophages are cultivated in either broth or agar cultures of young, actively growing bacterial cells.