Is it possible that one could generate specific monoclonal antibody in lab? If yes, then how much it is difficult ? I want to generate monoclonal antibody in lab but i didn't have any protocol giving detail step by step process. I will appreciate if someone send protocol or send link for that protocol. email ID is- [email protected]
Thanks
Hi, there are some informations and protocols:
UNIT 2.5 Production of Monoclonal Antibodies
http://onlinelibrary.wiley.com/doi/10.1002/0471142735.im0205s102/abstract?systemMessage=Wiley+Online+Library+will+be+unavailable+on+Saturday+3rd+September+2016+at+08.30+BST%2F+03%3A30+EDT%2F+15%3A30+SGT+for+5+hours+and+Sunday+4th+September+at+10%3A00+BST%2F+05%3A00+EST%2F+17%3A00+SGT+for+1+hour++for+essential+maintenance.+Apologies+for+the+inconvenience
http://www.umass.edu/vetimm/docs/Wagner_Hybridoma.pdf
http://www.cshlpress.com/pdf/sample/2013/Antibodies2/AB2Intro7Part1.pdf
Hey
The production of Monoclonal Abs not difficult so not easy ,its need special lab have facilities of cell culture,
,The technique of producing hybrid cell lines called “hybridomas” by the fusion of fusing a specific antibody-producing lymphocyte B cell with a myeloma cell that has an ability to grow in tissue culture .
Antibody-producing lymphocyte B cell isolated from the spleen or peripheral blood of animal immunized with Specific antigen
Single myeloma cell is a bone marrow tumour cell capable of multiplying indefinitely.
Fused hybrid cells or hybridoma have the antibody producing capability inherited from lymphocytes and have the ability to grow continuously or immortal like malignant cancer cells.
in briefly
Immunization of an animal such as mouse
Isolation of B cells from the spleen or peripheral blood
Cultivation of myeloma cells
Fusion of myeloma and B cells
Separation of cell lines
Screening of suitable cell lines
in vitro (a) or in vivo (b) multiplication
Harvesting
this technique at least 6 months need.
Find PDF attached described the protocol in detail
Thanks you very much both of you for providing information. Could we use supernatant of hybrid cells as antibody or else we need to do downstream process to get antibody?
You can use the supernatant, normally you need to screen a few thousands clones before you find your antibody, that s not doable if you have to purify them
But I emphasize again.... Hybridoma technologie is not something you do untrained. Its definitely something you just read the protocol and grow some cell lines. Especially the fusion process is tricky and to keep everything clean is another challenge. We have dedicated labspace where no one enters who is not trained. And... you are working with animals, so ethically you should know what you are doing so you are not wasting animal life because you are unskilled.
And from what I read here you sound pretty unskilled.
There are plenty of companies who will develop mAbs for you if you provide them the antigen, I can only advise you to look into outsourcing or find a partner who gives you proper training
I would suggest you to find a collaborating partner with experience and the right cell culture facilities. If you are in this kind of lab, you could do it without too many problems and be able to set up your own facilities in the future if you want. Once you have the hybridomas, ist not difficult to purify large amounts of the antibodies for downstream applications. Cell supernatant contains very low amounts of antibodies, although for some particular applications this is enough.
Alternatively, you could find a partner with experience in recombinant antibodies generation, for instance using phage display. Antibody discovery phase should be faster and animals are not required at all, but subsequent production (for instance using mammalian cells) can be very challenging for some antibodies.
The choice of one method or the other mainly depends on the expertise in your lab and in the collaborating labs.
Producing mAbs is not difficult but is laborious work, and you should have a mount concentration of antigen for immunize mice and a good myeloma. as a fusion partner and also an isolated Macrophages ( MQ) from Balb/C mause.
The added publication will give you an overview of the procedure to make monoconals in your own lab. I hope, that you will find this useful. All the best from Holland !
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