|Year : 2020 | Volume
| Issue : 2 | Page : 94-98
The relevance of complement C4d staining in renal allograft biopsies
Department of Pathology, ESIC Model Hospital, Gurugram, Haryana, India
|Date of Submission||04-Nov-2019|
|Date of Acceptance||15-Mar-2020|
|Date of Web Publication||06-Jul-2020|
Dr. Anju Khairwa
Department of Pathology, ESIC Model Hospital, Gurugram, Haryana
Source of Support: None, Conflict of Interest: None
In renal transplant, the allograft is affected by many triggering agents such as innate and adaptive immune mechanisms, either mediated by macrophages and lymphocytes, or by soluble components antibodies and the complement system, which can ultimately lead to graft rejection. Antibody-mediated rejection (AMR) is a predominant cause of allograft failure. Donor-specific antibodies, mostly reactive to human leukocyte antigen antigens, are now considered by pathologists and clinicians as a significant cause of early and late graft dysfunction and failure. Complement 4d (C4d) is a fragment of the classical complement pathway (that is a part of component C4), which is activated by antigen-antibody complexes. The diagnosis of AMR improves by detection of the complement fragment C4d in renal biopsy, and it has included for diagnosis of AMR in the year 2003. There is more development about C4d after inclusion in the diagnostic criteria of AMR. This review aims to evaluate pathogenesis and current relevance of C4d in AMR.
Keywords: Antibody-mediated rejection, complement 4d, donor-specific antibodies, positive
|How to cite this article:|
Khairwa A. The relevance of complement C4d staining in renal allograft biopsies. Indian J Transplant 2020;14:94-8
| Introduction|| |
Post kidney transplant, the allograft is affected by many stimulating agents such as antigen- antibody-mediated immunocomplexes and cellular immunity mediated by various cells (macrophages and lymphocytes), which could lead to graft rejection. Based on etiology, the Banff criteria divided the rejection broadly in antibody-mediated rejection (AMR), and cell-mediated rejection. AMR is a predominant cause of allograft failure. The clinicians and pathologists now consider antibodies against human leukocyte antigen (HLA) antigens defined as donor-specific antibodies (DSA) a remarkable cause of early and late graft dysfunction.
Complement 4d (C4d) (that is part of component C4) is a fragment of the classical complement pathway, which is activated by immune complexes. C4 is cleaved by C1s into C4a and C4b, exposing a sulfhydryl group. The active sulfhydryl group of C4b rapidly forms an amide or ester bond with nearby molecules containing hydroxyl or amino groups and forms C4d. C4d makes covalent bond with the tissue. It stays at the site of complement activation for a longer time, in comparison to other complement pieces. The diagnosis of AMR is improved by identification of the complement pieces C4d in renal biopsy, and it has been introduced first under diagnostic criteria of AMR in year 2003. There is more development about C4d after inclusion in diagnostic tests of AMR. This review aims to evaluate pathogenesis and current relevance of C4d in AMR.
| Pathogenesis of Complement 4d (Complement System and Complement 4d)|| |
Complement is an important component of innate and adaptive immunity in our body. Complement is the leading effector constituent of adaptive humoral immunity. There are three different pathways of complement activation and their activator, as shown in [Flow Diagram 1]. This flow diagram [Figure 1] is showing complement activation and C4d deposition in renal allograft biopsy.
|Figure 1: Complement 4d deposition in peritubular capillaries (diffuse, intense linear circumferential peritubular capillaries staining of complement 4d on IHC, ×200)|
Click here to view
In the classical pathway of complement kick over of C1 (which composed of C1q, C1r, and C1s) is begun by an interplay of C1q with immunoglobulin bound to epitopes on the graft endothelium. C1s splits C4 into C4a and C4b and react with exposing a sulfhydryl group. The ester or amide bond forms with interacting between sulfhydryl group of C4b and nearby molecules containing hydroxyl or amino groups in the tissue. C3 convertase (C4bC2a) of classical pathway is formed by combination C4b with enzymatically loose pieces of C2a. C4bC2a (C3 convertase) splits C3 into C3a and C3b. C3b has an active sulfhydryl group which is dative bound and settles in the immediate locality and forms C5 convertase (C4bC2aC3b). C5 convertase split C5 into C5a and C5b. Further, membrane attack complex (membrane-bound C5b-9) formed by the addition of C5b component of C5 convertase, that destroys cells. The lectin pathway of complement activates by mannan-binding lectin (MBL) and H-ficolin or L-ficolin. The MBL attach to the suitable carbohydrate on apoptotic cells or pathogens. MBL, H-ficolin, and is L-ficolin are homologous to C1q and fibrinogen, which are also activate C4 by their associated serine proteases, mannose-associated serine protease 1 (MASP-1 and MASP-2) (analogs to C1r and C1s). The C-reactive protein (CRP) also activates complement (C4) pathway. The CRP binds to carbohydrate and choline phosphate along with C1q. The complement components C3a and C5a (call as anaphylatoxin) formed during this pathway.
