99:
187:
similarities there are a few key differences between these culture methods. For example, though both adherent and suspension cell cultures can be maintained in standard flasks such as the T-75 tissue culture flask, suspension cultures need to be agitated to avoid settling to the bottom of the flask. While adherent cell cultures can be maintained in flat flasks with a lot of surface area (to promote cell adhesion), suspension cultures require agitation otherwise the cells will fall to the bottom of a flask, greatly impacting their access to nutrients and oxygen, eventually resulting in cell death. For this reason, specialized flasks (including the spinner flask and shaker flask, discussed below) have been developed to agitate media and keep the cells in suspension. However, the agitation of media subjects the cells to
246:. Suspension cells are often passaged outright without changing the media. In order to change the media for a suspension culture, all cells from the current container should be removed and centrifuged into a pellet. The excess media is then removed from the centrifuged sample, and the flask is refilled with fresh media before re-adding the cells to the flask. Media changes and subculturing are important to maintain cell lines, since cells will consume nutrients in media to expand. Cells will also grow exponentially until the environment becomes inhospitable due to lack of nutrients, extreme pH, or lack of space to grow.
258:, which are limited by the surface area provided for them to expand on, suspension cultures are limited by the volume of their container. Meaning, suspension cells can exist in much larger quantities in a given flask and are preferred when using cells to make products including proteins, antibodies, metabolites or just to produce a high volume of cells. However, there are far fewer mammalian suspension cell lines than mammalian adhesive cell lines. Most large scale suspension culture involves non-mammalian cells and takes place in bioreactors.
90:. While some cell lines are cultured in suspension, the majority of commercially available mammalian cell lines are adherent. Suspension cell cultures must be agitated to maintain cells in suspension, and may require specialized equipment (e.g. magnetic stir plate, orbital shakers, incubators) and flasks (e.g. culture flasks, spinner flasks, shaker flasks). These cultures need to be maintained with nutrient containing media and cultured in a specific cell density range to avoid cell death.
212:
208:
exchange. The magnetic spinner bar itself is typically suspended from a rod attached to the central cap so that it maximizes media circulation in the cell suspension. When culturing cells, the spinner flask containing cells is placed on a magnetic stir plate, inside of an incubator and the spinner parameters need to be adjusted carefully to avoid killing cells with shear forces.
225:
optimize cell culture proliferation, the revolutions per minute of the orbital shaker must be adjusted within an acceptable range depending on the cells and media used. The media must be allowed to stir, but cannot disturb the cells too much causing them excessive stress. Shaker flasks are often used for fermentation cultures with microorganisms such as yeast.
166:(cells derived directly from a subject) must first be removed from a subject, isolated (using digestion enzymes), and suspended in media before being cultured. However, this does not mean that these cells are compatible with suspension culture, as most mammalian cells are adherent and need to attach to a surface to divide.
224:
Shaker flasks are also used for suspension cultures, and appear similar to typical
Erlenmeyer flasks but have a semi-permeable lid to allow for gas exchange. During suspension cell culturing, shaker flasks are loaded with cells and the appropriate media before they are placed on an orbital shaker. To
177:
Immortalized mammalian cell lines (cells that are able to replicate indefinitely), plant cells, and insect cells can be obtained cryopreserved from manufacturers and used to start a suspension culture. To start a culture from cryopreserved cells, the cells must first be thawed and added to a flask or
233:
Passaging, or subculturing, suspension cell cultures is more straightforward than passaging adherent cells. While adherent cells require initial processing with a digestion enzyme, to remove them from the culture flask surface, suspension cells are floating freely in media. A sample from the culture
207:
Spinner flasks, which are used for suspension cultures, contain a magnetic spinner bar which circulates the media throughout the flask and keeps cells in suspension. Spinner flasks contain one central capped opening flanked by two protruding arms which are also capped and allow for additional gas
186:
Suspension cell cultures are similar to adherent cultures in a number of ways. Both require specialized nutrient containing media, containers that allow for gas transfer, aseptic conditions to avoid contamination, and frequent passaging to prevent overcrowding of cells. However, even within these
242:). Using this information, a portion of the current suspension culture will be transferred to fresh flask and supplemented with media. The passage number should be recorded, particularly if the cells are primary and not immortalized as primary cell lines will eventually undergo
31:
130:
using fresh plasma combined with saline solutions. Carrel went on to develop the first known cell line, a line derived from chicken embryo heart which was maintained continuously for 34 years. Though the "immortality" of the cell line was later challenged by
74:. The history of suspension cell culture closely aligns with the history of cell culture overall, but differs in maintenance methods and commercial applications. The cells themselves can either be derived from
1475:
Moreira, Ana Sofia; Silva, Ana Carina; Sousa, Marcos F. Q.; Hagner-McWhirterc, Åsa; Ahlénc, Gustaf; Lundgren, Mats; Coroadinha, Ana S.; Alves, Paula M.; Peixoto, Cristina; Carrondo, Manuel J. T. (April 2020).
