there are millions of proteins are immunogenic, however there are only several used as model antigens in immunological studies. One of the best known is chicken ovalbumin. Could someone tell me why ovalbumin is chosen as a model antigen? Which special properties of ovalbumin attracted the attention of scientists? Thanks!
In the beginning, when immunologists discovered antibodies, the first efforts were focused on understanding the mechanism of antigen recognition. At that time, Sigma did not offer the plethora of reagents it does now, and obtaining abundant purified protein for such studies relied on abundantly available eggs, which is the source of ovalbumin (OVA). Further, the amino acid sequence comparisons of OVA from different types of bird and fowl allowed immunologists to define conserved and antigen-specific sequences; By characterizing Ab binding to these proteins scientists were able to map the immunogenic epitopes of the OVA antigen. Because of its long history, the structure-function relationship of OVA is one of the best characterized and understood antigens, in which both Ab and T cell epitopes have been mapped for many strains of mice. Therefore, it was one of the first antigen genes used as a transgene to make transgenic mice. Many different transgenic mouse models are available - systemic OVA expression (driven by the ubiquitously expressed b-actin promoter), tissue-specific OVA expression (eg. using the insulin promoter to drive the transgene expression, for studying type I diabetes), or in different isoforms (secreted or cell-membrane bound ), and more recently as inducible transgenes. These models are well characterized, and have contributed to our understanding of immunogenicity and tolerance. In addition, because of OVA's immunogenicity, the transgene is often introduced into non-immunogenic tumor cell lines to study cancer immunology. Hope this gives you some perspective on OVA, and other similar non-threatening antigens, such as Hen Egg Lysozyme (HEL). (Although these proteins can be antigen targets of human allergy responses to eggs.)
In the beginning, when immunologists discovered antibodies, the first efforts were focused on understanding the mechanism of antigen recognition. At that time, Sigma did not offer the plethora of reagents it does now, and obtaining abundant purified protein for such studies relied on abundantly available eggs, which is the source of ovalbumin (OVA). Further, the amino acid sequence comparisons of OVA from different types of bird and fowl allowed immunologists to define conserved and antigen-specific sequences; By characterizing Ab binding to these proteins scientists were able to map the immunogenic epitopes of the OVA antigen. Because of its long history, the structure-function relationship of OVA is one of the best characterized and understood antigens, in which both Ab and T cell epitopes have been mapped for many strains of mice. Therefore, it was one of the first antigen genes used as a transgene to make transgenic mice. Many different transgenic mouse models are available - systemic OVA expression (driven by the ubiquitously expressed b-actin promoter), tissue-specific OVA expression (eg. using the insulin promoter to drive the transgene expression, for studying type I diabetes), or in different isoforms (secreted or cell-membrane bound ), and more recently as inducible transgenes. These models are well characterized, and have contributed to our understanding of immunogenicity and tolerance. In addition, because of OVA's immunogenicity, the transgene is often introduced into non-immunogenic tumor cell lines to study cancer immunology. Hope this gives you some perspective on OVA, and other similar non-threatening antigens, such as Hen Egg Lysozyme (HEL). (Although these proteins can be antigen targets of human allergy responses to eggs.)
Yes-basically it is an easily obtained and abundant protein when first used.
Another reason for the current popularity of OVA as a model antigen are research tools such as OT-1 and OT-II OVA-specific TCR transgenic mice and monoclonal antibodies such as 25D.1 that bind to MHC class I complexes plus an OVA peptide (Kb-SIINFEKL).
Very good point Pamela. The TCRs of OTI and OTII transgenic mice and epitope specific Abs recognize defined OVA epitopes presented by MHC class I (Kb) and II (IA) molecules. In addition there is the DO11.10 BALB/C transgenic mouse model that sees the same SIINFEKL peptide presented by the Kd molecule, but at a lower (and some argue more physiological) affinity.
Responding to Amin's question:
I believe that instead of isolating OVA from eggs, recombinant OVA protein can be isolated from transformed bacterial cultures. Because of that, immunologists became concerned regarding levels of contaminating LPS, which, as a TLR4 agonist, can itself activate the immune system and mask antigen-specific responses. But I am not an expert on this topic. Just be sure, for immunological studies, you get certified LPS-free OVA protein.
Essentially, yes - unless you buy Endotoxin(LPS)-free grade OVA from other vendors. It looks like Sigma may not make this - some report that the highest grade (V) OVA from Sigma still has endotoxin. I must correct my answer above - the OVA is still purified from egg whites, but because chicken coops are rife with bacteria, that seems to be the source of endotoxin contamination.
Here is a recent Plos One article - check the methods
http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0098648
www.hyglos.de
http://www.youzre.com/_d276299191.htm
I have a few follow-up questions if anyone is still following this thread. Where does exogenous ovalbumin localize? Is it cytoplasmic, secreted, or other? Also, does the expressing cell need to die in order for DC cross-presentation? Cheers.
Michael Davis The location of exogenous ovalbumin can vary depending on the stage of processing (e.g. early endosome, late endosome, lysosome etc.). It can be retained in phagosomes and either processed in the endosomes or escape into cytosool. I have seen that cells dye and still enhance cross-presentation, but most of the case, cells do not necessiarly need to die for DC cross-presentation. Thank you,
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