Not only do we not have a roller flask apparatus, we are unable to produce these antibodies via ascites method. I need to produce enough antibodies to do rounds of IHC's. Is there anything you can recommend?
An additional solution, giving increased yields compared to T-flasks, is the CELLline system: http://www.integra-biosciences.com/sites/celline_1_e.html
May be also stacks will work, see for example
http://www.labplanet.com/search/cellfactory/
An additional solution, giving increased yields compared to T-flasks, is the CELLline system: http://www.integra-biosciences.com/sites/celline_1_e.html
I can suggest another stack system: Corning Hyperflask (for automation) or Hyperflask-M for manual use: http://catalog2.corning.com/Lifesciences/en-US/Shopping/ProductDetails.aspx?productid=10020(Lifesciences)&categoryname=HYPERFlask+Vessels(Lifesciences)
Yes, CELLline system is amazing. 0,5 to 1g per week depending on the productivity of your cells. You only change your medium once a week. The flasks are a little bit expensive but you rapidly save your money if you use them for several productions.
We now use the CellLine system for pretty much all our mAb production:
http://www.integra-biosciences.com/sites/celline_1_e.html
Also see our Web site pages for more options and how to purify mAbs:
http://users.path.ox.ac.uk/~scobbold/tig/mprod.html
You can use CELLine 350 OR 1000 bioreactor flasks. very easy to make mABs in huge amount.
Mandy, I use TC150sq. cm flasks with 100ml medium laying on their sides. When they're growing well you can pour off 50ml a day and replenish with 50ml fresh medium. Some hybs grow a little slower so harvest intervals are longer.
I've had great success with the CELLine 1000. Initially I had problems with cell death in the cell compartment, but when I stopped 're-seeding' that compartment with each harvest, and simply removed all floating cells, the ones that remained flourished.
Hi Mandi:
I agree with Dr Mader. You can culture the hybridomas in T-flasks (T175: the large ones) In one T175 you may have up to 150ml of the hybridome cells.
My suggestion is also you can use bigger botles as you want. However you be carefull and start with the secretory cells adding medium step by step. I meant first day of dilution ( from cells come from smaller botles looking for adaptation of cells and expanded after. For example for big botles (175 cm2) begin with 50 mls of medium, after 2 or three days (depending adaptation of the cell to a new botle) add another 50 mls finally complete the volume to 150. Once the hybridomes are adapted to the roller botles, you can do 1:2 dilutions or 1:20 and grow faster your hybridomes. Remember if you add since the begining all the medium, some of the nutrients as glutamine expire before reach the grow required.
I may be able to produce ascites for you. Please contact [email protected]
Have you determined that the cell line is actually SECRETING the antibody? This can can be done in 24 well plates. Collect supernatants at 96 hrs and check by ELISA. If less than 10 ug/ ml, we subclone for a high producer. Some lines drift to making just light chain or fragments.
We use large flasks and do just fine....when the line is secreting.
I also have great experience with the celline.High antibody production can be reach in these flasks.
I agre with Dr Vitetta. Establishing that you have a secreting hybridoma producing the specificity and concentration of antibody desired is essential. Note that commercial hybridoma cultures do no always perform as described. Not geting production in
ascities model suggests your line may not producing secreting antibody.
You can produce large quantities of monoclonal antibodies (may be in mg) in form of ascites. If you are not getting production of ascites you please make sure if your mice inbred and you are using the right strain of mice.Which cell line you are using?
in hollow fiber systems (http://www.fibercellsystems.com/documents/ABL-Cadwell.pdf) you can reach IgM-concentrations up to 16 mg/mL. Simply to purify by euglobulin precipitation (at the IEP of the monoclonal antibody, by dialyzing vs. very low contented buffers [0.001 mol/L])
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