Dear Graeme, You will need to dissociate the lymph node, or at least a part of it. This should be fairly easy to do using scalpels and gently pressing the tissue through a steel mesh (60-100 gauge) using the plunger from a syringe. Or use the Medicons tissue dissociator. Or add some dilute collagenase (Type 4 @ 0.2mg/ml) and dissociate at 37oC for about 15 minutes. Cells will then need to be fixed, either 70% cold methanol; or 1% paraformaldehyde (5 mins), wash, 70% methanol 30mins+. Then stain cells using FITC conjugated EMA-type antibodies, or cytokeratin (CAM5.2, CK8,18,19) or using two stage staining with a second stage FITC conjugated antibody. This will enable epithelial cells to be identified by flow cytometry. If you wish, also add 50ug/ml propidium iodide, stain overnight and then do dual analysis with the FITC Ab (FL1) and DNA (FL2 or 3) to measure DNA ploidy and cell cycle of the positive cells. Best wishes, John.
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