174:
is typically the result of an inflammatory immune response and requires specific cell-cell interactions that should not occur in a suspension of a single type of white blood cell.
991:
191:
which can stress the cells and negatively impact growth. Although both adherent and suspension cell cultures require media, media used in suspension culture may contain a
1180:
770:
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cell culture techniques, including modifying the hanging drop technique for nerve cells and introducing aseptic technique to the culture process. Later in 1910,
98:
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434:
408:
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laid the groundwork for future tissue culture, by developing a saline buffer that was used to maintain living cells (chicken embryos) for a few days.
1049:
Kurtis Kasper, Milind Singh, F.; Mikos, Antonios G. (2013-01-01), Ratner, Buddy D.; Hoffman, Allan S.; Schoen, Frederick J.; Lemons, Jack E. (eds.),
178:
bioreactor containing media. Depending upon the cryoprotectant agent, the cells might need to be washed to avoid deleterious effects from the agent.
1083:
170:
can be taken from a subject and cultured in suspension, since they naturally exist in suspension in blood. Adhesion of white blood cells
1451:
195:
to protect cells from shear forces in addition to the amino acids, vitamins and salt solution contained in culture media such as
1144:
1110:
1062:
1028:
931:
879:
746:
1405:"Effects of oxygen on recombinant protein production by suspension cultures of Spodoptera frugiperda (Sf-9) insect cells"
196:
78:
or from heterogenous cell solutions. Suspension cell culture is commonly used to culture nonadhesive cell lines like
1544:
106:
The history of suspension cell culture is closely tied to the overall history of cell and tissue culture. In 1885,
17:
1303:"Recombinant therapeutic protein production in cultivated mammalian cells: current status and future prospects"
135:, this was a major breakthrough and inspired others to pursue creating other cell lines. Notably in 1952,
1132:
1098:
1050:
1016:
734:
896:
844:
972:
Alberts, Bruce; Johnson, Alexander; Lewis, Julian; Raff, Martin; Roberts, Keith; Walter, Peter (2002).
795:
317:
1302:
1015:
Freed, Lisa E.; Guilak, Farshid (2007-01-01), Lanza, Robert; Langer, Robert; Vacanti, Joseph (eds.),
234:
can then be taken and analyzed to determine the ratio of living to dead cells (using a stain such as
75:
1203:
Li, Feng; Vijayasankaran, Natarajan; Shen, Amy (Yijuan); Kiss, Robert; Amanullah, Ashraf (2010).
820:
481:
143:. While the other cell lines were adherent, HeLa cells were able to be maintained in suspension.
119:
111:
283:
Producing cell suspension cultures to support oncolytic adenovirus used in cancer immunotherapy
640:
67:
1539:
549:
515:
424:
307:
163:
1478:"Establishing Suspension Cell Cultures for Improved Manufacturing of Oncolytic Adenovirus"
8:
139:
and his assistant Mary
Kubicek cultured the first human derived immortalized cell line -
87:
70:. Suspension culture is one of the two classical types of cell culture, the other being
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1262:"Fundamentals of Immobilised Yeast Cells for Continuous Beer Fermentation: A Review"
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55:
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Matasci, Mattia; Hacker, David L.; Baldi, Lucia; Wurm, Florian M. (2008-09-01).
