What are the 4 groups of Eukaryotic Cells?
What is the cell theory?
- Smallest unit of life
- All organisms are made of cells
- New cells come from pre-existing cells
What is cell culture?
The process by which prokaryotic or eukaryotic cells are grown under controlled conditions
A homogenous population of cells derived from a single parental cell is called a ______?
What famous cell was derived from cancerous growth of Henreitta Lacks?
Current Cell culture techniques are used for?
- Biomedical technology
- Molecular Cell Biology
- Tissue Engineering
What are some future prospects for cell culture?
- Targeting diseases
What is a Primary Culture?
Cells taken directly from a tissue to a dish
What is a secondary culture?
- Cells taken from a primary and passed or divided in vitro
- Limited divisions (ends in apoptosis)
What is a cell line?
Cells that have mutated and won't undergo apoptosis
What is a transformed cell line?
A cell line that has been transformed by a tumor inducing virus or chemical
What is a hybrid cell line (hybridoma)?
Two cell types fused together with characteristics of each
What other disciplines is cell culture related with?
- Cell biology
- Genetic Engineering
- Protein Chemistry
What are the advantages of using cell culture?
- Use of animals reduced
- Cells from one cell line are homogenous and have same growth requirements
- In-vitro manipulated is easier that in vivo
What are the disadvantages of using cell culture?
- Cell characteristics may change over time (genotype and phenotype change)
- Cells adapt to different culture environments
- Nearly impossible to recreate the in vivo environment in cell culture
What is the purpose of designing and designating lab a working area?
- To avoid contamination
- Protect personnel
What are the two cell culture labs spaces?
- Sterile handling section (working, incubation, storage)
- Support service area (washing, preparing, sterilization)
What are the three classes of microbiological safety cabinets?
- I - Open fronted, low and moderate risk agents where product protection is not critical
- II - open fronted, protects from particulate contamination, low to moderate risk
- III - totally enclosed, gas tight, glove ports. High risk pathogens
What does an incubator do?
- Provides correct growth conditions
What is the most common temperature for ultra low freezers and what are they used for?
- Long term storage of tissue cells, DNA, RNA, and some growth medium
- (sometimes -80 for cell culture labs)
What does an autoclave do?
Sterilizes by killing microbes with superheated steam
What does a centrifuge do?
Seperate samples on density gradient
What does a spectrophotometer do?
Measures nucleic acids and protein concentration
What is the difference between a serological pipette and a micropipette?
- Serological - mL
- Micro - uL
What do animal and plants require in their media?
hormones and growth factors (occurs in-vivo) plus essential nutrients
What does developing cell culture media in animals depend on?
- Origin and type of cell/tissue
- Purpose of in-vitro culturing
What are the sources of cell culture media?
- In-house - ready made in the lab and stored as stock
- Commercial- liquid or solid and may need addition growth factors or antibiotics
What is the purpose of developing in-vitro growth media?
- To grow cells in conditions that mimics in vivo
- nutrients for the cell
- pH and aeration
- salt concentration
What are the general types of media?
- Serum based
- Serum free
- Animal free
- Protein free or chemically defined
What two components are in a complete culture medium?
- Basal medium (nutrients, salts and buffers)
- Supplements (hormone, growth factors, antibiotics)
What is balance salt solutions (BSS) required in basal media?
maintain pH and prevent osmotic imbalance
What is the primer nutrient in mammalian cell culture?
Glucose for energy
Why should essential amino acids be supplemented to cells grown in-vitro?
The cell doesn't produce it and it needed for growth
What are vitamins?
nutrients required for essential metabolic reactions (growth and multiplication)
What does the vitamin content of basal medium depend on?
The cell line for which it was designed
Why are cells cultured in a CO2 incubator?
Growing cells don't produce enough CO2 to maintain optimal pH
What is Hepes?
An organic buffer used to prevent pH change when culture is removed from CO2 incubator
What are the disadvantages of using Hepes?
- Toxic to some cells at elevated levels
- Higher cost
- No nutritional value
What is the most common pH indicator and what colors does is turn for acidic, optimal, and basic?
- Optimal - Red
- Acidic- Yellow
- Basic- Purple
What are the 5 types of growth factors
- Colony Stimulating Factors
Why are antibiotics often used in cell culture media?
Prevent or reduce microorganism contamination
Why aren't antibiotics recommended for routine sub-culturing and stock culture?
May mask low level of contamination
What are the most frequently used antibiotics?
- Mixtures of penicillin and streptomycin
Why shouldn't antibiotics be used continuously?
They may develop resistant microorganisms
What is a serum?
- Yellowish, clear liquid left over after fibrin and cells are removed from blood
- Or it is a supernatant of clotted blood
What is a serums function?
- Promotes attachment and spreading
- Supplies micronutrients and growth factor
- protects and acts as antitoxins, antioxidants, antiproteases
What are the sources of serum?
Fetal calf, calf, horse, and bovine
What is serum stored at and how long is it good at this temperature?
-20 and good for at least 5 years
What are the disadvantages of serum?
- High variability between batches
- Risk of contamination
What are the alternatives to the use of serum?
- Serum free media
- Synthetic serum