Introduction to Animal Cell Culture

Technical Bulletin
John A. Ryan, Ph.D.
Corning Incorporated
Life Sciences
900 Chelmsford St.
Loell, !A 0"#$"
Table of Contents
Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1
What is Cell and Tissue Culture? . . . . . . . . . . . . . . . . . . 1
How are Cell Cultures Obtained? . . . . . . . . . . . . . . . . . . 2
What Are Cultured Cells Lie? . . . . . . . . . . . . . . . . . . . . !
What Are "ome of the #roblems $aced b%
Cultured Cells? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . &
How to 'ecide if Cultured Cells Are (Ha))%*? . . . . . . . +
What is Cell Culture ,sed $or? . . . . . . . . . . . . . . . . . . . +
-eferences . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Introduction
Cell culture has become one of the ma/or tools used in the
life sciences toda%. This 0uide is desi0ned to ser1e as a basic
introduction to animal cell culture. It is a))ro)riate for lab2
orator% worers who are usin0 it for the first time3 as well as
for those who interact with cell culture researchers and who
want a better understandin0 of the e% conce)ts and termi2
nolo0% in this interestin0 and ra)idl% 0rowin0 field.
What is Cell and Tissue Culture?
Tissue Culture is the 0eneral term for the remo1al of cells3
tissues3 or or0ans from an animal or )lant and their subse2
4uent )lacement into an artificial en1ironment conduci1e
to 0rowth. This en1ironment usuall% consists of a suitable
0lass or )lastic culture 1essel containin0 a li4uid or semi2
solid medium that su))lies the nutrients essential for sur2
1i1al and 0rowth. The culture of whole or0ans or intact
or0an fra0ments with the intent of stud%in0 their continued
function or de1elo)ment is called Organ Culture. When
the cells are remo1ed from the or0an fra0ments )rior to3
or durin0 culti1ation3 thus disru)tin0 their normal relation2
shi)s with nei0hborin0 cells3 it is called Cell Culture.
Althou0h animal cell culture was first successfull% underta2
en b% -oss Harrison in 15673 it was not until the late 15&68s
to earl% 15968s that se1eral de1elo)ments occurred that made
cell culture widel% a1ailable as a tool for scientists. $irst3
there was the de1elo)ment of antibiotics that made it easier
to a1oid man% of the contamination )roblems that )la0ued
earlier cell culture attem)ts. "econd was the de1elo)ment of
2
Additional cell culture
terminolo0% and usa0e
information can be found
on the "ociet% for In :itro
Biolo0% web site at
ww w .si1 b .o r0 ;ed u<
terminolo0%.as).
the techni4ues3 such as the use of tr%)sin to remo1e cells from culture 1essels3 necessar% to
obtain continuousl% 0rowin0 cell lines =such as HeLa cells>. Third3 usin0 these cell lines3
scientists were able to de1elo) standardi?ed3 chemicall% defined culture media that made
it far easier to 0row cells. These three areas combined to allow man% more scientists to use
cell3 tissue and or0an culture in their research.
'urin0 the 15+68s and 15768s3 commerciali?ation of this technolo0% had further im)act on
cell culture that continues to this da%. Com)anies3 such as Cornin03 be0an to de1elo) and
sell dis)osable )lastic and 0lass cell culture )roducts3 im)ro1ed filtration )roducts and mate2
rials3 li4uid and )owdered tissue culture media3 and laminar flow hoods. The o1erall result
of these and other continuin0 technolo0ical de1elo)ments has been a wides)read increase in
the number of laboratories and industries usin0 cell culture toda%.
$i@ed and stained human
foresin e@)lants on the sur2
face of a 196 mm culture dish.
The e@)lants were cultured for
a))ro@imatel% two wees. Two
of the nine e@)lants =bottom
left and ri0ht corners> failed to
0row. The remainin0 e@)lants
show 0ood 0rowth. Aach
s4uare is a))ro@imatel% 2 cm
across.
How Are Cell Cultures Obtained?
Primary C%lt%re
When cells are sur0icall% remo1ed from an
or0anism and )laced into a suitable culture
en1ironment3 the% will attach3 di1ide and
0row. This is called a Primary Culture.