| Clinical Relevance of Complement 4d|| |
The conditions where complement 4d is positive
Following conditions are showing C4d positivity:,
- Acute AMR
- Chronic antibody-mediated rejection
- A, B, O blood group ABO incompatible grafts
The state where complement 4d is negative
The following conditions are showing C4d negativity:,,,,
- T-cell-mediated rejection (TCMR)
- A technical error like a type of fixatives, immunofluorescence (IF) versus immunohistochemistry (IHC)
- Fc receptor (FCR) on NK cells (FcRIIA) mediated rejection
- Antibodies unable to fix the complement
- Complement independent pathways of endothelial activation
- C4d deposition is a very low in quantity for the identification limits of IF/IHC
- Alloantibodies can direct endothelium injury to interact with major histocompatibility complex (MHC)
- Increased expression of endothelial transcripts causing endothelium injury.
The C4d-negative AMR occur due to alloantibodies to MHC molecules on endothelial cells, elicit strong responses that include proliferation and activation of intracellular signaling pathways, which leads to endothelium injury. The C4d-negative AMR also seen with high renal endothelial transcript expression in patients with alloantibody and is an indicator of active antibody-mediated allograft damage and poor graft outcome.
| Antibody-Mediated Rejection|| |
The AMR harms allograft kidney, chiefly in more sensitized patients. AMR accounts for up to 30% of all posttransplant rejection events and 20%–30% graft loss at 1 year in untreated cases. AMR can occur in three forms, namely hyperacute, acute, and chronic rejection. The presensitization (blood transfusion, pregnancy, and prior transplant) is a primary risk factor for hyperacute and acute rejection. Acute AMR is most commonly occurs due to expose of MHC Class I and II HLA antigens. Whereas, chronic AMR is largely associated with Class II DSA.
Besides, other antigens which are responsible for AMR included MHC antigen (polymorphic) MICA (MHC Class I-related chain A) and ABO blood group antigens., Pathological findings of AMR- grossly kidneys become swollen due to interstitial edema and hemorrhage. Microscopic findings of AMR- glomerulitis, peritubular capillaritis, fibrinoid necrosis of arteries, and acute tubular injury. Other diagnostic features of AMR requires the simultaneous presence of DSA and C4d deposition in peritubular capillaries (PTCs).
Hyperacute AMR is defined as rejection occurs within minutes or 24 h after transplantation, mostly due to preexisting DSA present in high titres present in a patient. Grossly, the graft rapidly becomes cyanotic and flaccid. Histomorphology characterized by arteritis, interstitial edema, and extensive cortical necrosis. In acute AMR, C4d is deposited in PTC and glomerular capillaries. In chronic AMR, histomorphology characterized by transplant glomerulopathy (TG) in renal biopsies. The TG specified as capillary basement membrane duplication or splitting and mesangial expansion of glomeruli, tubular atrophy and interstitial fibrosis, and fibrous intimal thickening in arteries. Sometimes, electron microscopy also showed peritubular capillary basement membrane multilayering., The effector mechanisms of AMR (1) Direct effects of antibody to MHC, (2) Complement fixation, and (3) Cellular FCRs., The DSA produced by plasma cells bind to the endothelium of donor PTC and glomerular capillaries and start the pathological progression of AMR. The C1q is a component of the complement of the classical pathway which ties to the endothelium-binding DSA, finally leading to graft injury and dysfunction. Diagnosis of acute AMR based on the identification of DSA in plasma, C4 complement component (C4d) in PTC, structural corroboration of AMR, such as kidney injury and graft dysfunction., Mixed cellular and humoral rejection are very hard to examine because it mostly occurs after months and years posttransplantation, and biopsy may show acute and chronic lesion. Here, it is tough to prove that cellular or humoral immune system responsible for the extracellular injury. There suggested that AMR is the chief cause of graft damage, which positive IHC for C4d in PTC in the combination of high levels of DSA in the serum than cellular rejection., If there is minimum or absent of c4d positivity in PTC and absent of DSA in plasma less likely AMR. Some studies had shown by utilizing microarray investigation showed excessive NK cells in biopsies of patients with acute (in the form of PTCs, glomerulitis) and chronic AMR (in the form of TG).
