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hybridoma cell question - (Apr/18/2005 )

I'm using hybridoma cells to produce monoclonal antibodies. However, some problems had occured during sub cloning.

During the cell fusion, could it be possible that more than one B-cells can fuse with a single myeloma cell? If so, would the hybridoma cell be stable and secret two different antibodies??

If this is the case, how can I prevent more than one B-cells to fuse with a single myeloma? Should I reduce fusion time?

-babydoll-

Possible - probably kind of rare ~ 0.5% type of thing - I have made a couple hybridoma that had a single heavy chain and multiple light chains and I am 100% sure that the cell line was clonal.


QUOTE (babydoll @ Apr 18 2005, 12:39 PM)
I'm using hybridoma cells to produce monoclonal antibodies. However, some problems had occured during sub cloning.

During the cell fusion, could it be possible that more than one B-cells can fuse with a single myeloma cell? If so, would the hybridoma cell be stable and secret two different antibodies??

If this is the case, how can I prevent more than one B-cells to fuse with a single myeloma? Should I reduce fusion time?

-tedley7-

hi,

I believe that it could be possible but, after cell fusion, these cells are not able no grown as a stable hybridoma cell line and they were died.

QUOTE (tedley7 @ Mar 6 2006, 07:24 PM)
Possible - probably kind of rare ~ 0.5% type of thing - I have made a couple hybridoma that had a single heavy chain and multiple light chains and I am 100% sure that the cell line was clonal.


QUOTE (babydoll @ Apr 18 2005, 12:39 PM)

I'm using hybridoma cells to produce monoclonal antibodies. However, some problems had occured during sub cloning.

During the cell fusion, could it be possible that more than one B-cells can fuse with a single myeloma cell? If so, would the hybridoma cell be stable and secret two different antibodies??

If this is the case, how can I prevent more than one B-cells to fuse with a single myeloma? Should I reduce fusion time?

-tonix37-

hi all it is possible that more than two one cell might fuse with the fusion partner, and the cell might become polyploid for a while (not a tetraploid as in case of hybridoma). but then the cell looses the extra chromosomes acquired due to fusion, during cell multiplication and becomes diploid again. however many mechanisms might occcur bet. theses chromosomes like crossing over, deletions duplications etc. this might be the cause of some of the well known problems like inconsistancy in the production of antibodies, variation in H and L chains etc.

-SHIVA KESHAVA-

hi,
It is quite possible that more than one B cell will fuse to single myeloma cell, as our members said these polyploid cells are not quite stable. it is not worth to have high hopes on them.

we can avoid this situation by selcting appropriate ratio of myeloma and splenocytes. since myeloma cells far bigger than B cell u should make a ratio based on the surface phenomenon.
i would suggest 1:5 to 1:10 ration (myeloma : splenocytes) works better.
i would not worry too much on fusion time.

gud luk


QUOTE (babydoll @ Apr 18 2005, 09:39 AM)
I'm using hybridoma cells to produce monoclonal antibodies. However, some problems had occured during sub cloning.

During the cell fusion, could it be possible that more than one B-cells can fuse with a single myeloma cell? If so, would the hybridoma cell be stable and secret two different antibodies??

If this is the case, how can I prevent more than one B-cells to fuse with a single myeloma? Should I reduce fusion time?

-payeli-