Diagnostic Accuracy of Direct Immunofluorescence Test on Paraffin-Embedded Blocks in Comparison with Frozen Section Blocks in Renal Biopsies

Table 2

Protocol for immunofluorescence on paraffin-embedded renal biopsies.

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Cut formalin-fixed paraffin-embedded tissue at 3 μ thickness on poly-L-Lysine coated slides

Deparrafinize and rehydrate tissue sections

Immerse in tris EDTA pH 9 for 30 min at room temperature

Perform enzymatic digestion with proteinase K 1.25 mg/mL (Sigma–Aldrich, United States) at room temperature for 20 min

Stop digestion by immersing in tris EDTA at 4°C

Leave in tris EDTA for 40 min at 4°C

Rinse in PBS for 10 min

Apply FTTC conjugated polyclonal rabbit antibodies directed against IgG, IgM, IgA, and C3 incubated for 1 h in a moist chamber in the dark

Rinse with PBS

Mount in glycerine

Examine slides under a dark field immunofluorescence microscope

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PBS: Phosphate buffered saline; FTTC: Fluorescein isothiocyanate.