| Transplant Glomerulopathy|| |
TG is a morphological change that is mostly seen in humoral rejection and related to immunological etiology., TG is a variant of chronic AMR and characterized by deposition of C4d. It proposed that TG may be one manifestation of humoral rejection of graft injury., Morphologically, TG consists of diffuse multilayering or duplication of the glomerular basement membrane (GBM). Their lack distinct de novo or repetitive glomerular lesion or evidence of thrombotic microangiopathies (TMA). GBM duplication best seen jones methenamine silver and periodic acid-Schiff stain. In TG double counter of GBM occurs due to accumulation of fluff such as material and mesangial cells are get deposited at subendothelial space.
IF and IHC show C4d deposition along the GBM presented in a minority of cases (6% in one series).
| Transplant Glomerulopathy Also Associated Multilayering of Peritubular Capillarie|| |
Ultra-structure features of TG consist of cytoplasmic vacuolation, broadening of subendothelial space, with the existence of electron-dense floccular material, probably due to stiffening of the lamina rara interna.
Interstitial fibrosis and tubular atrophy existed along with TG. Arterial intimal fibrosis new-onset, with leukocytic infiltration in intima and absence of internal elastic lamellae, favors chronic rejection with TG. TG was connected with the diagnosis of AMR, in the existence of either peritubular capillaritis or glomerulitis or both as well as with C4d expulsion in PTCs.
| Complement 4d detection in Renal Allograft Biopsies|| |
Two techniques are mostly used for identification of C4d IHC and IF in allograft biopsies of the kidney. IF technique applied to frozen sections whereas IHC technique used to paraffin-embedded tissue used routinely in various laboratories. In acute and chronic AMR-positive C4d stain with IF technique is described as “widespread, strong linear circumferential PTC staining in cortex or medulla, excluding scar or necrotic areas,” according to a consensus at 2003 Banff Conference. In IHC, C4d staining is crisp, linear, continuous, diffuse, and lying around in the PTC wall, while the strength typically is slighter and changeable, but it may have a finely granular pattern in high power. The scoring of C4d is significant by IF on frozen section ie C4d2 or C4d3 and by IHC C4d > 0 on paraffin sections. The circulating DSA are most specifically associated with C4d deposition in PTC and its interactivity with endothelial cells in the graft.
| Current Status of Complement 4d|| |
Devadass et al. reported C4d is not very sensitive marker of AMR, as it originally thought. Histologically, moderate to marked microvascular inflammation (MVI) correlated with dispersing C4d positive cases in contrast to focal C4d-positive cases.
C4d is significant staining in 1% or more of the PTC for formalin/IHC-IP, or 10% or more for frozen/frequency IF. Interobserver and interinstitutional reproducibility for C4d IHC staining in renal allografts was poor but improved with a binary scoring system (positive/negative).
Currently, C4d has become an essential investigation for AMR according to Banff update 2017. In this update, both C4d and validated transcripts/classifiers/molecular marker can serve as potential alternatives and complements to DSAs in the diagnosis of ABMR.
| Complement 4d negative Antibody-Mediated Rejection|| |
There are many causes for C4d-negative AMR, as mentioned above. The C4d-negative AMR was first demonstrated Feucht et al., and Edmonton group. The C4d-negative AMR is included in Banff 2013 classification. The morphology of C4d-positive and C4d-negative AMR having following similar features: (1) varying degrees of glomerulitis and peritubular capillaritis, (2) frequent TCMR, (3) both may occur early or late posttransplantation. However, C4d-negative AMR morphologically have higher intrarenal endothelial gene expression, alloantibodies expression, poor graft outcomes, usually occur after 1-year posttransplantation, and associated acute on chronic AMR.
There is another entity of AMR DSA negative AMR, which characterized negative DSA/lack of DSA detection, moderate MVI with (g + ptc) scores of ≥2) as per Banff 2013 with or without C4d positivity.