There are two basic methods for doin0 this.
$irst3 for Explant Cultures3 small )ieces of
tissue are attached to a 0lass or treated )lastic
culture 1essel and bathed in culture medium.
After a few da%s3 indi1idual cells will mo1e
from the tissue e@)lant out onto the culture
1essel surface or substrate where the% will
be0in to di1ide and 0row. The second3 more
widel% used method3 s)eeds u) this )rocess b%
addin0 di0estin0 =)roteol%tic> en?%mes3 such
as tr%)sin or colla0enase3 to the tissue fra02
ments to dissol1e the cement holdin0 the cells
to0ether. This creates a sus)ension of sin0le
cells that are then )laced into culture 1essels
containin0 culture medium and allowed to
0row and di1ide. This method is called
Enzymatic Dissociation.
S%&c%lt%ring
-emo1em
tissue
Bince orm
cho)
'i0est withm
)roteol%ticm
en?%mes
#lace in m
culture
An?%matic 'issociation
#rimar% culture from the fish
Poeciliopsis l%cida. Ambr%os
were minced and dissociated
with a tr%)sin solution. These
cells were in culture for about
1 wee and ha1e formed a
confluent monola%er.
When the cells in the )rimar% culture 1essel ha1e 0rown and filled u) all of the a1ailable
culture substrate3 the% must be Subcultured to 0i1e them room for continued 0rowth. This
is usuall% done b% remo1in0 them as 0entl% as )ossible from the substrate with en?%mes.
These are similar to the en?%mes used in obtainin0 the )rimar% culture and are used to brea
the )rotein bonds attachin0 the cells to the substrate. "ome cell lines can be har1ested b%
0entl% scra)in0 the cells off the bottom of the culture 1essel. Once released3 the cell sus)en2
sion can then be subdi1ided and )laced into new culture 1essels.
Once a sur)lus of cells is a1ailable3 the% can be treated with suitable cr%o)rotecti1e a0ents3
such as dimeth%lsulfo@ide ='B"O> or 0l%cerol3 carefull% fro?en and then stored at cr%o2
0enic tem)eratures =below 21!6CC> until the% are needed. The theor% and techni4ues for
cr%o)reser1in0 cells are co1ered in the Cornin0 Technical BulletinD Eeneral Euide for
Cr%o0enicall% "torin0 Animal Cell Cultures =-ef. 5>.
'%ying And 'orroing
An alternati1e to establishin0 cultures b% )rimar% culture is to bu% established cell cultures
from or0ani?ations such as the ATCC =ww w .atcc.or0>3 or the Coriell Institute for Bedical
-esearch =ccr.coriell.or0>. These two non)rofit or0ani?ations )ro1ide hi0h 4ualit% cell lines
that are carefull% tested to ensure the authenticit% of the cells.
Cornin0 culture dishes are
a1ailable in a 1ariet% of si?es
and sha)es for 0rowin0
anchora0e2de)endent cells.
Cornin0 culture flass are
used for 0rowin0 anchora0e2
de)endent cells.
Cornin0 s)inner 1essels are
used for 0rowin0 anchora0e2
inde)endent cells in
sus)ension.
$ibroblast2lie !T! cells deri1ed
from mouse embr%os
Bore fre4uentl%3 researchers will obtain =borrow> cell lines from other laboratories. While
this )ractice is wides)read3 it has one ma/or drawbac. There is a hi0h )robabilit% that the
cells obtained in this manner will not be health%3 useful cultures. This is usuall% due to )re2
1ious mi@2u)s or contamination with other cell lines3 or the result of contamination with
microor0anisms such as m%co)lasmas3 bacteria3 fun0i or %east. These )roblems are co1ered
in detail in a Cornin0 Technical BulletinD ,nderstandin0 and Bana0in0 Cell Culture
Contamination =-ef. 7>.
What Are Cultured Cells Lie?