973:
956:
157:
79:
51:
1365:
897:""Alexis Carrel's Tissue Culture Techniques." The Embryo Project Encyclopedia"
1533:
1503:
1428:
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1326:
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941:
845:"Alexis Carrel's Tissue Culture Techniques | The Embryo Project Encyclopedia"
635:
239:
123:
63:
59:
869:
1511:
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1477:
1452:"In vitro cell culture techniques: Adherent culture Vs. Suspension culture"
1381:
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620:
392:
107:
47:
1436:
1403:
Scott, Robert I.; Blanchard, John H.; Ferguson, Clare H. R. (1992-10-01).
1349:
1181:"Fundamental Techniques in Cell Culture - Laboratory Handbook 3rd Edition"
921:
708:"Fundamental Techniques in Cell Culture - Laboratory Handbook 3rd Edition"
536:
235:
188:
1350:"Engineering the plant cell factory for secondary metabolite production"
606:
585:
564:
529:
464:
437:
411:
385:
364:
1260:
Pilkington, P. H.; Margaritis, A.; Mensour, N. A.; Russell, I. (1998).
821:"Ross Granville Harrison (1870-1959) | The Embryo Project Encyclopedia"
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243:
192:
181:
83:
249:
630:
1474:
1259:
1131:
Link, H.; Weuster-Botz, D. (2011-01-01), Moo-Young, Murray (ed.),
1347:
1168:. www.invitrogen.com/cellculturebasics: Thermofisher Scientific.
668:. www.invitrogen.com/cellculturebasics: Thermofisher Scientific.
211:
1348:
Verpoorte, R.; van der
Heijden, R.; Memelink, J. (2000-07-01).
1097:
Klöckner, W.; Büchs, J. (2011-01-01), Moo-Young, Murray (ed.),
126:
to establish multiple tissue cultures that could be maintained
733:
Taya, M.; Kino-oka, M. (2011-01-01), Moo-Young, Murray (ed.),
238:) and the total concentration of cells in the flask (using a
680:"Cell Culture Basics: Equipment, Fundamentals and Protocols"
1205:"Cell culture processes for monoclonal antibody production"
502:
344:
140:
151:
1202:
274:
Secondary metabolite production for drugs in plant cells
30:
971:
280:
Bulk protein production for enzyme and vaccine research
1402:
1300:
182:
Suspension cell culture maintenance for laboratories
867:
122:adapted Harrison's technique and collaborated with
1048:
954:
250:Commercial applications of suspension cell culture
1449:
1017:"Chapter Eleven - Engineering Functional Tissues"
228:
1531:
1139:, Burlington: Academic Press, pp. 119–134,
1130:
1105:, Burlington: Academic Press, pp. 213–226,
1023:, Burlington: Academic Press, pp. 137–153,
1021:Principles of Tissue Engineering (Third Edition)
741:, Burlington: Academic Press, pp. 373–382,
287:
1051:"Chapter II.6.3 - Tissue Engineering Scaffolds"
277:Recombinant protein production in insect cells
1096:
955:Granger, D. Neil; Senchenkova, Elena (2010).
868:Jedrzejczak-Silicka, Magdalena (2017-05-10).
735:"2.27 - Bioreactors for Animal Cell Cultures"
1137:Comprehensive Biotechnology (Second Edition)
1103:Comprehensive Biotechnology (Second Edition)
1082:: CS1 maint: multiple names: authors list (
739:Comprehensive Biotechnology (Second Edition)
732:
54:or small aggregates of cells are allowed to
1133:"2.11 - Medium Formulation and Development"
1014:
271:Therapeutic protein production by CHO cells
992:"Cryopreservation of Mammalian Cells - US"
978:Molecular Biology of the Cell. 4th Edition
261:Some examples of suspension cell culture:
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705:
210:
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29:
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14:
1532:
1057:, Academic Press, pp. 1138–1159,
961:. Morgan & Claypool Life Sciences.