The accommodation is defined as C4d deposition in PTCs in the absence of active rejection with or without DSA positivity, mostly seen in ABO-incompatible graft transplantation. The TMA (thrombotic microangiopathy) is a complication of AMR in the absence of other causes characterized with fibrin thrombi, fragmented red blood cells, mesangiolysis, and muco-intimal thickening and injury of small vessels. According Banff diagnostic criteria, C4d staining in PTCs (at least 10% PTC positive is considered as significant) by IHC on paraffin sections Banff score C4d > 0 is significant. In contrast, by IFs on frozen sections Banff scores C4d2 or C4d3 is considered significant., Thus, it is indicated that C4d detection by IHC more specific and C4d detection by IF is more sensitive. The IHC method for C4d detection feasible in formalin-fixed, paraffin-embedded tissue, it is used when frozen sections facility not available, IHC also useful when small biopsy tissue or tissue not available for frozen sections. The disadvantage of C4d staining by IHC are lower sensibility, nonspecific background staining, more costly and more time consuming, and it needs external controls. IF method for C4d requires extra tissue and frozen sections facility.
| Conclusion|| |
The C4d-positive staining in PTC is a diagnostic marker as well as a prognostic marker for allograft. According to various studies worldwide, C4d appears to be a less sensitive marker than initially thought. The C4d as a biomarker has following PROS (1) provoked an enormous amount of insight in the diagnosis of allograft rejection, (2) core diagnostic tool to identify AMR, and (3) used for vast amount of research into the deposition patterns of C4d in different clinical settings such as pregnancy, thrombotic complications and CONS as follows: (1) difficulties of interpreting focal staining patterns, (2) relatively low sensitivity of C4d as a marker for AMR in late renal allograft biopsies, and (3) its lack of utility as a marker for antibody-mediated injury in biopsies of ABO-incompatible allografts).
However, in which cases C4d not helpful in diagnosis whereas molecular studies have furnished perceptiveness evocative of a complement-independent form of AMR or C4d-negative AMR. According to us, the detection of AMR should be best reported based on morphological features such as tubulointerstitial, vascular, and glomerular histological changes, with a piece of legislation to the presence or absence of C4d. Despite all pitfalls C4d is excellent marker for AMR.
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Conflicts of interest
There are no conflicts of interest.
| References|| |
Nankivell BJ, Alexander SI. Rejection of the kidney allograft. N
Engl J Med 2010;363:1451-62.
Mengel M, Sis B, Haas M, Colvin RB, Halloran PF, Racusen LC, et al
. Banff 2011 Meeting report: New concepts in antibody-mediated rejection. Am J Transplant 2012;12:563-70.
Tait BD, Süsal C, Gebel HM, Nickerson PW, Zachary AA, Claas FH, et al
. Consensus guidelines on the testing and clinical management issues associated with HLA and non-HLA antibodies in transplantation. Transplantation 2013;95:19-47.
Monsinjon T, Gasque P, Chan P, Ischenko A, Brady JJ, Fontaine MC. Regulation by complement C3a and C5a anaphylatoxins of cytokine production in human umbilical vein endothelial cells. FASEB J 2003;17:1003-14.
Chakravarti DN, Campbell RD, Porter RR. The chemical structure of the C4d fragment of the human complement component C4. Mol Immunol 1987;24:1187-97.
Racusen LC, Colvin RB, Solez K, Mihatsch MJ, Halloran PF, Campbell PM, et al
. Antibody-mediated rejection criteria-An addition to the Banff 97 classification of renal allograft rejection. Am J Transplant 2003;3:708-14.
Magil AB, Tinckam K. Monocytes and peritubular capillary C4d deposition in acute renal allograft rejection. Kidney Int 2003;63:1888-93.
Sun Q, Cheng Z, Cheng D, Chen J, Ji S, Wen J, et al
. De novo
development of circulating anti-endothelial cell antibodies rather than pre-existing antibodies is associated with post-transplant allograft rejection. Kidney Int 2011;79:655-62.
Kilgore KS, Schmid E, Shanley TP, Flory CM, Maheswari V, Tramontini NL, et al
. Sublytic concentrations of the membrane attack complex of complement induce endothelial interleukin-8 and monocyte chemoattractant protein-1 through nuclear factor-kappa B activation. Am J Pathol 1997;150:2019-31.
Orandi BJ, Alachkar N, Kraus ES, et al.
Presentation and Outcomes of C4d-Negative Antibody-Mediated Rejection After Kidney Transplantation. Am J Transplant. 2016;16:213-20. doi:10.1111/ajt.13434.
Haas M, Sis B, Racusen LC, Solez K, Glotz D, Colvin RB, et al
. Banff 2013 meeting report: Inclusion of c4d-negative antibody-mediated rejection and antibody-associated arterial lesions. Am J Transplant 2014;14:272-83.
