Is IgG monoclonal antibody better then IgM? why? - why everyone prefer IgG? (Sep/03/2010 )
Hi all,
I would like to generate a monoclonal antibody and I suspected that I mostly got IgM. Most people are talking about raising IgG monoclonals and I was wondering why is IgG better then IgM? I know that IgM is larger and more difficult to purify becasue it is less soluble and easier to denature compared to IgG. On the otherhand , IgM has higher affinity towards the antigen as it has more antigen binding site, is it correct? Thanks in advance ^^
Best Regards,
Vivian
I'd assume that IgG monocolonals are more popular because, as you said, they are easier to generate and purify and then more stable to work with.
IgM monoclonals may have more binding sites but surely only one antibody can bind to one antigen on the cell surface due to spatial flexibility? I think the multiple binding sites of IgM would only really matter if you were using the antibody to neutralise a substance that was in solution.
Loads of companies produce antibodes and a large number of these are IgG antibodies. I'd take from this that they've tried and tested their methods for Ab production and have ended up favouring IgG Abs. Though when it comes to raising an Ab for your specific antigen it could be wise to try both!
Piersgb on Mon Sep 13 10:18:49 2010 said:
IgM monoclonals may have more binding sites but surely only one antibody can bind to one antigen on the cell surface due to spatial flexibility? I think the multiple binding sites of IgM would only really matter if you were using the antibody to neutralise a substance that was in solution.
I would not necessarily agree. IgMs have 10 binding sites, which means that they replace low affinity with high avidity. That's why they are the first antibody isotype in response to an antigen. That being said, in our hands, IgMs can have quite high non-specific binding. This is much reduced with IgGs, that only have two binding sites. And because IgGs undergo affinity maturation, they usually have a much higher affinity, than the IgMs.
Another reason why IgGs are preferred is that they can be used in various assay types e.g. ELISA, WB, dot blot, IHC, etc... They bind protein A/G, so they can be used in immunoprecipitation, wheras IgMs can't.
In our experience if we obtain IgMs it's because the immune response as a whole was low. I hope you are using complete Freund's adjuvant for immunizations. What type of antigen do you have (protein, carbohydrate)?
Miha
BioMiha on Tue Sep 14 07:37:30 2010 said:
Piersgb on Mon Sep 13 10:18:49 2010 said:
IgM monoclonals may have more binding sites but surely only one antibody can bind to one antigen on the cell surface due to spatial flexibility? I think the multiple binding sites of IgM would only really matter if you were using the antibody to neutralise a substance that was in solution.
I would not necessarily agree. IgMs have 10 binding sites, which means that they replace low affinity with high avidity. That's why they are the first antibody isotype in response to an antigen. That being said, in our hands, IgMs can have quite high non-specific binding. This is much reduced with IgGs, that only have two binding sites. And because IgGs undergo affinity maturation, they usually have a much higher affinity, than the IgMs.
Another reason why IgGs are preferred is that they can be used in various assay types e.g. ELISA, WB, dot blot, IHC, etc... They bind protein A/G, so they can be used in immunoprecipitation, wheras IgMs can't.
In our experience if we obtain IgMs it's because the immune response as a whole was low. I hope you are using complete Freund's adjuvant for immunizations. What type of antigen do you have (protein, carbohydrate)?
Miha
Thank you very much for both for you ^^
I am using complete Freund's adjuvant and incomplete Freund's adjuvant for immunizations. I used purified protein as antigen. I have performed 6 immunizations in 3 months time so I think the class switching have completed? anyways, I guess I can only try more fusion and see if it helps to generate hybridoma that secrete IgG ... Thanks a lot.