Inoculum inoculum should be grown to a density of 4. Insect cell cultures are now commonly used in insect physiology, developmental biology, pathology, and molecular biology. Being eukaryotic, insect cells allow correct folding and posttranslation modification, and purified protein can. As the field has advanced from methods development to a standard procedure, so has the diversity of scientists using the technique. Insect cell culture, particularly the culture of lepidopteran cell lines, has rapidly evolved into a large research effort sinee grace established the first eell lines in. Lynn1 usdaars, insect biocontrol laboratory, barcwest, building 011a, room 214, beltsville, maryland 207052350. First, most insect cell culture media utilize phosphate buffers rather than carbonatebased buffers commonly used for mammalian cells.
This manual is included with sf9, sf21, and high five insect cells and provides general information. In 1917, just two years after goldschmidts pioneering attempts to culture insect cells, glaser 11 attempted to grow a polyhedral virus in cultures of insect hemocyte cells. The need for new and superior insect cell lines for expression of recombinant proteins. Cell culture has become one of the most fundamental techniques for modeling biological systems, and is of increasing importance in the biotechnology and pharmaceutical sectors as well as an essential. A confluent monolayer is an adherent cell culture dish, plate or flask in which the cells have formed a single layer over the entire surface area available for growth.
Also, since insects are coldblooded, the cells will grow at a much wider range of temperatures. The metabolic condition of the culture and growth medium used also can affect the timing of baculovirus infection. This paper describes techniques for the development of continuous cell lines from insect embryos and other tissues. Cell culture company custom mammalian and insect whole cell. Insects cell lines are commonly used in place of prokaryotic ones because posttranslational modifications of proteins are possible in insect cells whereas this mechanism is not present in. The relative advantages of different tissues embryos, ovaries, etc. A confluent monolayer is an adherent cell culture dish, plate or flask in which the cells have formed a.
Growth and maintenance of insect cell lines the wolfson centre. Cell culture company remains committed to meeting our customers needs. Both the cells lines are valuable choices, but sf9 cells that are a subclone of the sf21 cells are often preferred due to their faster growth rate and higher cell densities. Jan 29, 2016 high level recombinant protein production in insect cell culture kamal rashid, ph. In case of small cultures, shaker or spinner culture flasks may provide adequate starting materials but for bigger productions, the growth of insect cells must be carried out in bioreactors and the culture is constantly monitor with advanced.
In developing midgut epithelial cell lines, we found that traditional methods in. Insect cell culture technology in baculovirus expression. Goosen queens university, ontario, canada the primary development in largescale insect cell culture over the past year has been the continuing accumulation of documented evidence fundamental and applied that conventional aerated stirredtank and airlift bioreactors may be employed for insect cell cultivation. Using the advent of new techniques and instruments in molecular. Bishop the ability to produce large quantities of proteins relatively cheaply and easily, by. A great variety of insect cells protein expression systems are available with different vectors that allow different applications orbigen, bd, clontech, life technologies etc. The sf21, sf9, and the btitn5b14 or high five cells are the most commonly used insect cell lines in research. Insect cell culture gas mixer gas blender equipment gas. Insect cell culture is a common choice for heterologous protein expression. A process has been developed for recombinant production of selected polypeptides using transformed insect cells cultured in a horizontally rotating culture vessel modulated to create low shear conditions.
Insect cell cultures are widely used in viral diagnosis and biotechnology, for the production of recombinant proteins, viral pesticides and vaccines as well as in basic research in genetics, molecular. The field has grown to the extent that now over 500 insect. Establishment and maintenance of insect cell lines. High level recombinant protein production in insect cell culture. Lynn1 usdaars, insect biocontrol laboratory, barcwest, building 011a, room 214, beltsville, maryland 207052350 summary the success of insect cell culture is demonstrated by reports of over 500 established cell lines. Expression systems provides tools to support the baculovirus and host cell expression system including baculovirus vectors and kits to determine baculovirus production. The sf21, sf9, and the btitn5b14 or high five cells are. The insect cell culture based protein expression system is inexpensive and easy to scale up. Gibco insect media has been formulated for maximum growth and protein yields. The following morphological changes are typical of monolayer sf9 cells infected with recombinant acnpv. Pdf insect cell culture for industrial production of.
