culturing cells

Question 1

what did robert hooke do

Answer 1

he discovered and named cells

Question 2

what did antonie van leeuwenkoek do

Answer 2

he was the first person to observe and describe living cells

Question 3

who came up with two of the three tenets that make up cell theory

Answer 3

mattias schleiden and theodar schwann

Question 4

what two tenets did schleiden and schwann come up with

Answer 4

living organisms are made up of one or more cells, cell is the basic unit of life

Question 5

what did rudolph virchow do

Answer 5

showed that all cells come from pre-existing cells

Question 6

define culturing

Answer 6

refers to the process of growing cells in an environment that promotes their survival

Question 7

why are cells cultured

Answer 7

to study the properties of them + we use cells to produce lots of products (ie antibodies)

Question 8

what is 2D culturing

Answer 8

cells cultured on a flat surface

Question 9

what is 3D culturing

Answer 9

makes use of a hydrogel, where cells are growing on a 3D scaffold

Question 10

what two types of cell cultures are there

Answer 10

primary cell cultures + cell lines

Question 11

what are primary cell cultures prepared from

Answer 11

animal tissues (ie skin, kidney, liver)

Question 12

how are primary cell cultures prepared

Answer 12

individual cells are prepared by treatment with proteolytic enzymes and then put into the appropriate medium (trypsin+collegenase are used to break the cells away from each other)

Question 13

T or F: primary cell cultures are the most similar to cells found in an intact organism

Answer 13

true

Question 14

T or F: cells in a primary cell culture will keep dividing

Answer 14

false; they will stop after a certain point

Question 15

after how many divisions in a primary cell culture will cells stop dividing

Answer 15

30-50

Question 16

define cell senescence

Answer 16

when cells stop dividing

Question 17

why do cells in a culture stop dividing after 30-50 divisions

Answer 17

production of telomerase turns off, and this is an enzyme that allows cell division to keep happening by keeping the chromosomes at a certain length. without telomerase, chromosomes will get shorter and shorter

Question 18

define culture shock

Answer 18

a protective mechanism that cells have in place when cultured for too long – it makes them stop dividing

Question 19

define cell lines

Answer 19

cell types that will grow indefinitely in culture

Question 20

why do cell lines grow indefinitely

Answer 20

they don’t turn off telomerase production

Question 21

give an example of cell types that grow indefinitely

Answer 21

rodent cell types

Question 22

why do some rodent cell types grow indefinitely

Answer 22

they have a mutation that prevents culture shock from occuring

Question 23

how can we generate cell lines from other organisms (other than rodents)

Answer 23

by using cells from within a tumor

Question 24

define transformed cell line

Answer 24

cells from tumors

Question 25

describe characteristics of transformed cell lines (ie cell lines from the cells of a tumor)

Answer 25

transformed cell lines only slightly resemble the parent cell/tissue, they may have altered gene expression, and may have an abnormal karyotype. They can often grow without attaching to a surface, and they can grow to a high density

Question 26

what is the first human cell line

Answer 26

HeLa cells

Question 27

where did HeLa cells come from

Answer 27

henrietta lacks, who died of cervical cancer. Her cells were removed and distributed without her permission

Question 28

what medical discoveries did HeLa cells help with

Answer 28

used to test the polio vaccine, used to show HPV causes cancer, and used in testing vaccines for covid

Question 29

define serum

Answer 29

the liquid portion of blood (ie the plasma) where the cells have been removed and the proteins involved in clotting have been removed

Question 30

why is serum used to grow a culture

Answer 30

it's very nutritious; contains amino acids, proteins, vitamins, CHOs, lipids, hormones, growth factors, minerals

Question 31

what is the optimal pH range for mammalian cells

Answer 31

7.2-7.4

Question 32

define osmolality + osmole

Answer 32

osmolality is the number of osmoles of a solute in a kg of solvent. An osmole is the amount of moles of solute that contribute to the osmotic pressure of a solution

Question 33

what system is used to maintain the pH level

Answer 33

CO2/sodium bicarbonate buffering system

Question 34

describe the CO2/sodium bicarbonate buffering system in media

Answer 34

as cells grow and respire, CO2 is produced. this dissolved and combined with water to form carbonic acid. This forms H+, causing the pH to decrease. Adding sodium bicarbonate to the medium minimizes the pH drop

Question 35

how much sodium bicarbonate do we add to a media to maintain pH (and how much CO2 is there)

Answer 35

1.5g/L of sodium bicarbonate with 5% CO2

Question 36

why is phenol red incorporated into medium

Answer 36

it serves as a pH indicator

Question 37

explain how phenol red serves as a pH indicator

Answer 37

acidic=yellow, basic=red

Question 38

T or F: cell cultures are grown in an open system

Answer 38

true

Question 39

define an open system for growing cultures

Answer 39

flask is open to the environment so has exchange can occur, but we use a vented cap that small enough that microbes cannot get in

Question 40

how small are the vents on a vented cap of an open system (what is the purpose of this)

Answer 40

0.2um, small enough that microbes cannot get in

Question 41

what are adherent cells

Answer 41

anchorage-dependent

Question 42

define confluency

Answer 42

the percentage of the surface of the culture dish that is covered by adherent cells

Question 43

at what percentage of confluency do we subculture adherent cells

Answer 43

75-90

Question 44

what do we sometimes treat the surface of the culture dish with to benefit the adherent cells

Answer 44

treated with something they can adhere to: collagen, laminin, gelatin, fibronectin

Question 45

describe how we subculture adherent cells

Answer 45

treat them with protease to lift the cells off the substrate and away from each other (trypsin/EDTA is frequently used)

Question 46

why is EDTA commonly used when subculturing adherent cells

Answer 46

because EDTA binds to Ca2+ which is involved with cell adhesion, so this makes it easier to remove the cells

Question 47

what are suspension cells

Answer 47

anchorage-independent

Question 48

T or F: suspension cells are easier to subculture than adherent cells

Answer 48

true; because we don't need to use trypsin

Question 49

how do we visualize suspension cells

Answer 49

bright field microscope or phase-contrast microscope

Question 50

what two orientations will a microscope have

Answer 50

upright or inverted

Question 51

benefit to inverted microscope?

Answer 51

objective lenses are below the specimen, so we can look at larger objects and it's easier/faster to focus (and no risk to the objective lenses)

Question 52

which microscope is inverted

Answer 52

phase-contrast

Question 53

which microscope is upright

Answer 53

bright-field

Question 54

which cell line are we using in the lab

Answer 54

L929

Question 55

where do L929 cells come from (what organism)

Answer 55

mice

Question 56

how old are the L929 cells that we're using

Answer 56

100 days

Question 57

what is the morphology of the L929 cells

Answer 57

fibroblasts

Question 58

are l929 cells adherent or suspension cells

Answer 58

adherent