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I am studying the procedures of forming hybridoma cells for generating a large number of monoclonal antibodies.

Before the procedure of fusion (with multiple myeloma cells) happens, I would like to ask how to ensure all the plasma cells in the cell extract only contains those which produce monoclonal antibodies only but not polyclonal antibodies?

I have searched in Wiki, and it says

identifying antigen-specific plasma/plasmablast cells (ASPC) that produce antibodies specific to an antigen of interest

I would like to ask what methods are available for us to do this identification?

Thank you.

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You don't do this before the fusion, you do it afterwards. First you make the hybridoma cells, which are then seeded as single cells into multiwell plates and cultivated to get enough cells for testing and further culture. This way you get only one antibody type produced per well (which is coming from a single clone, hence it is monoclonal) which then can be isolated and tested for antigen specificity.

For a schematic see the image from this publication:

enter image description here

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  • $\begingroup$ Oh thanks, so using my words (to clarify my understanding), can I say: putting individual (different types of) hybridoma plasma cells into different wells, so 1 type of cell in 1 plate --> 1 specific antibody (hence prevent polyclonal Ab formed since there are no signalling of 1 cell to multiple plasma cell lines to produce them)? $\endgroup$
    – Questions
    Jun 2 at 7:59
  • $\begingroup$ And may I ask is the purpose of screening after fusion is to make all of them immortal before culturing (which takes a relatively long time)? $\endgroup$
    – Questions
    Jun 2 at 8:01
  • $\begingroup$ @Questions Your wording "...there are no signalling of 1 cell to multiple plasma cell lines to produce them)? " is confusing to me - what do you mean by "signaling"? $\endgroup$
    – Armand
    Jun 2 at 13:12
  • $\begingroup$ @Questions The purpose of screening is to identify which clonal line is making a desired antibody - remember each clonal line (each well) makes a different antibody since it is descended from a different B cell. The immortalization has already happened from the fusion with immortal myeloma cells. $\endgroup$
    – Armand
    Jun 2 at 13:17
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    $\begingroup$ @Questions Sorry for the delay, I had other stuff to do. You do the screening after the generation of the immortal cells because you can expand these (theoretically at least) forever. And generation of this cells is nothing which is 100% effective. This way you screen cells which are immortal, can be used for antibody production. If you do it the other way, you cannot be sure to get immortal cells from your screened cells. $\endgroup$
    – Chris
    Jun 3 at 5:41

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