Detection of BK Virus in Renal Allograft Biopsies by RNA In Situ Hybridization RNAscope<sup>®</sup> Assay
November 08, 2019 | 10:00 AM - 12:00 PM
Detection of BK Virus in Renal Allograft Biopsies by RNA In Situ Hybridization RNAscope® Assay
Transplantation: Clinical - Post-Transplant Complications
November 08, 2019 | Location: Exhibit Hall, Walter E. Washington Convention Center
Abstract Time: 10:00 AM - 12:00 PM
- 1902 Transplantation: Clinical
- Costigliolo, Francesca, Johns Hopkins University, Baltimore, Maryland, United States
- Lombardo, Kara Anne, Johns Hopkins Medical Institutions, Baltimore, Maryland, United States
- Arend, Lois J., Johns Hopkins Hospital, Baltimore, Maryland, United States
- Rosenberg, Avi Z., Johns Hopkins University, Baltimore, Maryland, United States
- Matoso, Andres, Johns Hopkins, Baltimore, Maryland, United States
- Carter-Monroe, Naima, Veterans Administration Maryland Health Care System, Baltimore, Maryland, United States
- Bagnasco, S.M., The Johns Hopkins School of Medicine, Baltimore, Maryland, United States
Kara Anne Lombardo,
Lois J. Arend,
Avi Z. Rosenberg,
BK polyomavirus associated nephropathy (BKpyVAN) remains a cause of graft loss in kidney transplant recipients on immunosuppressive therapy, and its diagnosis relies on identification of BK virus in the renal allograft biopsy based on positive immunohistochemical stain for the viral SV40 large-T antigen. Real time PCR (qPCR) and in situ hybridization (ISH) for BKv DNA can also be used to identify BKv in kidney tissue. Aim of this study was to evaluate RNAscope®, a novel-next generation technique for in situ hybridization with probes designed to increase the signal-to noise ratio to visualize RNA transcripts, for the detection of BKv RNA in allograft biopsies.
SV40 IHC stain (Santa Cruz, Ventana System) and RNAscope® ISH (following Advanced Cell Diagnostics manufacturer protocol), were performed on serial paraffin embedded tissue sections of kidney allograft biopsies. The number of tubules showing positive SV40 nuclear staining was compared to the number of tubules showing positive RNAscope® ISH signal in each biopsy (paired, 2 sides t-test, linear regression, Pearson).
From 2010 to 2018, a diagnosis of BKpyVAN was made on 32 allograft biopsies from 30 renal transplant recipients (66% Caucasian, 58% male, average age at biopsy 54 ± 13 years). Median time of diagnosis post-transplant was 366 days (range 21-1577), median serum creatinine at diagnosis was 1.6 mg/dl (range 0.84-3.48 mg/dl), range BKv viremia levels: <201 – 1,270,000 DNA copies. We excluded 3 biopsies with equivocal SV40 stain, low BKv viremia (296 and 2830 DNA copies) and negative RNAscope® ISH. Three biopsies from 2 patients with high BKv viremia (>100,000 DNA copies), basophilic inclusion, negative SV40, showed positive staining by RNAscope® ISH in more than 50 tubules. In the remaining 26 biopsies with BKpyVAN there was no significant difference (P=0.207) in the average number of BKv-positive tubules detected by SV40 (26 ± 33) or RNAscope® ISH (21 ± 44), with good correlation between the two methods (r2=0.798, P<0001). Weak correlation was seen between the level of BK viremia and the number of positive tubules detected by either SV40 (r2=0.250, P=0.012) or RNAscope® ISH (r2=0.255, P=0.014).
Our data suggest that RNAscope® ISH may be comparable to SV40 for detection of BKv in renal allograft biopsies.