Q: assay for an immune status in mice

[dyk2]

Hi,

 

I am looking for a generic and simple method to evaluate an immune status/potential of mice. I am not an Immunology person (my field is DNA repair), so any advice will be much appreciated. Basically, I want to see if immune system is activated/enhanced after low dose gamma-irradiation. This treatment leads to lower frequency of spontaneous tumors and longer latency times. But DNA repair assays failed to show any enhancement. I am therefore wondering if it is immune system that causes the effect. I am interested in a general immune status assessment rather that specific pathways. Thanks.

 

Dmitry

[jimmydasaint]

I take it you want to examine immune response in terms of blood and lymph antibodies and then examine celllular responses from cell 'munchers' e.g. macrophages.

 

There is a really simple system for measuring the antibody responses to the molecules that cause an immune response called antigens. This is called radioimunoprecipitation. Firstly, choose a particularly obvious marker from tumour cells. Next, radio-label it using easily purchased reagents (e.g. Bolton hunter reagent to label lysine amino acids on proteins or protein-containing antigens).

 

Next perform an antibody extraction using serum from the affected animals. concentrate it if necessary and then add whole serum to the radiolabelled antigen (cell surface marker). After incubation, separate antibodies from captured antigens using SDS or mercaptoethanol and run an SDS-PAGE gel of all the antigens identified. Use a lane of the gel for common molecular weight markers and also include a negative control and a pre-treatment control. Any extra antibody response can be picked up quite nicely.

 

If you want to examine non-cellular responses, this is a bit more complicated. tell me if this advice makes any sense because I suspect you may be looking at purely non-specific responses.

[ABD]

Hi All.

 

I have one question for the forum.

Is there any body experienced in differentiating apoptosis and necrosis by using microscope and flow cytometry? Please,I need some explanation about it.

 

Thanks