Validation of digital pathology imaging for primary histopathological diagnosis.

AIMS: Digital pathology (DP) offers advantages over glass slide microscopy (GS), but data demonstrating a statistically valid equivalent (i.e. non-inferior) performance of DP against GS are required to permit its use in diagnosis. The aim of this study is to provide evidence of non-inferiority. METHODS AND RESULTS: Seventeen pathologists re-reported 3017 cases by DP. Of these, 1009 were re-reported by the same pathologist, and 2008 by a different pathologist. Re-examination of 10 138 scanned slides (2.22 terabytes) produced 72 variances between GS and DP reports, including 21 clinically significant variances. Ground truth lay with GS in 12 cases and with DP in nine cases. These results are within the 95% confidence interval for existing intraobserver and interobserver variability, proving that DP is non-inferior to GS. In three cases, the digital platform was deemed to be responsible for the variance, including a gastric biopsy, where Helicobacter pylori only became visible on slides scanned at the ×60 setting, and a bronchial biopsy and penile biopsy, where dysplasia was reported on DP but was not present on GS. CONCLUSIONS: This is one of the largest studies proving that DP is equivalent to GS for the diagnosis of histopathology specimens. Error rates are similar in both platforms, although some problems e.g. detection of bacteria, are predictable.

Human leukocyte antigen antibody-incompatible renal transplantation: excellent medium-term outcomes with negative cytotoxic crossmatch.

BACKGROUND: Human leukocyte antigen (HLA) antibody-incompatible renal transplantation has been increasingly performed since 2000 but with few data on the medium-term outcomes. METHODS: Between 2003 and 2011, 84 patients received renal transplants with a pretreatment donor-specific antibody (DSA) level of more than 500 in a microbead assay. Seventeen patients had positive complement-dependent cytotoxic (CDC) crossmatch (XM), 44 had negative CDC XM and positive flow cytometric XM, and 23 had DSA detectable by microbead only. We also reviewed 28 patients with HLA antibodies but no DSA at transplant. DSAs were removed with plasmapheresis pretransplant, and patients did not routinely receive antithymocyte globulin posttransplant. RESULTS: Mean follow-up posttransplantation was 39.6 (range 2-91) months. Patient survival after the first year was 93.8%. Death-censored graft survival at 1, 3, and 5 years was 97.5%, 94.2%, and 80.4%, respectively, in all DSA+ve patients, worse at 5 years in the CDC+ve than in the CDC-ve/DSA+ve group at 45.6% and 88.6%, respectively (P<0.03). Five-year graft survival in the DSA-ve group was 82.1%. Rejection occurred in 53.1% of DSA+ve patients in the first year compared with 22% in the DSA-ve patients (P<0.003). CONCLUSIONS: HLA antibody-incompatible renal transplantation had a high success rate if the CDC XM was negative. Further work is required to predict which CDC+ve XM grafts will be successful and to treat slowly progressive graft damage because of DSA in the first few years after transplantation.

Double filtration plasmapheresis in antibody-incompatible kidney transplantation.

Double filtration plasmapheresis (DFPP) was used in preference to plasma exchange in our program of antibody-incompatible transplantation, to treat higher volumes of plasma. Forty-two patients had 259 sessions of DFPP, 201 pre-transplant and 58 post-transplant. At the first treatment session, the mean plasma volume treated was 3.81 L (range 3-6 L), 55.5 mL/kg (range 36.2-83.6 mL/kg). Serum IgG fell by mean 59.4% (SD 10.2%), and IgM by 69.3% (SD 16.1%). Nine patients did not require increases in plasma volumes treated, and six did not tolerate higher plasma volumes. In the remaining patients, the mean maximum plasma volume treated pre-transplant was 6.67 L (range 4-15 L), 96.1 mL/kg (range 60.2-208.9 mL/kg). The complement dependent cytotoxic crossmatch was positive in 14 cases pre-treatment, and remained positive in six (42.8%) cases. The flow cytometric crossmatch was positive in 29 cases pre-treatment, and in 21 (72.4%) after DFPP. Post-transplant, DFPP was ineffective at reducing donor specific antibody levels during periods of rapid donor specific antibody synthesis. Post-transplant, the one year graft survival rate was 94%, although there was a high rate of early rejection. In summary, DFPP enabled the treatment of plasma volumes that were almost double those that would have been feasible with plasma exchange. Despite this, most patients were transplanted with a positive crossmatch, and DFPP post-transplant was unable to control rising antibody levels.

The histological development of acute antibody-mediated rejection in HLA antibody-incompatible renal transplantation.

