Glyoxal acid-free as a safer and reliable alternative to glutaraldehyde for renal ultrastructural pathology

Abstract Background Renal ultrastructural diagnosis still relies on glutaraldehyde fixation, but this fixative is toxic and has limited tissue penetration. We evaluated whether acid-free glyoxal can provide adequate preservation for renal transmission electron microscopy and whether diagnostically useful information can be recovered from paraffin-embedded tissue. Methods Normal human renal cortical tissue from nephrectomy specimens was fixed in acid-free glyoxal for 24, 48, or 72 hours and processed for transmission electron microscopy; tissue fixed in 2.5% buffered glutaraldehyde served as a control. Additional acid-free glyoxal-fixed paraffin-embedded samples were recovered and reprocessed. Preservation of podocyte foot processes, slit diaphragms, endothelial fenestrations, and the glomerular basement membrane were assessed. Conventional histology, automated immunohistochemistry, and exploratory immunofluorescence were also evaluated. Results Fixation for 24 hours preserved the main ultrastructural components of the glomerular filtration barrier, although the basement membrane appeared less compact than after glutaraldehyde fixation. Preservation improved markedly after 48 and 72 hours, with clear slit diaphragms, endothelial fenestrations, and balanced image contrast. Tissue recovered from paraffin blocks also yielded adequate ultrastructural detail for common diagnostic questions. Conventional histology was preserved, whereas automated immunohistochemical reactivity was reduced for the markers tested. Conclusions With appropriate fixation times, acid-free glyoxal provides good to excellent preservation of renal glomerular ultrastructure and allows recovery of diagnostically useful transmission electron microscopy findings from paraffin-embedded tissue, supporting its use as a safer and more versatile alternative to glutaraldehyde in renal diagnostic pathology.