Hello,
I would like to perform immunofluorescent staining of IgG and C3 (immune complexes) in the kidneys of lupus mice. I was wondering, whether perfusion of the mice is required for this, before kidney harvest. Would the background in non-perfused kidneys interfere with the fluorescent signals of the immune complexes, or would perfusion not make a significant difference, since maybe the immune complexes are deeper in the renal tissue?
In case this is dependent on the fixation strategy, we want to fix the kidneys with either 4% PFA or acetone.
I would greatly appreciate any feedback!
Erythrocytes and hemoglobin from lysed erythrocytes exhibit strong autofluorescence, esp., in the green and red channels (Fluorescein and alike). If you're making thin sections, you might get away without perfusion.