Long-term storage of cultured cells
Cryopreservation of mammalian cells is extremely valuable and common in biological research. Fear of losing a cell line to contamination or incubator failure is frequently the impetus for making archival storage of cells a high priority after receiving or generating a new cell line. Once transferred from growth media to freezing medium, the cells are usually frozen at a controlled rate and stored in liquid nitrogen vapor or at -130°C in a mechanical deep freeze. Although freezing a cell line is a commonly performed procedure, problems arise when suitable freezers are not available, or unwanted variables are introduced by the presence of serum, extra-wash or complicated freezing algorithms.
Save time while saving your cells
The cell freezing media BambankerTM permits cryopreservation of cells at -80°C (or in liquid nitrogen), obviating the need for an additional and expensive ultralow freezer and avoiding time consuming and complicated controlled freezing protocols. Simply (1) harvest cells, (2) aspirate medium, (3) resuspend in BambankerTM (4) transfer to a cryovial and (5) store at -80°C. No programmed or sequential freezing is required! BambankerTM is a serum-free cryopreservation medium that is delivered ready-to-use and can be kept in the refrigerator for up to two years. Convenient 20 ml bottles are available, making BambankerTM freezing medium ideal for use by individual members of your lab. Today this innovative cell freezing medium BambankerTM is the market leader in Japan and characterized by many different published articles with very sensitive cell lines all over the world.
Serum adds variation to long-term storage
All BambankerTM products are produced with no serum. Cryopreservation media which contain sera have the disadvantage of fluctuations in recovery rates and undefined composition. Reproducibility of experiments with cells which were frozen in a serum-containing-medium, could be affected by lot-to-lot variation of the serum since the composition and concentration of proteins and other biological molecules varies with each batch of serum. This may result in issues when thawing and using cells from such serum-containing media. Breathe easy, as every ingredient of BambankerTM is precisely defined so that you can be confident that cells stored at different times will all behave and recover similarly.
Universal – even for preservation of stem cells
All common cell lines can be preserved resulting in a high number of intact cells after thawing. Recovery rates of even sensitive cells is much higher compared to regular media. For example the JCRB cell bank stores over 1,400 different cell lines with great success with BambankerTM.
There a four different version of BambankerTM:
BambankerTM prevents undesired differentiation
Cell viability and ALP staining of pluripotent stem cells. Upper row: A great number of cells are detected two days after thawing. The cells show no morphological change after thawing. Lower row: BambankerTM does not cause cell differentiation as all stem cells frozen down are still producing high levels of alkaline phosphatase, a reporter for pluripotent stem cells.
Comparison of BambankerTM with another cryopreservation medium for the cultivation of 1,400 different cell lines
The JCRB cell bank handles approximately 1,400 different cell lines. A low survival rate after thawing frozen cell lines (KHYG-1, KAI3, HL60, OVMANA) has let them to test BambankerTM and compare it to the up to then used preservation medium for the four cell lines. The growth efficiency after thawing was compared for cells stored with the currently used commercially available preservation medium and BambankerTM.
All cultured cells were harvested in the logarithmic growth phase. 1 ml preservation media was added to approximately 1 x 106 cells in a storage tube. The cells were stored for 2 weeks at -80°C. The frozen cells were thawed in an 37°C water bath and incubated at 37°C and 5% CO2 in a 96-well plate. Every day the viable cell number was determined.
The survival rate after thawing of the four cell lines (KHYG-1, HL60, KAI3, OVMANA) is with the currently used commercially available product and Bambanker™ very low. However, after thawing, all four cell lines cell proliferation was improved with Bambanker™ when compared to the currently used commercial product.
→ The JCRB cell bank has been using BambankerTM since 2014 for all their cell lines.
→ Download here AppNote for more Information
Successful frozen cells
The JCRB cell bank stores over 1,400 different cell lines with great success with BambankerTM.
- Hatoya, S. et al. Effect of co-culturing with embryonic fibroblasts on IVM, IVF and IVC of canine oocytes. Theriogenology 66, 1083–1090 (2006).
- Hikichi, T. et al. Differentiation potential of parthenogenetic embryonic stem cells is improved by nuclear transfer. Stem Cells 25, 46–53 (2007).
- Huang, Y. H., Yang, J. C., Wang, C. W. & Lee, S. Y. Dental Stem Cells and Tooth Banking for Regenerative Medicine. J. Exp. Clin. Med. 2, 111–117 (2010).
- Lechner, S. M. Interaktionen von Inseminatbestandteilen mit Epithelzellen und Leukozyten im Uterus des Rindes. (2008).
- Liu, D. et al. Relation between human decay-accelerating factor (hDAF) expression in pig cells and inhibition of human serum anti-pig cytotoxicity: Value of highly expressed hDAF for xenotransplantation. Xenotransplantation 14, 67–73 (2007).
- Mieno, S. et al. Effects of diabetes mellitus on VEGF-induced proliferation response in bone marrow derived endothelial progenitor cells. J. Card. Surg. 25, 618–625 (2010).
- Mieno, S. et al. Characteristics and Function of Cryopreserved Bone Marrow-Derived Endothelial Progenitor Cells. Ann. Thorac. Surg. 85, 1361–1366 (2008).
- Naito, H. et al. The advantages of three-dimensional culture in a collagen hydrogel for stem cell differentiation. J. Biomed. Mater. Res. – Part A 101, 2838–2845 (2013).
- Shimizu, Y. et al. Impaired tax-specific t-cell responses with insufficient control of HTLV-1 in a subgroup of individuals at asymptomatic and smoldering stages. Cancer Sci. 100, 481–489 (2009).
- Takata, Y. et al. Generation of iPS Cells Using a BacMam Multigene Expression System. Cell Struct. Funct. 36, 209–222 (2011).
- Tamai, Y. et al. Potential contribution of a novel Tax epitope-specific CD4+ T cells to graft-versus-Tax effect in adult T cell leukemia patients after allogeneic hematopoietic stem cell transplantation. J. Immunol. 190, 4382–92 (2013).
- Zaidi, S. K. et al. Runx2 deficiency and defective subnuclear targeting bypass senescence to promote immortalization and tumorigenic potential. Proc. Natl. Acad. Sci. U. S. A. 104, 19861–19866 (2007).
Does the media contain DMSO?
Yes, the only Bambanker that does not contain DMSO is Bambanker DMSOfree (BBF01, BBF02)
Can we thaw cells directly in culture medium, without a washing step?
Yes, you do not need a washing step.
Can Bambanker be used for Drosophila?
As long as single cells are used, there is no problem. We do not have experience for whole organism.
Can it be used for secondary cells.
This is no problem.
What is the minimal volume which can be used?
1 ml is recommend for 10^6 cells, you can make different downsizing 0.5 for 5×10^5 and so on.
Can cells which were frozen in Bambanker can be used for RNA isolation?
Yes. But the low amount of 10^6 cells can make a problem in the RNA Isolation and will lead to a low amount of RNA.
How can I increase the surviaval rate of Organoids by using Bambanker? We use 20 mg of stomach and colon tissue.
You can use normal Bambanker for animal tissue. Also we would recommend to make the tissue finer than 20 mg. Also you can place the tissue in Bambanker at 4°C for 10 minutes before freezing it.
For further information, please take a look in these papers:
Can I use Bambanker for freezing viruses in infected cells?
This is no problem, as long as the cells keep healthy.
Can I use Bambaker directly without bringing it back to RT?
This is no problem. Use it directly.