Once in culture3 cells e@hibit a wide ran0e of beha1iors3 characteristics and sha)es. "ome
of the more common ones are described below. Fohn #aul discusses these issues in detail in
Cha)ter ! of Cell and (iss%e C%lt%re =-ef. !>.
Cell C%lt%re Systems
Two basic culture s%stems are used for 0rowin0 cells. These are based )rimaril% u)on the
abilit% of the cells to either 0row attached to a 0lass or treated )lastic substrate =Monolayer
Culture Sytems> or floatin0 free in the culture medium =Suspension Culture Systems>.
Bonola%er cultures are usuall% 0rown in tissue culture treated dishes3 T2flass3 roller bottles3
Cell"TACG
H
Culture Chambers3 or multi)le well )lates3 the choice bein0 based on the
num2 ber of cells needed3 the nature of the culture en1ironment3 cost and )ersonal
)reference.
"us)ension cultures are usuall% 0rown eitherD
1. In ma0neticall% rotated s)inner flass or shaen Arlenme%er flass where the cells are
e)t acti1el% sus)ended in the mediumI
2. In stationar% culture 1essels such as T2flass and bottles where3 althou0h the cells are not
e)t a0itated3 the% are unable to attach firml% to the substrate.
Ban% cell lines3 es)eciall% those deri1ed from normal tissues3 are considered to be
Anchorage-Dependent that is3 the% can onl% 0row when attached to a suitable substrate.
"ome cell lines that are no lon0er considered normal =fre4uentl% desi0nated as Trans!ormed
Cells" are fre4uentl% able to 0row either attached to a substrate or floatin0 free in sus)ensionI
the% are Anchorage-#ndependent$ In addition3 some normal cells3 such as those found in
the blood3 do not normall% attach to substrates and alwa%s 0row in sus)ension.
(ypes of Cells
Cultured cells are usuall% described based on their mor)holo0% =sha)e and a))earance> or
their functional characteristics. There are three basic mor)holo0iesD
1. A)ithelial2lieD cells that are attached to a substrate and a))ear flattened and )ol%0onal
in sha)e.
2. L%m)hoblast2lieD cells that do not attach normall% to a substrate but remain in
sus)ension with a s)herical sha)e.
!. $ibroblast2lieD cells that are attached to a substrate and a))ear elon0ated and bi)olar3
fre4uentl% formin0 swirls in hea1% cultures.
It is im)ortant to remember that the culture conditions )la% an im)ortant role in determin2
in0 sha)e and that man% cell cultures are ca)able of e@hibitin0 multi)le mor)holo0ies.
,sin0 cell fusion techni4ues3 it is also )ossible to obtain h%brid cells b% fusin0 cells from two
different )arents. These ma% e@hibit characteristics of either )arent or both )arents. This
techni4ue was used in 1579 to create cells ca)able of )roducin0 custom tailored mono2
clonal antibodies. These h%brid cells =called %ybridomas> are formed b% fusin0 two differ2
ent but related cells. The first is a s)leen2deri1ed l%m)hoc%te that is ca)able of )roducin0
the desired antibod%. The second is a ra)idl% di1idin0 m%eloma cell =a t%)e of cancer cell>
that has the machiner% for main0 antibodies but is not )ro0rammed to )roduce an% anti2
bod%. The resultin0 h%bridomas can )roduce lar0e 4uantities of the desired antibod%. These
antibodies3 called Monoclonal Antibodies due to their )urit%3 ha1e man% im)ortant clini2
cal3 dia0nostic3 and industrial a))lications with a %earl% 1alue of well o1er a billion dollars.
A)ithelial2lie cell line =Cl25>
deri1ed from rat li1er. The
mitotic cells indicates this
culture is acti1el% 0rowin0.
CHO2G1 cells J a widel% used
continuous =transformed> cell
line deri1ed from adult
Chinese hamster o1ar% tissue
in 1597.
#hotomicro0ra)h of a low
le1el %east infection in a li1er
cell line =#LHC213 ATCC K C-L2
2&6+>. Buddin0 %east cells can
been seen in se1eral areas
=arrows>. At this low le1el of
contamination3 no medium
turbidit% would be seenI
howe1er3 in the absence of
antibiotics3 the culture
medium will )robabl% become
turbid within a da%.