Bian H, Reed EF. Alloantibody-mediated class I signal transduction in endothelial cells and smooth muscle cells: Enhancement by IFN-gamma and TNF-alpha. J Immunol 1999;163:1010-8.
Sis B, Jhangri GS, Bunnag S, Allanach K, Kaplan B, Halloran PF. Endothelial gene expression in kidney transplants with alloantibody indicates antibody-mediated damage despite lack of C4d staining. Am J Transplant 2009;9:2312-23.
de Kort H, Willicombe M, Brookes P, Dominy KM, Santos-Nunez E, Galliford JW, et al
. Microcirculation inflammation associates with outcome in renal transplant patients with de novo
donor-specific antibodies. Am J Transplant 2013;13:485-92.
Puttarajappa C, Shapiro R, Tan HP. Antibody-mediated rejection in kidney transplantation: A review. J Transplant 2012;2012:193724.
Abbas AK, Lichtman AH, Pober JS. Transplantation immunology. In: Cellular and Molecular Immunology. 4th
ed. Philadelphia: W.B. Saunders Co; 2000. p. 369-85.
Scornik JC, LeFor WM, Cicciarelli JC, Brunson ME, Bogaard T, Howard RJ, et al
. Hyperacute and acute kidney graft rejection due to antibodies against B cells. Transplantation 1992;54:61-4.
Issa N, Cosio FG, Gloor JM, Sethi S, Dean PG, Moore SB, et al
. Transplant glomerulopathy: Risk and prognosis related to anti-human leukocyte antigen class II antibody levels. Transplantation 2008;86:681-5.
Li L, Chen A, Chaudhuri A, Kambham N, Sigdel T, Chen R, et al
. Compartmental localization and clinical relevance of MICA antibodies after renal transplantation. Transplantation 2010;89:312-9.
Montgomery RA, Lonze BE, Jackson AM. Using donor exchange paradigms with desensitization to enhance transplant rates among highly sensitized patients. Curr Opin Organ Transplant 2011;16:439-43.
Porter KA, Marchioro TL, Starzl TE. Pathological changes in 37 human renal homotransplants treated with immunosuppressive drugs. Br J Urol 1965;37:250-73.
Nickeleit V. The pathology of kidney transplantation. In: Ruiz P, editors. Transplantation Pathology. New York: Cambridge University Press; 2009. p. 45-110.
Nickeleit V, Klimkait T, Binet IF, Dalquen P, Del Zenero V, Thiel G, et al
. Testing for polyomavirus type BK DNA in plasma to identify renal-allograft recipients with viral nephropathy. N
Engl J Med 2000;342:1309-15.
Kohler TR, Tilney NL. Microangiopathic hemolytic anemia associated with hyperacute rejection of a kidney allograft. Transplant Proc 1982;14:444-7.
Regele H, Böhmig GA, Habicht A, Gollowitzer D, Schillinger M, Rockenschaub S, et al
. Capillary deposition of complement split product C4d in renal allografts is associated with basement membrane injury in peritubular and glomerular capillaries: A contribution of humoral immunity to chronic allograft rejection. J Am Soc Nephrol 2002;13:2371-80.
Zhang X, Reed EF. Effect of antibodies on endothelium. Am J Transplant 2009;9:2459-65.
Rotman S, Collins AB, Colvin RB. C4d deposition in allografts: Current concepts and interpretation. Transplant Rev 2005;19:65-77.
Yabu JM, Higgins JP, Chen G, Sequeira F, Busque S, Tyan DB. C1q-fixing human leukocyte antigen antibodies are specific for predicting transplant glomerulopathy and late graft failure after kidney transplantation. Transplantation 2011;91:342-7.
Klein J, Sato A. The HLA system. First of two parts. N
Engl J Med 2000;343:702-9.
Stegall MD, Chedid MF, Cornell LD. The role of complement in antibody-mediated rejection in kidney transplantation. Nat Rev Nephrol 2012;8:670-8.
Everly MJ, Everly JJ, Susskind B, Brailey P, Arend LJ, Alloway RR, et al
. Bortezomib provides effective therapy for antibody- and cell-mediated acute rejection. Transplantation 2008;86:1754-61.
Hidalgo LG, Sellares J, Sis B, Mengel M, Chang J, Halloran PF. Interpreting NK cell transcripts versus T cell transcripts in renal transplant biopsies. Am J Transplant 2012;12:1180-91.
Gloor JM, Sethi S, Stegall MD, Park WD, Moore SB, DeGoey S, et al
. Transplant glomerulopathy: Subclinical incidence and association with alloantibody. Am J Transplant 2007;7:2124-32.