Researchers familiar with vertebrate cell culture should be aware of a few differences that relate to the differences in the biology of insects as compared with mammals. Goosen queens university, ontario, canada the primary development in largescale insect cell culture over the past. Your use of this pdf, the bioone complete website, and all posted and associated content indicates. It is a testament to their scientific insights that these media still form the basis of most mammalian cell culture research being performed today. Cell lines can be successfully established from tissues or cells of insect larvae or pupae. Insect cell culturebaculovirus expression jennifer griffiths 22306 general information about insect cell culture there are many advantages to using baculovirus for gene expression, the foremost of which is. Pdf baculovirus and insect cell expression haitham amer. Using the advent of new techniques and instruments in molecular biology as well as the advancement in biotechnology, the indigenous cell lines might prove useful in the development of alternatives to conventional chemical pesticides for agriculture and in the creation of. The most widely used insect cells are the sf9 and sf21 cell lines.
Graces insect cell culture medium, supplemented instructions for use product description graces insect cell culture medium is commonly used to support cell growth, protein expression, and. Following krp singhs pioneering research in 1967, a large number of cell lines from diptera, hemiptera, and lepidopteran insects were. The sf9, sf21, and high fivez cell lines are suitable for use in expressing recombinant proteins with baculovirus and other insect expression systems e. The use of insect cells in baculovirus expression vectors described elsewhere in this book has proven to be a blessing to the whole field of insect cell culture by creating a reliable market for insect cell culture media. Proteolysis is a problem of the bevs system due to its lytic nature, and can, therefore, be a critical issue in insect cell bioprocessing. Guide to baculovirus expression vector systems bevs and. Cell culture company custom mammalian and insect whole. Contract mammalian cell banking, cell culture, purification and analytics. Insect cell culture medium additional products expression systems offers a variety of media formulations for insect and mammalian cell culture. In particular, insect cell culture is an indispensable tool for the study of insect viruses. This means a co 2 atmosphere is not necessary and thus simplifies the incubators needed for insect cells. For largescale production or basic research, insect cells are able to express large quantities of protein with complex posttranslational modifications. Insect cell cultures grow as suspension or as monolayer cultures at 30c.
The culture of insect cells in the absence of serum is reaching maturity, and promising serum substitutes hydrolysates, new growth and productionenhancing factors are being evaluated. Cell lines introduction this manual is included with sf9, sf21, and high fivez insect cells and provides general information about the growth and maintenance of insect cell cultures. To initiate cell culture immediately upon receipt, see page 10. Tissue culture treated dishes for seeding insect cells appropriate complete insect cell culture medium e. Insect cell culture technology in baculovirus expression systems lain r. Growth and maintenance of insect cell lines product information this manual provides general information about the growth and maintenance of insect cell cultures. In particular, insect cell culture is an indispensable tool for the study. Design of an efficient medium for insect cell growth and recombinant. For largescale production or basic research, insect cells are able to express large quantities of protein with complex post. Wo1994026087a3 recombinant protein production and insect. Jul 01, 2017 a great number of cell lines from hemipteran, lepidopteran, and dipteran insects have been developed and characterized. The formulations created by cell culture pioneers like eagle, dulbecco, ham, and others can be found below.
The insect cellbaculovirus vector system has become one of the favorite platforms for the expression of viral vectors for vaccination and gene therapy purposes. High level recombinant protein production in insect cell culture kamal rashid, ph. High level recombinant protein production in insect cell. A great number of cell lines from hemipteran, lepidopteran, and dipteran insects have been developed and characterized. Most mammalian recombinant glycoproteins produced by baculovirus expression vectors in insect cell culture are produced with mannoseterminated oligosaccharides or otherwise less.
Aav production culture characteristics in the present study, we investigated aav production using regularand high cell density cultures. Graces insect cell culture medium, supplemented instructions. Graces insect medium with lglutamine, without sodium. Graces antheraea eucalypti moth cells grow as suspension cultures. Nlinked glycosylation of a baculovirusexpressed recombinant. The sf9, sf21, and high fivez cell lines are suitable for use in expressing recombinant proteins with. This paper describes methods that are effective for maintaining various insect cell lines. Insect cell cultures are widely used in studies on insect cell physiology, developmental biology and microbial pathology. Pdf insect cell culture and biotechnology researchgate. Cell culture is the process by which prokaryotic, eukaryotic or plant cellsare grown under controlled conditions in practice it refers to the culturing of cells derived from animal cells.