)%nctional Characteristics
The characteristics of cultured cells result from both their ori0in =li1er3 heart3 etc.> and how
well the% ada)t to the culture conditions. Biochemical marers can be used to determine if
cells are still carr%in0 on s)eciali?ed functions that the% )erformed in 1i1o =e.0.3 li1er cells
secretin0 albumin>. Bor)holo0ical or ultrastructural marers can also be e@amined =e.0.3
beatin0 heart cells>. $re4uentl%3 these characteristics are either lost or chan0ed as a result of
bein0 )laced in an artificial en1ironment. "ome cell lines will e1entuall% sto) di1idin0 and
show si0ns of a0in0. These lines are called &inite. Other lines are3 or become immortalI
these can continue to di1ide indefinitel% and are called Continuous cell lines. When a
(normal* finite cell line becomes immortal3 it has under0one a fundamental irre1ersible
chan0e or (transformation*. This can occur s)ontaneousl% or be brou0ht about intentional2
l% usin0 dru0s3 radiation or 1iruses. Trans!ormed Cells are usuall% easier and faster 0row2
in03 ma% often ha1e e@tra or abnormal chromosomes and fre4uentl% can be 0rown in sus2
)ension. Cells that ha1e the normal number of chromosomes are called Diploid cellsI those
that ha1e other than the normal number are Aneuploid. If the cells form tumors when the%
are in/ected into animals3 the% are considered to be 'eoplastically Trans!ormed.
What Are "ome of the #roblems $aced b% Cultured Cells?
A*oiding Contamination
Cell culture contamination is of two main t%)esD chemical and biolo0ical. Chemical contam2
ination is the most difficult to detect since it is caused b% a0ents3 such as endoto@ins3 )lasti2
ci?ers3 metal ions or traces of chemical disinfectants3 that are in1isible. The cell culture
effects associated with endoto@ins are co1ered in detail in the Technical BulletinD Andoto@ins
and Cell Culture =-ef. 16>. Biolo0ical contaminants in the form of fast 0rowin0 %east3 bacte2
ria and fun0i usuall% ha1e 1isible effects on the culture =chan0es in medium turbidit% or )H>
and thus are easier to detect =es)eciall% if antibiotics are omitted from the culture medium>.
Howe1er3 two other forms of biolo0ical contamination3 m%co)lasmas and 1iruses3 are not
eas% to detect 1isuall% and usuall% re4uire s)ecial detection methods.
There are two ma/or re4uirements to a1oidin0 contamination. $irst3 )ro)er trainin0 in and
use of 0ood ase)tic techni4ue on the )art of the cell culturist. "econd3 )ro)erl% desi0ned3
maintained and sterili?ed e4ui)ment3 )lasticware3 0lassware3 and media. The careful and
selecti1e =limited> use of antibiotics desi0ned for use in tissue culture can also hel) a1oid
culture loss due to biolo0ical contamination. These conce)ts are co1ered in detail in a
Cornin0 Technical BulletinD ,nderstandin0 and Bana0in0 Cell Culture Contamination
=-ef. 7>.
)inding A +,appy- .n*ironment
To cell culturists3 a (ha))%* en1ironment is one that does more than /ust allow cells to sur2
1i1e in culture. ,suall%3 it means an en1ironment that3 at the 1er% least3 allows cells to
increase in number b% under0oin0 cell di1ision =mitosis>. A1en better3 when conditions are
/ust ri0ht3 some cultured cells will e@)ress their (ha))iness* with their en1ironment b% car2
r%in0 out im)ortant in 1i1o )h%siolo0ical or biochemical functions3 such as muscle contrac2
tion or the secretion of hormones and en?%mes. To )ro1ide this en1ironment3 it is im)or2
tant to )ro1ide the cells with the a))ro)riate tem)erature3 a 0ood substrate for attachment3
and the )ro)er culture medium. Ban% of the issues and )roblems associated with ee)in0
cells (ha))%* are co1ered in the Cornin0 Technical BulletinD Eeneral Euide for Identif%in0
and Correctin0 Common Cell Culture Erowth and Attachment #roblems =-ef. .>.
