Kidney Biopsy Techniques to Differentiate Calcium Oxalate and Dihydroxyadenine Tubular Crystals

To the Editor:

One of the most challenging aspects of kidney pathology is the differential diagnosis of crystal deposition. Some crystals (calcium oxalate [CaOx] crystals and 2,8-dihydroxyadenine [DHA] crystals) show bright birefringence under polarized light; however, further differential diagnosis is difficult. We had earlier reported that CaOx crystals dissolve in Michel transport medium (MTM). Frozen sections of tissue that were exposed to MTM do not have birefringent CaOx crystals.¹ Herein, we report that DHA crystals do not dissolve in MTM, suggesting that incubation in MTM may be a diagnostic tool to differentiate between CaOx and DHA crystals in a kidney biopsy specimen.

A patient with kidney failure secondary to DHA deposition had early recurrence of the disease in a kidney allograft. Brown, CaOx-like (Fig 1A), brightly birefringent crystals were noted in the tubules in both formalin-fixed paraffin sections and frozen sections (Fig 1B and C). Of note, the biopsy core submitted for immunofluorescence was exposed to MTM for 24 hours (additional incubation in MTM did not affect the birefrigent crystals, Fig 1D). We incubated kidney biopsy cores (3 patients) with CaOx nephropathy, (Fig 1E), that had never been exposed to MTM, in MTM for 24 hours. In the sections of paraffin embedded tissue (Fig 1F) and sections of fresh frozen tissue sections (Fig 1G), numerous birefringent CaOx crystals were seen. After the 24-hour exposure to MTM, the birefringent crystals disappeared from the frozen sections in all 3 cases. However, incubation of biopsy cores in MTM is unpractical; therefore, we incubated unstained sections of frozen tissue that were not exposed to MTM before freezing from a case with CaOx crystals and sections of frozen tissue from the case with DHA crystals in MTM for 2, 8, and 24 hours. Incubation in MTM for 2, 8, and 24 hours resulted in the dissolution of CaOx crystals (Fig 1H) but not DHA crystals (Fig 1D).

Fig 1.

Histology of tubular crystal deposits. (A) Light microscopy images of 2,8-dihydroxyadenine crystal tubular deposits in a kidney biopsy specimen. Darker crystals are seen in the tubules (arrows). Acute tubular necrosis is seen in the proximal tubules. Hematoxylin and eosin (H&E) stain. Original magnification, ×200. (B) Birefringent 2,8-dihydroxyadenine crystals are seen in the sections of paraffin-embedded tissue. H&E stain, polarized light. Original magnification, ×100. (C) Birefringent 2,8-dihydroxyadenine crystals are seen in the sections of frozen tissue that were exposed to Michel transport medium (MTM) for 24 hours before cutting. H&E stain, polarized light. Original magnification, ×100. (D) Birefringent 2,8-dihydroxyadenine crystals are seen in the sections of frozen tissue that were exposed on the slide to MTM for 24 hours. H&E stain, polarized light. Original magnification, ×100. (E) Light microscopy images of calcium oxalate crystal tubular deposits in a kidney biopsy specimen. Translucent light-colored crystals are seen in the tubules (arrows). Acute tubular necrosis is seen in the proximal tubules. H&E stain. Original magnification, ×200. (F) Birefringent calcium oxalate crystals are seen in the sections of paraffin-embedded tissue. H&E stain, polarized light. Original magnification, ×100. (G) Birefringent calcium oxalate crystals are seen in the sections of frozen tissue that were not exposed to the MTM. H&E stain, polarized light. Original magnification, ×100. (H) No birefringent calcium oxalate crystals are present in the sections of frozen tissue after incubation of the slide in the MTM for 2 hours. H&E stain, polarized light. Original magnification, ×100.

We suggest that exposure to MTM may be helpful in patients with birefringent crystals in frozen tissue. If tissue was not exposed to MTM, then such birefringent crystals could be either CaOx or DHA. In such cases, incubation of unstained sections or entire biopsy cores in MTM will help in the differential diagnosis between CaOx and DHA crystals. If birefringent crystals are not seen after MTM incubation, then such crystals most likely are CaOx. However, if birefringent crystals are still seen in the tissue exposed to MTM, then such crystals could represent DHA deposition. Our data suggest that incubation in MTM even for 2 hours is sufficient to dissolve CaOx crystals from sections of frozen tissue.