919:
152:Isolating cells and starting a culture
684:Cell Science from Technology Networks
487:Late stage (20–24 hours old) embryos
1055:Biomaterials Science (Third Edition)
923:The immortal life of Henrietta Lacks
794:Hoffman, R.M. (September 26, 2016).
771:"Subculturing Suspension Cells - US"
765:
763:
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1266:Journal of the Institute of Brewing
1172:
1158:
958:Leukocyte–Endothelial Cell Adhesion
24:
1307:Drug Discovery Today: Technologies
1279:10.1002/j.2050-0416.1998.tb00970.x
1166:Gibco Cell Culture Basics Handbook
666:Gibco Cell Culture Basics Handbook
102:SH-SY5Y cells adhered to a surface
25:
1556:
894:
796:"The Beginning of Tissue Culture"
760:
723:
696:
655:
265:Antibody production by hybridomas
202:
1450:admin.facellitate (2022-06-08).
1099:"2.17 - Shake-Flask Bioreactors"
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1409:Enzyme and Microbial Technology
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926:. New York: Crown Publishers.
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837:
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268:Fermentation cultures for beer
229:Passaging (subculturing) cells
13:
1:
648:
288:List of suspension cell lines
1421:10.1016/0141-0229(92)90095-6
7:
1319:10.1016/j.ddtec.2008.12.003
614:
10:
1561:
215:Orbital laboratory shaker.
155:
93:
1309:. Protein therapeutics.
920:Skloot, Rebecca (2010).
800:Elsevier SciTech Connect
1545:Cell culture techniques
1366:10.1023/A:1008966404981
871:History of Cell Culture
482:Drosophila melanogaster
147:Methods and maintenance
120:Montrose Thomas Burrows
114:in 1907 then developed
112:Ross Granville Harrison
34:CHO cells in suspension
1495:10.1002/biot.201900411
1221:10.4161/mabs.2.5.12720
641:Cell adhesion molecule
322:Chinese Hamster Ovary
216:
103:
35:
1482:Biotechnology Journal
550:Spodoptera frugiperda
542:Spodoptera frugiperda
516:Spodoptera frugiperda
508:Spodoptera frugiperda
379:Embryonic stem cells
214:
101:
33:
996:www.thermofisher.com
974:"Cell-Cell Adhesion"
775:www.thermofisher.com
425:Asian tiger mosquito
27:Type of cell culture
1354:Transgenic Research
358:Cervical carcinoma
402:White blood cells
355:Cervix epithelium
349:"Henrietta Lacks"
217:
104:
76:homogenized tissue
44:suspension culture
36:
1146:978-0-08-088504-9
1112:978-0-08-088504-9
1064:978-0-12-374626-9
1030:978-0-12-370615-7
933:978-1-4000-5217-2
881:978-953-51-3134-2
748:978-0-08-088504-9
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256:adherent cultures
168:White blood cells
66:, thus forming a
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133:Leonard Hayflick
72:adherent culture
50:in which single
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1488:(4): e1900411.
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1415:(10): 798–804.
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1360:(4): 323–343.
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901:embryo.asu.edu
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530:Cellosaurus
496:Cellosaurus
475:Schneider 2
465:Cellosaurus
438:Cellosaurus
412:Cellosaurus
386:Cellosaurus
365:Cellosaurus
338:Cellosaurus
331:Epithelial
236:trypan blue
84:plant cells
1534:Categories
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1152:2021-10-30
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649:References
308:Morphology
295:Cell line
244:senescence
193:surfactant
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600:Prostate
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615:See also
571:SH-SY5Y
325:Hamster
298:Meaning
128:in vitro
116:in vitro
60:multiply
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312:Links
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