Tem)erature is usuall% set at the same )oint as the bod% tem)erature of the host from which
the cells were obtained. With cold2blooded 1ertebrates3 a tem)erature ran0e of 1.C to 29CC
is suitableI most mammalian cells re4uire !+C to !7CC. This tem)erature ran0e is usuall%
maintained b% use of carefull% calibrated3 and fre4uentl% checed3 incubators.
Basic en1ironmental
-e4uirements for
(Ha))%* CellsD
Controlled
tem)erature
Eood substrate
for cell
attachment
A))ro)riate medium
and incubator that
maintains the correct
)H and osmolalit%
Cornin0H Transwell )ermeable
su))orts are used to stud% cell
trans)ort and mi0ration.
"us)ension and microcarrier
cultures can be 0rown in 0lass
and )lastic s)inner 1essels.
CHO2G1 cells 0rowin0 on a
microcarrier bead
Anchora0e2de)endent cells also re4uire a 0ood substrate for attachment and 0rowth. Elass
and s)eciall% treated )lastics =to mae the normall% h%dro)hobic )lastic surface h%dro)hilic
or wettable> are the most commonl% used substrates. Howe1er3 Attachment &actors3 such
as colla0en3 0elatin3 fibronectin and laminin3 can be used as substrate coatin0s to im)ro1e
0rowth and function of normal cells deri1ed from brain3 blood 1essels3 idne%3 li1er3 sin3
etc. Often normal anchora0e2de)endent cells will also function better if the% are 0rown on a
)ermeable or )orous surface. This allows them to )olari?e =ha1e a to) and bottom throu0h
which thin0s can enter and lea1e the cell> as the% do in the bod%. Transwell
H
inserts are
Cornin0 1essels with membrane2based )ermeable su))orts that allow these cells to de1elo)
)olarit% and ac4uire the abilit% to e@hibit s)ecial functions such as trans)ort. Ban% s)ecial2
i?ed cells can onl% be trul% (ha))%* =function normall%> when 0rown on a )orous substrate
in serum2free medium with the a))ro)riate mi@ture of 0rowth and attachment factors.
Cells can also be 0rown in sus)ension on beads made from 0lass3 )lastic3 )ol%acr%lamide and
cross2lined de@tran molecules. This techni4ue has been used to enable anchora0e2de)endent
cells to be 0rown in sus)ension culture s%stems and is increasin0l% im)ortant for the manu2
facture of cell2based biolo0icals.
The culture medium is the most im)ortant and com)le@ factor to control in main0 cells
(ha))%*. Besides meetin0 the basic nutritional re4uirement of the cells3 the culture medium
should also ha1e an% necessar% 0rowth factors3 re0ulate the )H and osmolalit%3 and )ro1ide
essential 0ases =O
2
and CO
2
>. The Lfood8 )ortion of the culture medium consists of amino
acids3 1itamins3 minerals3 and carboh%drates. These allow the cells to build new )roteins and
other com)onents essential for 0rowth and function as well as )ro1idin0 the ener0% neces2
sar% for metabolism. $or additional information on this to)ic3 see the articleD Construction
of Tissue Culture Bedia b% C. Wa%mouth in /roth, 0%trition and !eta&olism of Cells in
C%lt%re3 :olume 13 =1572I -ef. 9>.
The 0rowth factors and hormones hel) re0ulate and control the cells8 0rowth rate and func2
tional characteristics. Instead of bein0 added directl% to the medium3 the% are often added in
an undefined manner b% addin0 9 to 26M of 1arious animal sera to the medium. ,nfortu2
natel%3 the t%)es and concentration of these factors in serum 1ar% considerabl% from batch
to batch. This often results in )roblems controllin0 0rowth and function. When 0rowin0
normal functional cells3 sera are often re)laced b% s)ecific 0rowth factors.
