Optimal Cutoff Point for Immunoperoxidase Detection of C4d in the Renal Allograft: Results From a Multicenter Study

Abstract

Background Although C4d deposition in peritubular capillaries has been identified as a strong risk factor for subsequent renal allograft loss, the optimal cut-off for the fraction of peritubular capillaries needed to establish a positive stain in formalin-fixed paraffin-embedded material has not been systematically defined. The objective of this study was to establish the threshold for positive staining that best predicts renal outcome in renal biopsies in a multicenter study in which local and central pathology were compared. Methods Unstained renal biopsy slides were obtained from 296 patients. The percentage of peritubular capillaries staining positively for C4d was detected by immunoperoxidase staining. Results The percentage C4d deposition ranged from 0% to 90% with 44% (129/296) having a positive percentage of C4d staining. The median for positive cases was 25%. Local C4d+ results were reported qualitatively, with 28% recorded as positive for C4d. Using a centrally-determined cut-off of 10%, tests for agreement of local and central C4d staining were fair (Kappa 0.40, 95% CI 0.29-0.51). Raising the centrally-determined cut-off to 25% or 50% did not change the Kappa values (0.44 and 0.41, respectively). By Cox proportional hazards model, C4d positivity (centrally-determined assessment) using a cut-off of 10% was the strongest predictor of time to graft loss (HR 2.66, 95% CI [1.68, 4.21]). Centrally-determined C4d positivity correlated with Banff scores indicative of acute inflammation, but not with scores indicative of fibrosis/atrophy or transplant glomerulopathy. Conclusions Our findings indicate that C4d positivity, defined as ≥10% by immunoperoxidase, is a strong predictor of graft loss.

A systematic review of the role of C4d in the diagnosis of acute antibody-mediated rejection

“…Five companion reports were used for supplementary data. 9,10,[32][33][34] Figure 1 presents the flow diagram of search results. A total of 3485 indication and 868 surveillance biopsies were included.…”

Sequential analysis of donor-specific antibodies and pathological findings in acute antibody-mediated rejection in a rat renal transplantation model

“…However, the incidence of C4d-positive AMR in grafts varies depending on the proportion of sensitized patients, as has been shown in previous reports. [27][28][29][30][31] The reason for using C4d, the split product of complement component 4 (C4), as a marker for AMR is based on its role in the complement cascade. 1 There are three main pathways for complement activation: the classical, lectin, and alternative pathways.…”

Clinical and pathological correlations of C4d immunostaining and its infl uence on the outcome of kidney transplant recipients

“…The percentage of C4d staining that accurately predicts kidney transplantation outcome is still debated, and even a cutoff of 10% for C4d positivity was reported to be a strong predictor of renal graft loss. 27 Several studies had compared IF and IHC on frozen and paraffin sections for C4d identification and grading in kidney graft biopsies. Some of them showed higher accuracy for frozen-IF with monoclonal antibody in relation to paraffin-IHC, 14,15,28 but other authors reported an acceptable sensitivity (87.5%) and specificity (98%) of paraffin-IHC with anti-C4d polyclonal antibody.…”