Quiz 2 Slides (Lecs. 5-7) Flashcards
Blast cells
immature, undifferentiated cells that become less active and mature into cytes
Cyte cells
mature, differentiated cells that can revert back into blasts
What are the components of cell culture media? (9)
Water, Balanced Salts, energy molecules, buffering agents, pH indicators, anti bacterial substances, amino acids, vitamins, growth factors
What is the function of balanced salts in cell culture media?
provide a physiologic ionic environment and maintain osmotic balance
What is the function of buffering agents in cell culture media?
maintain pH of cells around 7.4
What is the function of phenol red?
pH Change indicator
What is the purpose of a biosafety cabinet during cell culture?
prevent airborne particles from contaminating cells
What is the purpose of a culture flask?
maintain a sterile environment and give cells a substrate to bind to
Why use an inverted microscope to view cells?
can look directly at cells instead of looking at humidity and other molecules on the top of the glass slide
What are the 4 common cell culture contaminants?
bacteria, yeast, fungi (mold), viruses/mycoplasma
What is the bicarbonate buffer system?
acid-base mechanism involving the balance of carbonic acid, bicarbonate, and CO2 in order to maintain pH
What is the most common cell detachment solution?
trypsin; an enzyme which breaks down proteins and dissociates adherent cells
What is trypan blue?
dye used to differentiated between live and dead cells
What is cell proliferation?
process that results in an increase in cell number
What are the pros of in vivo cell culture?
native 3D environment and all relevant signals present
What are the cons of in vivo cell culture?
many variables leading to noisy data and animal rights concerns
What are the pros of in vitro cell culture?
simplified models, observe as function of time, study parameters independently
What are the cons of in vitro cell culture?
unnatural 2D enviroment, and may lack important signals
What is an advantage of 3D in vitro cell culture?
more accurate representation of in vivo cell growth. and more in vivo like w/o animal rights concerns
What factors affect cell proliferation?
temperature, nutrients, waste removal, cell type, culture type and in vitro vs in vivo
primary cultures
cells isolated directly from living organism, will eventually die off
Cell lines
derived from primary cells, shows exponential growth
Why is the constrained growth model a better approximation of proliferation?
takes into account things that constrain cell growth such as contact inhibition
Stem cells
undifferentiated cells with self renewal capabilities
reprogrammed cells
differentiated cells that have been altered to become less differentiated
What is passaging?
removal of the medium and transfer of cells from a previous culture into new culture with fresh medium
What is expansion during passaging?
moving cells from one dense flask into a larger flask or multiple flasks
When are cells passaged in an adherent culture?
when the cells cover all the available substrate and have no more room to grow
When are cells passaged in a suspension culture?
when the cells exceed the nutritional capacity of the medium
What is the term P1 cell culture
once cells have been re-plated they are termed starting with P1 then P2
what is senescene?
when cells reach their lifetime limit
What is Hayflick limit?
the number of doublings that a cell can undergo (typ. 50-70)
What are the 3 common cell harvesting methods?
bone marrow aspirate, uroepithelial flush out, tissue biopsies
Bone marrow aspirate
use large needle w/ local anesthesia to withdraw about 2mL of marrow
Uroepithelial cell flush out
no anesthesia required, repeated filling/emptying w/ isotonic salt solution
Tissue biopsies
take punch from skin, arthroscopy to harvest, or fluoroscopy and catheter
Mechanical disruption
vortex w/ digestion buffer, pipet vigorously, dice w/ a scalpel
Enzymatic disruption
disrupt cell-ecm or cell-cell connections
Selective adhesion
isolated desired cell types by manipulating adhesion characteristics based on expression of cell surface receptors or adhesion speed
Cell selection techniques
selective adhesion, differential centrifugation, gradient centrifugation, antibody driven, FACS
Differential centrifugation
separation of cells entirely based on size and mass
isopycnic gradient centrifugation
for separation as a function of density only, high density gradient
rate zonal gradient centrifugation
for separation of particles primarily based on size and shape
antibody driven cell selection
select a target antigen on cell surface
fluorescent activated cell sorting
label cells w/ fluorescent antibody and flow through machine to sort
adherent cells
anchorage dependent and must be cultured while attached to a solid or semi solid substrate
suspension cells
anchorage independent, can be grown floating in the medium
Embryonic stem cells
cultured of isolated inner cell mass and/or primordial germ cells
embryonic stem cell limitations
propensity to form teratomas
Somatic cell nuclear transfer
take normal oocyte and remove nucleus then replace with nucleus from a somatic cell
totipotent stem cells
give rise to any of the 220 cell types found in an embryo as well as the placenta
pluripotent stem cells
give rise to all cell types but not the placental cells
multipotent stem cells
develop into a limited number of cell types in a particular lineage
Induced pluripotent stem cells
artificially created stem cells that are forced to express certain genes
progenitor cells
tendency to differentiate into a specific type of cell but is more specific than a stem cell
what is stemness
refers to an essential characteristic of a stem cell that distinguishes it from ordinary cells
chimera
single organism composed of cells from different zygotes
pros of using stem cells in tissue engineering
stem cells typically reduce and/or eliminate tissue rejection or failure
cons of using stem cells in tissue engineering
they differentiate too quickly and they can be tumorigenic
reprogramming cells
goal is to induce differentiated cell to revert back to pluripotency
transdifferentiation
cells regress to a point where they can switch lineages allowing them to differentiate into another cell type
dedifferentiation
terminally differentiated cell reverts back into a less differentiated stage
What factors influence differentiation?
biochemical reagents, changing culture media
stem cell niches
the microenvironment; specific anatomic location which regulates how they participate in tissue generation/maintenance
Cell function assays
adhesion, metabolism, migration, proliferation
transwell membrane assay
used to assess cell migration towards a chemotatic stimulus;
scratch assay
artificial gap is created on cell monolayer and cells move towards it
Laminar flow
no mixing between layers; Re < 2100
turbulent flow
irregular movement of particles; Re > 2400
Alamar blue reaction
used to assess cell metabolism; add alamar blue, incubate, and measure absorbance
Calcein assay
measurement of cell viability; cells are incubated in solution and fluorescent cells are imaged