The medium also controls the )H ran0e of the culture and buffers the cells from abru)t
chan0es in )H. ,suall% a CO
2
2bicarbonate based buffer or an or0anic buffer3 such as
HA#A"3 is used to hel) ee) the medium )H in a ran0e from 7.6 to 7.& de)endin0 on the
t%)e of cell bein0 cultured. When usin0 a CO
2
2bicarbonate buffer3 it is necessar% to re0ulate
the amount of CO
2
dissol1ed in the medium. This is usuall% done usin0 an incubator with
CO
2
controls set to )ro1ide an atmos)here with between 2M and 16M CO
2
=for Aarle8s
salts2based buffers>. Howe1er3 some media use a CO
2
2bicarbonate buffer =for Hans8 salts2
based buffers> that re4uires no additional CO
2
3 but it must be used in a sealed 1essel =not
dishes or )lates>. $or additional information on this to)ic3 see the articleD The Easeous
An1ironment of the Cell in Culture b% W.$. BcLimans in /roth, 0%trition and
!eta&olism of Cells in C%lt%re =1572I -ef. 9>.
$inall%3 the osmolalit% =osmotic )ressure> of the culture medium is im)ortant since it hel)s
re0ulate the flow of substances in and out of the cell. It is controlled b% the addition or sub2
traction of salt in the culture medium. A1a)oration of culture media from o)en culture 1es2
sels =dishes3 etc.> will ra)idl% increase the osmolalit% resultin0 in stressed3 dama0ed or dead
cells. $or o)en =not sealed> culture s%stems3 incubators with hi0h humidit% le1els to reduce
e1a)oration are essential. $or additional information3 see article b% C. Wa%mouthD Osmolalit%
of Bammalian Blood and of Bedia for Culture of Bammalian Cells =1576I -ef. +>.
A@aminin0 the mor)holo0% of
these stained roller bottles
=containin0 B-C29 human
fibroblasts> is a 0ood wa% to
chec if the cells are (ha))%*.
The bottle on the left was
rotated at too hi0h a s)eed
resultin0 in )oor attachment
and 0rowth and 1er%
(unha))%* cells.
A colon% of fi@ed and stained
human fibroblast cells
HT" TranswellH22& )lates are
used for to@icit% testin0 and
dru0 trans)ort studies
How to 'ecide if Cultured Cells Are (Ha))%*
A1aluatin0 the 0eneral health or (ha))iness* of a culture is usuall% based on four im)ortant
cell characteristicsD mor)holo0%3 0rowth rate3 )latin0 efficienc% and e@)ression of s)ecial
functions. These same characteristics are also widel% used in e1aluatin0 e@)erimental results.
The Morphology or cell sha)e is the easiest to determine but is often the least useful.
While chan0es in mor)holo0% are fre4uentl% obser1ed in cultures3 it is often difficult to
relate
these obser1ations to the condition that caused them. It is also a 1er% difficult characteristic
to 4uantif% or to measure )recisel%.
Often3 the first si0n that somethin0 is wron0 with a culture occurs when the cells are micro2
sco)icall% e@amined and )oor or unusual )atterns of cell attachment or 0rowth are obser1ed.
When )roblems are sus)ected3 stainin0 the culture 1essels with cr%stal 1iolet or other sim)le
histolo0ical stains ma% show 0rowth )atterns indicatin0 a )roblem. These 0rowth )roblems
are discussed in detail in the Cornin0 Technical BulletinD Eeneral Euide for Identif%in0 and
Correctin0 Common Cell Culture Erowth and Attachment #roblems =-ef. .>.
Cell countin0 and other methods for estimatin0 cell number3 on the other hand3 allow the
determination of the (ro)th *ate3 which is sensiti1e to ma/or chan0es in the culture
en1ironment. This allows the desi0n of e@)eriments to determine which set of conditions
=culture media3 substrate3 serum3 )lasticware> is better
for the cells3 i.e.3 the conditions )roducin0 the best 0rowth rate. These same or similar
techni4ues can also be used to measure cell sur1i1al or death and are often used for in 1itro
c%toto@icit% assa%s.
