Compartment-specific flow cytometry for the analysis of TNF-mediated recruitment and activation of glomerular leukocytes in murine kidneys

Cytokines of the TNF superfamily, particularly TNF itself, are important mediators of inflammatory leukocyte recruitment and activation in parenchymal organs. In inflammatory kidney diseases, leukocytes accumulate in glomeruli and the tubulointerstitium, leading to glomerulonephritis and tubulointerstitial nephritis, respectively. In particular, glomeruli can be the target of organ-threatening leukocyte-mediated inflammation. As microvasculatures of the glomerulus and the tubulointerstitium differ markedly in their structural and functional properties, recruitment and subsequent activation of leukocytes to these sites occur via distinct mechanisms. To understand the pathways and mediators of leukocyte-driven inflammation in the kidney it is therefore essential to analyze glomerular and tubulointerstitial leukocyte recruitment in a compartment-specific way. The protocol presented here describes an easy and rapid technique that allows compartment-specific quantitation and qualitative analysis of leukocytes present in glomeruli and tubulointerstitial tissue by flow cytometry after separation of these tissue compartments.