The implications and value of insect tissue cultures in the study of insect viruses were realized very early. Pdf on jan 1, 2010, spiros n agathos and others published insect cell culture find, read and cite all the research you need on researchgate. Current methods of insect cell culture have produced a limited variety of cell types in an ever expanding list of insect cell lines. Insect cell culture for industrial production of recombinant. Building on earlier research, insect cell culture began with the successful establishment of one cell line from pupal ovarian tissue. Methods for the development of cell lines from insects. We are fully operational and prioritizing production of covid19 testing materials. Pdf insect cell culture and applications to research and.
Graces insect cell culture medium, supplemented instructions for use product description graces insect cell culture medium is commonly used to support cell growth, protein expression, and baculovirus vector production for spodoptera frugiperda insect cells such as sf9 and sf21. Introduction to sf9 insect culture in a stirredtank. As the field has advanced from methods development to a standard. Key words insect cell lines, tissue culture, cell culture. Cell culture basics handbook thermo fisher scientific. The metabolic condition of the culture and growth medium used also can affect the timing of. New approaches to insect tissue culture springerlink. Development and characterization of insect cell lines. Startup culture from freezer stocks sterile cell culture technique must be used at all times. About your presenter kamal has over thirty years of academic experience in both research and biotechnology educational program development plus an additional focus on training at universities. Pdf baculovirus and insect cell expression haitham.
Confluency confluency is a marker for when to subculture your cells. Insects cell lines are commonly used in place of prokaryotic ones because posttranslational modifications of proteins are possible in insect cells whereas this mechanism is not present in prokaryotic systems. Insect cells used in conjunction with the baculovirus expression vector system bevs are gaining ground rapidly as a platform for recombinant protein production. In order to start a culture from a frozen stock follow the steps below. Insect cell culture baculovirus expression jennifer griffiths 22306 general information about insect cell culture there are many advantages to using baculovirus for gene expression, the foremost of which is that baculovirus expression permits folding, posttranslational modification and oligomerization in manners that are often. Dec 15, 2010 insect cell cultures are widely used in studies on insect cell physiology, developmental biology and microbial pathology. Find the right classical media formulation for your mammalian cell culture. Insect cell culture an overview sciencedirect topics. Cell culture cell culture is one of the major tools used in cellular and molecular biology, providing excellent model systems for studying the normal physiology and biochemistry of cells e. The first continuously growing insect cell cultures were established from lepidopteran insects around 1960. Insect cells in culture are currently commanding great interest as superior hosts for the efficient production of biologicals with applications in health care and in.
Insect cell cultures are widely used in viral diagnosis and biotechnology, for the production of recombinant proteins, viral pesticides and vaccines as well as in basic research in genetics, molecular biology, biochemistry, endocrinology and virology. The complete strategy for maximizing the yield of recombinant proteins from insect cell culture must include an optimization of the culture conditions during the growth phase as. Sf9 and sf21 cells are very similar and both derive from the pupal ovarian tissue of the fall army worm spodoptera frugiperda. The use of insect cells in baculovirus expression vectors described elsewhere in this. Methods include how to obtain aseptic primary tissues, how to dissociate the tissue, and then how to maintain the primary culture until cells grow consistently. Sf9 or sf21 trans it insect transfection reagent poet transfer plasmid or other compatible vector containing gene of interest. Insect cell culture as said before, bevs involves the use of a specific virus, capable of infecting only lepidopteran cells. The sf9 cell line is one of the most commonly used lines in insect cell culture. Growth and maintenance of insect cell lines user guide pub. Insect cells are suitable for both small and largescale cultures.
Similarly, most of the commercially available insect cell lines grow well in monolayer or suspension culture. Consolidating and expanding current, fundamental notions of virology and animal cell cultivation, this practical reference examines the development of insect cell culture techniques for the production of recombinant proteins and insect pathogenic virusesresolving onthejob problems such as sparging cell damage and reduced infectivity cells, insect cell culture engineering. Pdf the continued development of new cell culture technology is essential for the future growth and application of insect cell and baculovirus. In developing midgut epithelial cell lines, we found that traditional methods in insect cell culture failed to provide healthy cells from mature tissues. The need for new and superior insect cell lines for expression of recombinant proteins became important and research in insect cell culture gained renewed interest. The sf9, sf21, and high five cell lines are suitable for use in expressing recombinant proteins with baculovirus and other insect expression systems e. May 20, 2002 insect cell cultures are now commonly used in insect physiology, developmental biology, pathology, and molecular biology. Insect cell culture has been useful in the studies of morphogenesis, virology, pathology, biochemistry, genetics and many other fields of biology and medicine. Insect cell culture oers major reference works wiley.