Plating E!!iciency is a testin0 method where small numbers of cells =26 to 266> are )laced
in a culture 1essel and the number of colonies the% form is measured. The )ercenta0e of
cells formin0 colonies is a measure of sur1i1al3 while the colon% si?e is a measure of 0rowth
rate. This testin0 method is similar in a))lication to 0rowth rate anal%sis but is more sensi2
ti1e to small 1ariations in culture conditions.
The final characteristic3 the Expression o! Specialized &unctions3 is usuall% the most dif2
ficult to obser1e and measure. ,suall% biochemical or immunolo0ical assa%s and tests are
used. While cultured cells ma% 0row 1er% well in subobtimal conditions3 hi0hl% s)eciali?ed
functions usuall% re4uire near )erfect culture conditions and are often 4uicl% lost when
cells are )laced in culture.
What is Cell Culture ,sed $or?
Cell culture has become one of the ma/or tools used in cell and molecular biolo0%. "ome of
the im)ortant areas where cell culture is currentl% )la%in0 a ma/or role are briefl% described
belowD
!odel Systems
Cell cultures )ro1ide a 0ood model s%stem for stud%in0 1> basic cell biolo0% and biochem2
istr%3 2> the interactions between disease2causin0 a0ents and cells3 !> the effects of dru0s on
cells3 &> the )rocess and tri00ers for a0in03 and 9> nutritional studies.
(o1icity (esting
Cultured cells are widel% used alone or in con/unction with animal tests to stud% the effects
of new dru0s3 cosmetics and chemicals on sur1i1al and 0rowth in a wide 1ariet% of cell t%)es.
As)eciall% im)ortant are li1er2 and idne%2deri1ed cell cultures.
Cancer Research
"ince both normal cells and cancer cells can be 0rown in culture3 the basic differences
between them can be closel% studied. In addition3 it is )ossible3 b% the use of chemicals3
1iruses and radiation3 to con1ert normal cultured cells to cancer causin0 cells. Thus3 the
mechanisms that cause the chan0e can be studied. Cultured cancer cells also ser1e as a test
s%stem to determine suitable dru0s and methods for selecti1el% destro%in0 t%)es of cancer.
7
Cornin0H roller bottles are
widel% used for )roducin0
1iral 1accines.
The Cornin0 CellCubeH
bioreactor s%stem is ideal
for mass )roduction of
anchora0e2de)endent cells.
Cornin0
H
Arlenme%er flass are
often used for 0rowin0 insect
cells in sus)ension.
Cornin0 micro)lates are widel%
used for dru0 screenin0.
2irology
One of the earliest and ma/or uses of cell culture is the re)lication of 1iruses in cell cultures
=in )lace of animals> for use in 1accine )roduction. Cell cultures are also widel% used in the
clinical detection and isolation of 1iruses3 as well as basic research into how the% 0row and
infect or0anisms.
Cell3'ased !an%fact%ring
While cultured cells can be used to )roduce man% im)ortant )roducts3 three areas are 0en2
eratin0 the most interest. The first is the lar0e2scale )roduction of 1iruses for use in 1accine
)roduction. These include 1accines for )olio3 rabies3 chicen )o@3 he)atitis B and measles.
"econd3 is the lar0e2scale )roduction of cells that ha1e been 0eneticall% en0ineered to )ro2
duce )roteins that ha1e medicinal or commercial 1alue. These include monoclonal antibodies3
insulin3 hormones3 etc. Third3 is the use of cells as re)lacement tissues and or0ans. Artificial
sin for use in treatin0 burns and ulcers is the first commerciall% a1ailable )roduct. How2
e1er3 testin0 is underwa% on artificial or0ans such as )ancreas3 li1er and idne%. A )otential
su))l% of re)lacement cells and tissues ma% come out of wor currentl% bein0 done with both
embr%onic and adult stem cells. These are cells that ha1e the )otential to differentiate into
a 1ariet% of different cell t%)es. It is ho)ed that learnin0 how to control the de1elo)ment of
these cells ma% offer new treatment a))roaches for a wide 1ariet% of medical conditions.
