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Adjuvants

immunization

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6 replies to this topic

#1 neuron

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Posted 18 October 2013 - 01:42 AM

Hi,

 

I don't know much about immunology, just remember very few basic things, that I had read long backrolleyes.gif . For some of my work, I was reading about generation of antibodies. Adjuvants play a major role in this. And in mice/rabbit I read that mostly FCA is used. From my basic understanding, I know that adjuvant helps immune system in recognition and elicit immune response. Now I have a question, what if the antigen itself is strong enough to be recognized by immune system, do we still need adjuvant?

 

Also I was reading about various types of adjuvants, and came across to CpG. It is not emulsion based and just a dinucelotide. If antigen is very labile and mixing with adjuvant is a problem (as it needs vigorous mixing), can this CpG be used?

 

Please correct me if I am wrong somewhere and clear my doubts.

 

 

Thankssmile.png



#2 Missle

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Posted 18 October 2013 - 04:10 AM

There are many different adjuvants that can be used, including IFA & CFA.  I believe adjuvants have less to do with recognition and more with recruitment.  They have also been classified as 'danger signals' as they kind of put the immune system on high alert, which can boost your response.  If the antigen is immunogenic enough on it's own then you don't have to have an adjuvant but you will likely get a much stronger response if you do have one.

 

CpG can be used.  In nature, it elicits a stronger innate response than humoral but has some success in vaccines.  Here's a reference: http://www.ncbi.nlm....pubmed/21506647



#3 neuron

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Posted 20 October 2013 - 09:39 PM

Thanks Missle! for clearing my doubtssmile.png. I agree that adjuvant is required for recruitment, as you said that if antigen is strong enough then there is no need for adjuvant. But then I was wondering, if anybody has ever done that? 

 

Thanks for the reference also, I have seen this paper, here also I have an argument. They say that CpG can be used as an adjuvant directly, I mean that rigorous mixing is not required, then this should be the best adjuvant. Isn't it? I thought many people would be using it nowadays, but I dint find many references. Here is one-

 

http://onlinelibrary...1/cei.12146/pdf



#4 Missle

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Posted 23 October 2013 - 04:12 AM

Hi.  Sorry for the delayed response.  I have another reference (a commercial companies website) http://www.invivogen...ccine-adjuvants that provides a nice review on adjuvants and TH1 vs. TH2 immune response.  I don't think that mixing vigorously is usually much of a problem.  If the protein becomes aggregated - that's actually good for acheiving a good immune response & the immune system will break it apart anyway.  If it causes a little fragmentation, that is usually ok too.  A traditional alum or emulsion adjuvant has the additional benefit of serving as a 'time release', releasing the antigen a little at a time to increase presentation.  Look over the website linked above, I think you'll like it - it has solid references and science but is clear an concise too (love that combination!).



#5 neuron

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Posted 23 October 2013 - 08:09 PM

Thanks Misslesmile.png , I just found out this link, and you also sent. Thanks a lot. I completely agree that mixing should not be a problem, as this is the way it has been done. But as I told earlier also, I am not at all an immunologist, so my questions could be silly..so please bear with mesmile.png . What if you want an antibody against a particular confirmation of a protein or you want an antibody against interacting protein? How would you inject? Mixing with adjuvant can change the confirmation of protein or it can break the interactions between proteins? I was looking for some reference, where somebody would have done that, but I couldn't find that. Though as you said that alum adjuvant has benefit of slow release of antigen, but look at this reference-

 

http://www.fasebj.or.../26/3/1272.long

 

Isn't is contradictory?



#6 Missle

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Posted 24 October 2013 - 04:02 AM

Well, lets separate the two goals of an antibody with a desired conformational epitope and that of one against an interacting protein.  The native conformation does not have to absolutely be maintained in order to acheive a desired conformational epitope - it all comes down to the screening of the hybridoma clones.  The screening methodology has to be capable of identifying the antibodies that are to your desired conformational epitope.  As long as some conformation is maintained or some molecules of the protein maintain the targeted epitope conformation then it is possible to generate the antibodies you want - you just have to be able to fish them out.

I've never made an antibody against an interacting protein.  Do you mean two proteins bound non-covalently and you want the epitope of the antibody to be at their juncture?  That would be tricky.  You would either need to find a way to maintain that interaction or synthetically produce a protein with the desired conformation.  Tricky.

 

I agree that your reference does cast doubt regarding the 'time release' nature of alum.....I don't think that it speaks to what is a better adjuvant and, personal opinion, that is not possible looking at only one (or even 2 or 3 or 4) antigen.



#7 neuron

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Posted 27 October 2013 - 07:58 PM

Ok for confrontational antibody I agree that it depends on how we screen the hybridoma, but if we have to generate antibody against interacting protein? As you said its really tricky. Suppose some small molecule interacts with  2-3 proteins to initiate some cascade of events, but these interactions are really transient, if the proteins dissociate with this molecule they come back to their original confirmation. Did I make sense?unsure.png

 

And if the total molecular weight of small molecule-protein complex is in 1000s kDa, there should not be any need to adjuvant? Because adding of adjuvant can lead to loose the interaction?






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