Our team is currently using footpad swelling (measured with a caliper) to determine the disease progression of L. major in infected mice. We also monitor wheter there is necrosis on the footpad or not. However, we would like to compare the level of necrosis between the individuals.
I was thinking about using the HMGB1 protein as a marker for necrosis (detection with PE anti-HMGB1 antibodies in FACS), since necrosis cells release this protein. However, I am not sure whether this would be a good marker or not, since I have read that recent studies have shown that also apoptotic cells can release this?
Hello, HMGB1 is passively released by cells that have died in a traumatic, way necrosis. Several receptors for HMGB1 exist, and upon binding HMGB1 they alert leukocytes to extravasate from the blood into the affected tissue, trigger adaptive immunity and promote the migration and proliferation of cells to repair the damaged tissue. Significantly, apoptotic cells modify their chromatin so as to bind HMGB1, which is also can happen.
Hope this info will help you
Good luck!
To do regular Flow cytometry, the marker needs to be in your cells of interest (either on the surface or intracellularly). HMGB1 is released from the cells when they die and I don’t think this release is specific to necrosis. Even if HMGB1 was a specific marker for necrosis, to measure something that is released from the cells is going to be problematic esp. using regular Flow Cytometry.
If you can find a way to lyse the foot pad sample without losing the released HMGB1, you could in theory measure it with bead based Flow Cytometry (provided you can find an HMGB1 CBA kit). Your best bet would be to do histology or do Flow using one of the apoptosis/necrosis kits (reagents binding to cell surface/intracellular markers) that are available commercially.
- Badges
- Science topic
- Similar topics
- Biological Science
- Immunology
- Antibodies