Monoclonal Antibody Production and the Culturing of Mouse Hybridoma Cells.

Introduction

The production and maintenance of a hybridoma cell begins with the fusion of a specific antibody producing B cell, to a cancer B cell called a myeloma, which does not produce an antibody by itself. Fusion results in an immortalized line called a hybridoma that will faithfully produce a specific antibody against a single epitope called a monoclonal antibody. Once produced, proper maintenance and culturing is required to maximize the performance and continued production of the antibody in question. Have you ever wondered how this is done? Are you fusing cells to produce your own monoclonal antibodies or wondering about your culturing options. This is your opportunity to ask questions about monoclonal antibody production and the culturing of mouse hybridoma cells.

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Question 2

I am looking into protocols for making hybridomas and notice that some people grow their myeloma cells in 8-azaguanine and other don’t. What is the purpose?

Some people grow their myeloma cells (Sp2/0, Sp2/0-ag14, NS-0 and NS-1 cells for example) in 8-azaguanine or 6-thioguanineThe reason for this is to ensure that the cell line is 8-azaguanine or 6-thioguanine and HGPRT negative. Azaguinine and 6-thioguanine are competitors of guanine. This phenotype is necessary for the myeloma cells to be sensitive to the … Continued

Question 4

I am looking for the most inexpensive way to culture hybridoma cells in serum free conditions. I am starting with classic media, is there anything that is serum free that I can add to classic media to allow for serum free culture or do I need to use a pre-made serum free medium for hybridoma cells.

Moving cells from serum-containing to serum-free media is not so easy since adaptation is required. To my knowledge there are no additives such as a non-animal derived substitute that can be added to a basel (classic) medium that will make it perform like a serum containing medium. For your information there are media such as … Continued

Question 5

Do i need to inject a 100% pure antigen in order to get a desired results or can I use a 80% purity antigen for injection, as the antigen I am dealing with will only grow along with the bacteria. I’m also using non spf female balb/c mices for the injection. I’m wondering if all these factors may affect to get the desired results.

About your antigen, if you were making polyclonal antibodies the purity would be a problem. Technically, by making monoclonal antibodies you can still get the one you want. But I do envision some problems. Obviously your search for the MAb you want will be made more difficult due to the 20% non-antigen directed Abs. The … Continued