Detection of immunoglobulins and complement components in formalin fixed and paraffin embedded renal biopsy material by immunoflourescence technique

Mubarak, Muhammed; Kazi, Javed Iqbal; Kulsoom, Umme; Ishaque, Muhammed

doi:10.5812/nephropathol.7518

Cited by 10 publications

(9 citation statements)

References 14 publications

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“…The efficacy of IF-FFPE in renal biopsies has been studied in human medicine. In one study, three methods of AR (heating with a Tris buffer of unknown pH, heating with CB, and pronase digestion) were tested, and it was demonstrated that heating with Tris buffer was most effective, allowing the evaluation of ICs in renal biopsy specimens [ 9 ]. The efficacy of dual MW-heating methods using an EDTA buffer (pH 8.0) has also been previously reported [ 8 ].…”

Section: Discussionmentioning

confidence: 99%

“…Using human renal biopsies, routine IF-F and IF-FFPE have been compared in several reports [ 7 , 10 – 12 ]. Although one report suggested that IF-FFPE could not replace IF-F in renal biopsy assessment [ 12 ], most studies demonstrated good agreement between the two methods [ 7 , 9 – 11 ]. In this study, 12 biopsy samples were used to compare IF-FFPE after Try-30 and MW-TEB pretreatment to routine IF-F.…”

Section: Discussionmentioning

confidence: 99%

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Paraffin immunofluorescence for detection of immune complexes in renal biopsies: an efficient salvage technique for diagnosis of glomerulonephritis in dogs

et al. 2017

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BackgroundRenal biopsy is an essential tool for the diagnosis of proteinuric kidney diseases in dogs, and evaluation of immune complexes (IC) by immunofluorescence (IF) of frozen sections (IF-F) is required for the diagnosis of IC-mediated glomerulonephritis (ICGN). However, the use of frozen sections from renal biopsies can have limitations. The aim of this study was to develop a reliable IF method using formalin-fixed and paraffin-embedded (FFPE) sections to detect ICs in dog ICGN.MethodsRenal biopsy specimens were obtained from dogs with protein-losing nephropathies. FFPE sections were prepared, and eight antigen retrieval pretreatment protocols were performed: digestion with trypsin, microwave (MW) heating in citrate buffer (MW-CB; pH 6.0), MW heating in Tris-EDTA buffer (MW-TEB; pH 9.0), as well as combinations of the above, and a non-treated control.ResultsA combination of trypsin for 30 min (Try-30) and MW-TEB; pH 9.0 was the most effective antigen retrieval pretreatment, with clear positive signals for IgG, IgA, IgM, and C3 detected by IF-FFPE. Granular signals, an important diagnostic indicator of ICGN, were clearly observed by both IF-F and IF-FFPE after combined pretreatment with Try-30 and MW-TEB, and IgG, IgA, IgM, and C3 signals were almost completely matched in all samples by IF-F and IF-FFPE.ConclusionIF-FFPE with Try-30 and MW-TEB pretreatment is a valuable technique for the diagnosis of renal diseases in dogs. This method could be an efficient tool when standard IF-F cannot be used, or does not provide useful results due to lack of glomeruli in the specimens for IF-F.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Paraffin immunofluorescence for detection of immune complexes in renal biopsies: an efficient salvage technique for diagnosis of glomerulonephritis in dogs

et al. 2017

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“…Because formalin fixation induces protein cross-linking, which generally blocks antigenicity, an antigen-retrieval step that allows for increased penetration of antibodies to the antigens “masked” by formalin fixation is required in IF-P. 5 , 6 This step involves incubating the paraffin sections with a proteolytic enzyme or heating the sections before incubation with fluorescein isothiocyanate–conjugated antibodies against Igs and complement components. Multiple proteolytic enzymes have been used in IF-P, including trypsin, 7 , 8 , 9 , 10 , 11 , 12 pronase E (protease XIV), 13 , 14 , 15 , 16 , 17 proteinase XXIV, 18 and proteinase K. 13 , 19 , 20 , 21 , 22 Successful results were also obtained by heat treatment with Tris or citrate buffers 15 and with dual microwave heating in EDTA antigen-retrieval solution. 23 In our laboratory, we use the pronase technique, which was originally described by Fogazzi et al.…”

Section: Methodologies Of Paraffin Immunofluorescencementioning

confidence: 99%

“… 37 (vi) The various IF-P techniques described are based on formalin fixation. 4 , 9 , 14 , 15 , 20 , 21 The performance characteristics of IF-P on non–formalin-based fixatives (Bouin, Duboscq-Brasil, and other fixatives) used in approximately 25% of renal pathology laboratories 38 , 39 are not known.…”

Section: Methodologies Of Paraffin Immunofluorescencementioning

confidence: 99%

Paraffin Immunofluorescence: A Valuable Ancillary Technique in Renal Pathology

Nasr

Fidler

Said

2018

Kidney International Reports

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Immunofluorescence on frozen tissue is the gold standard immunohistochemical technique for evaluation of immune deposits in the kidney. When frozen tissue is not available or lacks glomeruli, immunofluorescence can be performed on paraffin tissue after antigen retrieval (paraffin immunofluorescence). Excellent results can be obtained by paraffin immunofluorescence in most immune complex–mediated glomerulonephritides and dysproteinemia-associated kidney lesions, and thus this technique has become a valuable salvage technique in renal pathology. Furthermore, new data have emerged suggesting that paraffin immunofluorescence can be used as an unmasking technique, as it is more sensitive than frozen tissue immunofluorescence in some kidney lesions, such as crystalline light chain proximal tubulopathy and is needed to establish the diagnosis of certain unique lesions, such as membranous-like glomerulopathy with masked IgG kappa deposits and membranoproliferative glomerulonephritis with masked monotypic Ig deposits. However, it is important to recognize and be aware of the limitations and pitfalls associated with paraffin immunofluorescence. These include poor sensitivity for detection of C3 deposits and for the diagnosis of primary membranous nephropathy. Here, we summarize the available techniques of paraffin immunofluorescence, review its role and performance as a salvage and unmasking technique in renal pathology, address its limitations and pitfalls, and highlight unusual forms of glomerulopathy that require paraffin immunofluorescence for diagnosis.

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“…Several laboratory techniques have been used to detect immunoglobulin complexes, such as ELISA for soluble complexes [8] and immunohistochemistry to trace their deposition in tissues [9]. Recently, flow cytometry is applied to detect immune complexes on the surface of erythrocytes.…”

Section: Introductionmentioning

confidence: 99%

Delivery route determines the presence of immune complexes on umbilical cord erythrocytes

Lima

Franco

Sarmiento

et al. 2016

The Journal of Maternal-Fetal & Neonatal Medicine

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Detection of immunoglobulins and complement components in formalin fixed and paraffin embedded renal biopsy material by immunoflourescence technique

Abstract: In conclusion, it is possible to establish the diagnosis in most cases of immune complex-mediated glomerular diseases with IF on paraffin embedded tissue specimens.

Cited by 10 publications

References 14 publications

Paraffin immunofluorescence for detection of immune complexes in renal biopsies: an efficient salvage technique for diagnosis of glomerulonephritis in dogs

Paraffin immunofluorescence for detection of immune complexes in renal biopsies: an efficient salvage technique for diagnosis of glomerulonephritis in dogs

Paraffin Immunofluorescence: A Valuable Ancillary Technique in Renal Pathology

Delivery route determines the presence of immune complexes on umbilical cord erythrocytes

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