1. Introduction
Chapter 1

2. What is tissue culture? Tissue culture = organ culture + cell culture
Organ culture – three dimensional culture of undisaggregated tissue retaining some or all histological features of tissue in vivo.
Cell culture – derived from dispersed cells taken from original tissue by enzymatic, mechanical or chemical disaggregation.

3. Historical background (table 1.1)
Devised in 1907 – by Harrison - undisaggregation of cells
Disaggregation of cells and subsequent plating – Rous and Jones, 1916
L929 – first cloned cell strain – Sanford et al., 1948
HeLa – first continuous human cell line – Gey et al., 1952
Introduction of antibodies and different media for culture.

4. Sources of tissues Frog tissue Embryonated hen’s egg/chicken embryo
Rodent tissue
Human cells/tissues – HeLa cells
Insect cell lines-Baculovirus

5. Various tissue culture applications

6. Advantages of Tissue Culture
Consistency and reproducibility of results
Reduced statistical analysis of variance
Control physico-chemical environment
Control of physiological conditions
Change of selective media
Cryopreservation

7. Advantages of Tissue Culture
Characterization by immunostaining
Control of contamination
Reduced volumes of reagents
Control of dose, concentration and time
Microtitration and robotics
Less moral and ethical issues

8. Limitations of tissue culture
Chemical and bacterial contamination
Stringent aseptic procedures
Disposal of wastes
Quantity and cost – medium, pipets etc
Genetic instability
Phenotypic instability - Dedifferentiation

9. Limitations of tissue culture
Identification of cell type is difficult
Markers are not always expressed
Histology is difficult to recreate

10. In Vitro versus In Vivo cultures
Three-dimensional geometry is lost
Cultured on substrates
Specific cell interactions are lost
Cells spread out – proliferate
Lack systemic components involved in regulation of homeostasis
Energy metabolism - Glycolysis

11. Types of Tissue Culture (Organ)
Embryonic organs, adult tissue fragments
High effort
Characterization is easy
Histology is informative
Biochemical differentiation

12. Organ culture No propagation High intersample variation
Quantitation is difficult

13. Explant culture Tissue fragments Moderate effort Cytology and markers
Histology is difficult
Biochemical differentiation is heterogeneous

14. Explant culture Propagation possible from outgrowth
High intersample variation
Quantitation is difficult

15. Cell Culture Disaggregated tissue or primary cell line Less effort
Biochemical, molecular, immunological and cytological assays
No histology
Biochemical differentiation is lost

16. Cell Culture Propagation is possible Low intersample variation
High studies of quantitation

17. Propagation - Monolayer
Monolayer – Cells will attach to substrate
Anchorage dependence – serum derived glycoproteins, conditioning factors secreted by cells, and cell surface glycoproteins

18. Propagation - Suspension culture
Survive and proliferate without attachment
Anchorage independent
Hematopoietic cells, transformed cell lines and cells from malignant tumors

19. Cell Culture

20. Histotypic culture/Histoculture
Tissue culture of one cell type
High density

21. Organotypic culture Restricted to organ culture
More than one cell interaction

22. Summary Historical background Advantages and limitations
Types of tissue culture
Propagation of cell lines
Histotypic and Organotypic culture
Examination questions

23. Grant awarded program
This project is funded by a grant awarded under the President’s Community Based Job Training Grant as implemented by the U.S. Department of Labor’s Employment and Training Administration (CB ). NCC is an equal opportunity employer and does not discriminate on the following basis:
against any individual in the United States, on the basis of race, color, religion, sex, national origin, age disability, political affiliation or belief; and
against any beneficiary of programs financially assisted under Title I of the Workforce Investment Act of 1998 (WIA), on the basis of the beneficiary’s citizenship/status as a lawfully admitted immigrant authorized to work in the United States, or his or her participation in any WIA Title I-financially assisted program or activity.

24. Disclaimer
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