BACKGROUND: The aim of this study was to examine the development of acute antibody-mediated rejection in HLA antibody-incompatible renal transplantation in relation to the Banff 07 histological classification. METHODS: Renal biopsies were scored using the Banff 07 diagnostic criteria, and paraffin-embedded sections were stained with the pan-leucocyte marker CD45. RESULTS: Thirty-six patients had 72 renal biopsies. In biopsies performed 30 min after graft reperfusion, the mean number of CD45+ cells per glomerulus was higher than in control grafts (P < 0.04) and was associated with the donor-specific antibody (DSA) level at transplantation measured by microbeads (P < 0.01), and eight out of nine patients with greater than five CD45+ cells per glomerulus had early post-transplant rejection or oliguria, compared to 11 out of 20 with less than five cells per glomerulus (P < 0.01). In the first 10 days post-transplant, although peritubular capillary (PTC) leucocyte margination grade 3 and C4d deposition were specific for rejection, their sensitivities were low. PTC C4d staining was only seen in two out of 11 biopsies taken in the first 5 days after transplant, even in the presence of rejection, but was present in the majority of later biopsies with rejection. In biopsies stained for CD3, CD68 and CD20, it was notable that CD20+ cells were not seen during acute rejection, the infiltrates comprising CD3+ and CD68+ leucocytes. CONCLUSIONS: Glomerular margination of leucocytes occurred early after transplantation and was associated with DSA level and early graft dysfunction. The Banff 07 PTC margination scoring system was easy to apply, especially when CD45 staining was used, and PTC margination grade 3 was always associated with clinical rejection.

Rises and falls in donor-specific and third-party HLA antibody levels after antibody incompatible transplantation.

BACKGROUND: After human leukocyte antigen (HLA) antibody-incompatible transplantation, donor specific and third party HLA antibodies may be found, and their levels fall in a donor-specific manner during the first month. However, these changes have not been previously described in detail. METHODS: Donor-specific HLA antibody (DSA) and third-party HLA antibody (TPA) levels were measured using the microbead method in 44 presensitized patients who had renal transplantation. RESULTS: DSA+TPA fell in the first 4 days after transplantation, and greater falls in DSA indicated absorption by the graft. This occurred for class I (57.8% fall compared with 20.2% for TPA, P<0.0005), HLA DR (63.0% vs. 24.3%, P<0.0004), and for HLA DP/DQ/DRB3-4 (34% vs. 17.5%, P=0.014). Peak DSA levels occurred at a mean of 13 days posttransplant, and they were higher than pretreatment in 25 (57%) patients and lower in 19 (43%) patients (P=ns). The risk of rejection was associated with peak DSA levels; 15 of 25 (60%) patients with DSA at median fluorescence intensity (MFI) more than 7000U experienced rejection, compared with 4 of 7 (57%) patients with peak DSA MFI 2000 to 7000U, and 2 of 12 (17%) patients with peak DSA MFI less than 2000U (P<0.02). DSA levels subsequently fell in a donor specific manner compared to TPA. CONCLUSION: DSA levels may change markedly in the first month after antibody incompatible transplantation, and the risk of rejection was associated with higher pretreatment and peak levels.

Blood levels of donor-specific human leukocyte antigen antibodies after renal transplantation: resolution of rejection in the presence of circulating donor-specific antibody.

BACKGROUND: Accommodation to antibody is an important mechanism in successful ABO-incompatible transplantation, but its importance in human leukocyte antigen (HLA) antibody-incompatible transplantation is less clear, as sensitive techniques facilitating daily measurement of donor-specific HLA antibodies (DSAs) have only recently been developed. METHODS: We report 24 patients who had HLA antibody-incompatible kidney transplantation (21 living donors, 3 deceased), 21 of whom had pretransplant plasmapheresis. Eight had positive complement-dependent cytotoxic (CDC) crossmatch (XM) pretransplant plasmapheresis, nine had positive flow cytometric (FC) XM, and seven had DSA detectable by microbead analysis only. After transplant, DSA levels were monitored closely with microbead assays. RESULTS: Rejection occurred in five of eight (62.5%) CDC-positive cases, in three of nine (33%) FC-positive cases, and in two of seven (29%) of microbead-only cases at a median of 6.5 days after transplantation. Resolution occurred at a median of 15 days after transplantation, in 8 of 10 cases when the microbead level of DSA had median fluorescence intensity (MFI) >2000 U, in 6 of 10 when the microbead MFI >4000 U. In 8 of 10 cases, the microbead MFI at the time of resolution was greater than at the onset. DSA did not always cause clinical rejection. In five cases with a posttransplant DSA peaking at MFI >2000 U on microbead assay, rejection did not occur. CONCLUSION: These data suggest that the dominant method of successful transplantation was function of the transplant in the presence of circulating DSA, and they also define the period during which this occurred.