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Optical Clearing and Imaging of Immunolabeled Kidney Tissue
Published on: July 22, 2019
Comparison of C4d immunostaining methods in renal allograft biopsies
Megan L Troxell1, Lauren A Weintraub, John P Higgins
1Oregon Health and Science University, Department of Pathology, L471, 3181 SW Sam Jackson Park Road, Portland, OR 97239-3098, USA. mltroxell@yahoo.com
Insights
Immunohistochemical staining of formalin-fixed, paraffin-embedded renal allograft biopsies for C4d deposition is a reliable method for diagnosing humoral rejection. This technique shows high specificity and acceptable sensitivity compared to immunofluorescence on frozen tissues.
Area of Science:
- Nephrology
- Immunology
- Pathology
Background:
- C4d deposition in renal allografts is a key marker for humoral rejection.
- Current diagnostic methods primarily use immunofluorescence on frozen tissue sections.
- Limited data exists comparing different C4d staining methods, especially with formalin-fixed, paraffin-embedded tissues.
Purpose of the Study:
- To compare the diagnostic performance of two C4d immunostaining methods in renal allograft biopsies.
- To evaluate immunohistochemistry (IHC) on formalin-fixed, paraffin-embedded (FFPE) tissue against immunofluorescence (IF) on frozen tissue.
Main Methods:
- Direct comparison of two methods in 138 renal allograft biopsies.
- Method 1: Monoclonal antibody (mAb) with indirect immunofluorescence (IF) on frozen sections.
- Method 2: Polyclonal antibody with immunohistochemical (IHC) detection on FFPE tissue.
Main Results:
- Complete agreement between methods in 97% of initial cases (104/107).
- Overall positivity rate of 8.4% across all biopsies; higher in clinically indicated (15%) and acute cellular rejection (24%) cases.
- IHC method demonstrated 98% specificity and 87.5% sensitivity compared to IF.
Conclusions:
- IHC staining of FFPE tissue for C4d is a viable alternative to IF on frozen sections.
- The IHC method offers acceptable sensitivity and high specificity for diagnosing C4d deposition in renal allografts.
- This finding supports the broader use of FFPE tissues for C4d evaluation in transplant pathology.
Abstract:
Immunostaining of renal allograft biopsies for C4d deposition has become an important diagnostic tool in the recognition of humoral-mediated graft rejection. The majority of studies have been performed on frozen tissue sections with one of several commercially available antibody reagents. However, only a single small series that compared reagents or methods, including staining of formalin-fixed, paraffin-embedded tissue, has been published. Two different staining methods in 138 renal allograft biopsies were compared directly: A mAb (Quidel, San Diego, CA) on frozen tissue sections with indirect immunofluorescence (IF) and a polyclonal antibody (Biomedica Gruppe, distributed by ALPCO, Windham, NH) applied to formalin-fixed, paraffin-embedded tissue with immunohistochemical (IHC) detection. An initial data set of 107 consecutive cases showed complete agreement between staining methods in 104 (97%) cases. Overall, nine of 107 cases were positive with one or both methods, representing 8.4% of all allograft biopsies tested, 15% of clinically indicated biopsies, and 24% of biopsies with a histologic diagnosis of acute cellular rejection. A second set of 31 cases included 17 cases that were positive by either method, with concordance in 29 of 31 cases. Combining the two data sets, the overall specificity of the IHC method compared with IF was 98%, and sensitivity was 87.5%. Direct comparison demonstrates that IHC staining of formalin-fixed, paraffin-embedded tissue with anti-C4d polyclonal antibody has acceptable sensitivity and specificity, as compared with IF staining of frozen tissue with the Quidel mAb.