I think ELISA technique will help Annabel Curle

Samuel Oluyomi Ayodele thank you for your answer. Would you mind explaining this further please? I would like to strip and collect the surface antigen in order to expose the DCs to the antigen prior to the immunogenicity experiment.

Annabel - I am not clear about why you want to strip surface antigens off cells and what cells you want to do this to? I have worked in the fields of immunolgy and hematology for a number of years, and looked at inducing cellular and antibody responses to antigens, viruses and cells, and stripping off cell surface antigens isn't something that is done routinely.

But there is a technique for removing cell surface proteins from cells of interest: digestion with the protease bromelain, which is derived from pineapples. This was done to expose phosphoryl choline residues on red blood cells, which were then used to test the binding of anti-phosphoryl choline antibodies.

I personally have never done this, so I know nothing of the conditions necessary for the treatment. The literature in this field is old - 1960s - 1980s. Search the old literature for the technique if you are interested.

Gary Lee Gilmore thank you very much for your response.

I would like to pre-expose the DCs to the surface proteins of the cells of interest prior to live cell addition. Thank you for the suggestion of bromelain, I will look into this further.

I really appreciate your advice.

Best,

Anna

Ah. What you are interested in is antigen loading of DCs. I know a little bit about this subject. The primary work in this field was done by Ralph Steinman. A literature search of him would give you lots of information on this topic.

A couple of key points - there are two types of DCs - lymphoid and myeloid. From Steinman's work, one class of DCs [probably the lymphoid] derive from specific cells found in the epidermis called Langerhans cells. If the Langerhans cells encounter antigen while in the epidermis, they ingest it, change morphology, and migrate to the lymph nodes, where they present antigen to T lymphocytes. So this group of cells need to encounter antigen in the epidermis in order to present it. I do not know if they can process antigens for presentation in the DC form.

This is in contrast to the myeloid DCs, which derive from macrophages or monocytes. These cells are widely distrubed throughout the body. They actively phagocytose antigen and process it for presention to T lymphocytes. They can migrate to the lymph nodes and assume DC morphology. I suspect these cells can be antigen-loaded at any time.

The reason I know this is from my work making allogeneic chimeric mice - basically a C57BL6 immune system in a BALB/c [actually CB.17scid] enviroment. The Langerhan cells were BALB -derived, and the DCs in the lymph nodes were predominantly B6-derived [~67%, if I recall correctly].

Gary Lee Gilmore that's great, thanks very much