Basic knonwledge on cell cultures

Question 1

The extracellular environment depends on:

Answer 1

1- Nature of substrate (solid(monolayer), liquid(in suspension))
2- Culture medium
3- Gaseous phase and Temperature
4- Adhesion factors

Question 2

1- Choice the Nature of substrate (solid(monolayer), liquid(in suspension)) depends on:

Answer 2

• Cellular yied
• Growth modality (adh, susp)
• Type of atmsph in recipient
• Type of analysis to be performed
• Costs

Question 3

2- Culture medium

Answer 3

• Maintenance of suitable chemico-physical charact (pH& osmolarity)
• Supply of nutrients: sub able to enter the cells & used as COFACTORS/ substrate:
  
  • Essential nutrients:
    BF: AA,VIT, Iones
    Antibiotics
    Serum
    Glutamine

Question 4

Essential components of culture medium

Answer 4

• Inorg salts(K+,Na+,Ca2+): maintain electric potential of memb+ medium osmolarity
• Carbohydrates( glucose: source of carbon ions in cell metab)
• AA(13 here are essential but media contain 21 to avoid eccessive use of metab energy for cell)
• B grp vit
• Water (pyrogen free & tridistilled water(powder)
• Animal serum (bovine = contains growth factors/ components that cannot be substituted with synthetic compounds)

Question 5

Components of Serum!

Answer 5

• Growth factors:
  
  • Cel line-specific (diff stage)
  • Cell-Aspecific for many cell lines
• Hormones
• Drug Transporters

Question 6

How to isolate cells from a tissue

Answer 6

• Explant
• Enzymatic digestion
• Treatment with Ca2+ binder
• Mechanically
• All of those together

Question 7

Requirement to culture cells in vitro

Answer 7

• Possess intrinsic proliferative capacity
• Not be senescent/ close to it
• Microbio unharmed
  => Embryonic/foetal, adult with staminal fraction and adult with proliferative capicity despite the complete diff status : Are the cellular types that can be isolated from a donor

Question 8

From what kind of tissues, cell cultured in vitro can derive from ?

Answer 8

1- Normal tissues:
* Studies of “Cell biology, biochemistry and physiology (strct and funct genral and aspecific properties)
* Evaluation of the “Response to envnmtal changes” that can lead to permanent/transient variation of their status ( Models of Lesion)
- Genetic Manipulation (Engineering)
2- Pathological tissues:
Charac: evaluation of “Structure, functional, general and aspecific prop” compared to cells belonging to the same TISSUE but HEALTHY
=> Study of the causes / Biomolecular mechanisms at the basis of pathological phenotype
=> Correct (weaken) the anomalies at the basis of a definite pathological phenotype (THERAPY)

Question 9

TISSUE / ORGAN CULTURE

Answer 9

• Maintains cell-cell and cell-matrix Interactions
• Maintains for a Longer period of time, the biochemical and hystological Differentiation
• After the explant and an initial necrosis, it Remains in a NOT Replicative condition for certain period of time
• Cannot be Propagated
• High Variability among Different experiments (with consequent limitations in execution of replicas)

Question 10

Cell Culture characteristics

Answer 10

• Absent organized structure
• Loss of Hystiotypic Architecture
• PArtial loss of Specific biochemical ACTIvities
• Good PRopagation
• Accuracy among exp
• Cryoperservation
• Purification in selective media
• Separation in specific subpopulations
• Clonability

Question 11

Evolution of cells in Culture

Answer 11

1- Ph1: Iary culture: cells with a slow duplication rate
2- Ph2: Cell line: constant and rapid duplication time
3- Ph3: Senescent cells: Extension of duplication time, cell Death