/enetic Co%nseling
Amniocentesis3 a dia0nostic techni4ue that enables doctors to remo1e and culture fetal cells
from )re0nant women3 has 0i1en doctors an im)ortant tool for the earl% dia0nosis of fetal
disorders. These cells can then be e@amined for abnormalities in their chromosomes and
0enes usin0 ar%ot%)in03 chromosome )aintin0 and other molecular techni4ues.
/enetic .ngineering
The abilit% to transfect or re)ro0ram cultured cells with new 0enetic material ='NA and
0enes> has )ro1ided a ma/or tool to molecular biolo0ists wishin0 to stud% the cellular effects
of the e@)ression of theses 0enes =new )roteins>. These techni4ues can also be used to )ro2
duce these new )roteins in lar0e 4uantit% in cultured cells for further stud%. Insect cells are
widel% used as miniature cells factories to e@)ress substantial 4uantities of )roteins that the%
manufacture after bein0 infected with 0eneticall% en0ineered baculo1iruses.
/ene (herapy
The abilit% to 0eneticall% en0ineer cells has also led to their use for 0ene thera)%. Cells can
be remo1ed from a )atient lacin0 a functional 0ene and the missin0 or dama0ed 0ene can
then be re)laced. The cells can be 0rown for a while in culture and then re)laced into the
)atient. An alternati1e a))roach is to )lace the missin0 0ene into a 1iral 1ector and then
(infect88 the )atient with the 1irus in the ho)e that the missin0 0ene will then be e@)ressed
in the )atient8s cells.
Dr%g Screening and De*elopment
Cell2based assa%s ha1e become increasin0l% im)ortant for the )harmaceutical industr%3 not
/ust for c%toto@icit% testin0 but also for hi0h throu0h)ut screenin0 of com)ounds that ma%
ha1e )otential use as dru0s. Ori0inall%3 these cell culture tests were done in 5+ well )lates3
but increasin0 use is now bein0 made of !.& and 19!+ well )lates.
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-eferences
1. Culture of Animal Cells3 A Banual of Basic Techni4ue =155&> -. Ian $reshne%3 !rd
edition3 Alan -. Liss3 Inc.3 New Por.
2. Bethods in An?%molo0%D Cell Culture3 :ol. 9.3 =1575> W. B. Facob% and I. H. #asten3 eds.
Academic #ress3 New Por.
!. Cell and Tissue Culture =1579> Fohn #aul3 9th edition3 Churchill Li1in0stone3 Adinbur0h.
&. Animal Cell Culture Bethods3 :olume 973 =155.> F. Bather and '. Barnes3 eds. Bethods in
Cell Biolo0%3 Academic #ress3 "an 'ie0o3 155..
9. Erowth3 Nutrition and Betabolism of Cells in Culture =1572> E. H. -othblat and :. F. Cristofalo
eds. :olumes 12! b% Academic #ress3 New Por.
+. Osmolalit% of Bammalian Blood and of Bedia for Culture of Bammalian Cells3 =1576>.
C. Wa%mouth3 In :itro3 :olume +D 1652127.
7. ,nderstandin0 and Bana0in0 Cell Culture Contamination Cornin0 Life "ciences Technical
Bulletin. This is a1ailable on the Cornin0 Life "ciences web site at ww w .corn in0.com;lifesciences.
.. Eeneral Euide for Identif%in0 and Correctin0 Common Cell Culture Erowth and Attachment
#roblems Cornin0 Life "ciences Technical Bulletin. This is a1ailable on the Cornin0 Life "ciences
web site at ww w .cornin0.com; lifesciences.
5. Eeneral Euide for Cr%o0enicall% "torin0 Animal Cell Cultures Cornin0 Life "ciences Technical
Bulletin. This is a1ailable on the Cornin0 Life "ciences web site at ww w .corn in0.com;lifesciences.
16. Andoto@ins and Cell Culture Cornin0 Life "ciences Technical Bulletin. This is a1ailable on the
Cornin0 Life "ciences web site at ww w .cornin0.com;li fesciences.
$or additional )roduct or technical information3 )lease 1isit )) ) $corning$com+
li!esciences or call 1..66.&52.1116. Outside the ,nited "tates3 )lease call
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