Fotheringham J, Angel CA, McKane W. Transplant glomerulopathy: Morphology, associations and mechanism. Nephron Clin Pract 2009;113:c1-7.
Zollinger HU, Moppert J, Thiel G, Rohr HP. Morphology and pathogenesis of glomerulopathy in cadaver kidney allografts treated with antilymphocyte globulin. Curr Top Pathol 1973;57:1-48.
Habib R, Zurowska A, Hinglais N, Gubler MC, Antignac C, Niaudet P, et al
. A specific glomerular lesion of the graft: Allograft glomerulopathy. Kidney Int Suppl 1993;42:S104-11.
Yilmaz A, Miskulin D, Gedeon I, Burama A, YilmazS, Supanj F, et al.
Peritubular capillary basement membrane reduplication in allografts and native kidney disease. Transplantation 2001;71:1390-3.
Wavamunno MD, O'Connell PJ, Vitalone M, Fung CL, Allen RD, Chapman JR, et al
. Transplant glomerulopathy: Ultrastructural abnormalities occur early in longitudinal analysis of protocol biopsies. Am J Transplant 2007;7:2757-68.
Haas M. C4d-negative antibody-mediated rejection in renal allografts: Evidence for its existence and effect on graft survival. Clin Nephrol 2011;75:271-8.
Steinmetz OM, Panzer U, Kneissler U, Harendza S, Lipp M, Helmchen U, et al
. BCA-1/CXCL13 expression is associated with CXCR5-positive B-cell cluster formation in acute renal transplant rejection. Kidney Int 2005;67:1616-21.
Mannon RB, Matas AJ, Grande J, Leduc R, Connett J, Kasiske B, et al
. Inflammation in areas of tubular atrophy in kidney allograft biopsies: A potent predictor of allograft failure. Am J Transplant 2010;10:2066-73.
Devadass CW, Vanikar AV, Nigam LK, Kanodia KV, Patel RD, Vinay KS, et al
. Evaluation of renal allograft biopsies for graft dysfunction and relevance of C4d staining in antibody mediated rejection. J Clin Diagn Res 2016;10:EC11-5.
Couzi L, Perera R, Manook M, Barnett AN, Shaw O, Kessaris N, et al
. Incidence and outcome of C4d staining with tubulointerstitial inflammation in blood group-incompatible kidney transplantation. Transplantation 2015;99:1487-94.
Haas M, Loupy A, Lefaucheur C, Roufosse C, Glotz D, Seron D, et al
. The banff 2017 kidney meeting report: Revised diagnostic criteria for chronic active T cell-mediated rejection, antibody-mediated rejection, and prospects for integrative endpoints for next-generation clinical trials. Am J Transplant 2018;18:293-307.
Feucht HE, Schneeberger H, Hillebrand G, Burkhardt K, Weiss M, Riethmüller G, et al
. Capillary deposition of C4d complement fragment and early renal graft loss. Kidney Int 1993;43:1333-8.
Sis B, Halloran PF. Endothelial transcripts uncover a previously unknown phenotype: C4d-negative antibody-mediated rejection. Curr Opin Organ Transplant 2010;15:42-8.
Gupta A, Broin PÓ, Bao Y, Pullman J, Kamal L, Ajaimy M, et al
. Clinical and molecular significance of microvascular inflammation in transplant kidney biopsies. Kidney Int 2016;89:217-25.
Solez K, Colvin RB, Racusen LC, Haas M, Sis B, Mengel M, et al
. Banff 07 classification of renal allograft pathology: Updates and future directions. Am J Transplant 2008;8:753-60.
Loupy A, Haas M, Solez K, Racusen L, Glotz D, Seron D, et al
. The banff 2015 kidney meeting report: Current challenges in rejection classification and prospects for adopting molecular pathology. Am J Transplant 2017;17:28-41.
Roufosse C, Simmonds N, Clahsen-van Groningen M, Haas M, Henriksen KJ, Horsfield C, et al
. A 2018 reference guide to the banff classification of renal allograft pathology. Transplantation 2018;102:1795-814.
Santos A, Viana H, Galvão MJ, Carvalho F, Nolasco F. C4d detection in renal allograft biopsies: Immunohistochemistry vs immunofluorescence. Port J Nephrol Hypert 2012;26:272-7.
Cohen D, Colvin RB, Daha MR, Drachenberg CB, Haas M, Nickeleit V, et al
. Pros and cons for C4d as a biomarker. Kidney Int 2012;81:628-39.