iPSCs can't be grown with FBS since the growth factors and other proteins present in the serum will cause them to differentiate. KOSR provides similar nutrients that the cells need, but without the serum.
FBS quality varies a lot from batch to batch as it is a biological material. If you would like to use it on your iPSC ( from mouse I guess?), it is possible but you need to test each batch (lot) to check if it support the growth of your cells (number of cells, morphology) and does not cause differentiation. You can ask your supplier for ES tested serum, it can help. KO Serum replacement (KSR) is a synthetic medium and thus is less variable from lot to lot. I would recommand to use KSR.
My cells are human iPSCs and till now I have been using the ES cell serum .Based on your recommendation , I think it is better to be exchanged by KSR.
then if your cells are from human origin, yes for sure you should forget serum because human iPSC are very sensitive. Depending on your iPSC, you may also test your batch of KSR. Be carefull with your feeders. One other good alternative is to grow your human iPSC cells on matrigel in mTESR meddium: no more KSR batch to test, no more feeders.
I agree with Dr. Marie Wattenhofer-Donzé in that human iPSCs are very sensitive and you should consider to stay away from serum, due to many factors but most importantly the variabilities involved, which has caused problems for many researchers.
You can consider the mTeSR1 which is serum-free and feeder-free. It is a very widely used media to grow and maintain hESC and iPSCs in their undifferentiated state. Many cell lines and iPSCs has been cultured with mTeSR1, and the media has been featured in more than 400 publications as of date.
Here is a link for the protocol:http://www.stemcell.com/~/media/Technical%20Resources/B/C/A/2/B/29106MAN_3_0_0.ashx
Please feel free to let me know if you need more technical information.
Best regards,
Dunstan
Thank you very much dear Dr.Marie Wattenhofer-Donzé and Dunstan. Your comments were very good and helped me to made up my mind.
I would like to ask if that apply on mouse ES & iPS cells as human or not ? Thanks in advance
sir, I have to work with rat spermatoonial stem cells and have to maintain them in undifferentiated state for a while ? at present i am growing on MEF , I would like to know whether matrigel can be used for SSCs also ?
Pratibha, why don't you isolate your own rat Sertoli cell lines from juvenile testes and maintain your spermatogonia on them? It worked for mouse. If Sertoli keep proliferating it is because these clones usually secrete LIF. If you need to, you can supplement some LIF.
If you want to induction the cells to differentiation , It is possible to grow iPS cells in medium containing FBS . but the kos used to support the growth of undifferentiated ES therefore we use to maintenance of iPS cells in an undifferentiated state.
Normally human ESC and iPSC are grown in KO Serum replacement for extended periods of time. FBS is not effective for human cultures as the undefined and uncontrolled components actually can trigger spontaneous differentiation. FBS is still used when working with mouse ESC. KO Serum replacement is used when deriving and maintaining your human pluripotent stem cells on mouse feeders, or alternatively in a xeno-free manner on human feeders. Please see below for available options:
http://www.lifetechnologies.com/search/global/searchAction.action?query=knockout+serum+replacement&resultPage=1&resultsPerPage=15&autocomplete=true
Alternatively, there is a chemically defined, feeder free and Xeno free media system to culture and derive iPSC using the Esseintial 8 media systems. Please see the following link
http://www.lifetechnologies.com/order/catalog/product/A1517001?ICID=search-product
Hi,
KnockOut Serum Replacement it is used as a substitute for fetal bovine serum (FBS). It is used when we want to eliminate animals factor in the cell culture or when the serum interacts with analyzed compounds.
KnockOut Serum Replacement "is a more defined" it's mean that each bottle of the medium which we buy contains exactly the same quantity of ingredients, while fetal bovine serum (FBS) is obtained from animals, so serum obtained from different animals are different.
Fetal bovine serum (FBS) as a product contains always the same compounds but their amount is variable between each part of the product.
It is the reason why iPS are more stable at the undifferentiated stage at the medium contain KnockOut Serum Replacement.
Composition of Knockout Serum Replacement contains:
1. Amino Acids
Glycine, L-histidine, L-isoleucine, L-methionine, L-phenylalanine, L-proline, L-hydroxyproline, L-serine, L-threonine, L-tryptophan, L-tyrosine, L-valine
2. Vitamins/Antioxidants
Thiamine, reduced glutathione, ascorbic acid 2-PO4
3. Trace Elements
Ag+, Al3+, Ba2+, Cd2+, Co2+, Cr3+, Ge4+, Se4+, Br−, I−, F−, Mn2+, Si4+, V5+, Mo6+, Ni2+, Rb+, Sn2+, Zr4+
4. Proteins
Transferrin (iron-saturated), insulin, lipid-rich albumin (AlbuMAX)
Can anybody tell me what is the meaning of "Knock out" when we use this terminology for media or serum like Knockout DMEM or Knock out Serum Replacement?
Sadhan,
KnockOut Serum replacement was developed a long time ago (in the last millenium) in order to provide a replacement for serum when growing mouse embryonic stem cells, as serum drives the differentiation of ES cells. As these ES cells were mainly used within the process of Knockout mice development those days, this serum replacement was called KnockOut Serum Replacement (commonly called KSR or KOSR). In the meantime it was recognized not only to support mice ES cell growth but equally human ES and iPSC growth. Also media like Knockout DMEM were developed to support the growth of mouse ES cells within this workflow of KnockOut mice generation, hence the name.
Nowadays researchers are tending more and more to use feeder free options for culturing of their iPSCs and ESCs.
Does anyone have a referance to a paper for the composition and amounts used for KOSR?
Onur Oncer have you find some info about composition and amounts of KOSR?
I found it from some one (Justyna Augustyniak). It may be helpful.
Composition of Knockout Serum Replacement contains:
1. Amino Acids
Glycine, L-histidine, L-isoleucine, L-methionine, L-phenylalanine, L-proline, L-hydroxyproline, L-serine, L-threonine, L-tryptophan, L-tyrosine, L-valine
2. Vitamins/Antioxidants
Thiamine, reduced glutathione, ascorbic acid 2-PO4
3. Trace Elements
Ag+, Al3+, Ba2+, Cd2+, Co2+, Cr3+, Ge4+, Se4+, Br−, I−, F−, Mn2+, Si4+, V5+, Mo6+, Ni2+, Rb+, Sn2+, Zr4+
4. Proteins
Transferrin (iron-saturated), insulin, lipid-rich albumin (AlbuMAX)