Classification Of Amyloidosis Research Articles

Diagnosis and Classification of Amyloidosis in Abdominal Subcutaneous Fat Aspiration Specimens Using Mass Spectrometry Based Proteomics

Abdominal subcutaneous fat aspiration is one of the most practical, sensitive and specific methods for the diagnosis of systemic amyloidosis. One limitation of this method, compared to more invasive tissue biopsy based approaches, remains the technical difficulties in further classification of the amyloidosis as commonly used methods, such as immunohistochemistry, are not readily applicable to fat aspiration specimens. To overcome these difficulties we developed a method using nano-flow liquid chromatography electrospray tandem mass spectrometry (LC-MS/MS) that could identify amyloid subtypes in freshly obtained Congo Red positive fat aspirate specimens with great accuracy.Abdominal subcutaneous fat aspirate specimens were obtained from 73 patients with clinical suspicion for systemic amyloidosis. One half of the specimen was stained with Congo red and used for diagnosis of amyloidosis and the other half was processed and enzyme digested for LC-MS/MS analysis. The resulting LC-MS/MS data was correlated to theoretical fragmentation patterns of tryptic peptide sequences from the Swissprot database using Scaffold (Mascot, Sequest, and X!Tandem search algorithms). Peptide identifications were accepted if they could be established at greater than 90.0% probability and protein identifications were accepted if they could be established at greater than 90.0% probability and contain at least 2 identified spectra. The identified proteins were subsequently examined for the presence or absence of amyloid related peptides. Of the 73 cases studied, 41 were positive for Congo red consistent with systemic amyloidosis. In Congo red positive cases, LC-MS/MS peptide profiles consistent with AL-lambda (28/31), AL-kappa (6/7), and ATTR (2/3) were observed. Only one case in the Congo red negative control group (31/32) gave a kappa light chain profile which was attributed to a high level of kappa in the serum (285 mg/dL). Of the 35 out of 41 cases of systemic amyloidosis successfully classified by LC-MS/MS, additional clinical and pathology data validating the amyloid type was available. In each of these cases the MS/MS results accurately predicted the amyloid type.In conclusion, LC-MS/MS proteomic analysis of abdominal subcutaneous fat aspiration specimens involved by amyloidosis provides a highly specific (97% specificity) and sensitive (>85% sensitivity) method for diagnosis and classification of amyloidosis. The method is rapid and readily applicable in a clinical setting and will greatly improve the clinical management of amyloidosis patients.

Typing of amyloidosis in renal biopsies: diagnostic pitfalls.

Abstract Context.—Amyloidosis represents a group of diseases with extracellular deposition of congophilic fibrils of similar morphology but differing chemical composition. The types commonly involving the kidney are AL (light chain amyloid) and AA (serum amyloid A). Familial amyloidosis can also affect the kidney, but we have not encountered such a case during the study period. Distinguishing between the AL and AA forms of amyloid is clinically important because of the different treatments and outcomes. The classification of amyloidosis is made by immunostaining with antibodies to κ and λ immunoglobulin light chains and for serum amyloid A protein. Objective.—To draw attention to the nonspecific immunofluorescence staining patterns in renal biopsies with amyloidosis, causing potential diagnostic pitfalls. Design.—Renal biopsies from 15 patients, including 13 cases of AL and 2 cases of AA amyloidosis, were studied. Immunofluorescence staining with routine antibody panel and immunoperoxidase staining for am...

Typing of Amyloidosis in Renal Biopsies: Diagnostic Pitfalls

Amyloidosis represents a group of diseases with extracellular deposition of congophilic fibrils of similar morphology but differing chemical composition. The types commonly involving the kidney are AL (light chain amyloid) and AA (serum amyloid A). Familial amyloidosis can also affect the kidney, but we have not encountered such a case during the study period. Distinguishing between the AL and AA forms of amyloid is clinically important because of the different treatments and outcomes. The classification of amyloidosis is made by immunostaining with antibodies to kappa and lambda immunoglobulin light chains and for serum amyloid A protein. To draw attention to the nonspecific immunofluorescence staining patterns in renal biopsies with amyloidosis, causing potential diagnostic pitfalls. Renal biopsies from 15 patients, including 13 cases of AL and 2 cases of AA amyloidosis, were studied. Immunofluorescence staining with routine antibody panel and immunoperoxidase staining for amyloid A were performed. Of the 13 cases of AL amyloidosis, 2 cases showed little difference in staining intensity between kappa and lambda light chains (2+ and 3+, respectively) and 4 cases showed only moderate intensity (2+) of the predominant light chain. The 2 cases diagnosed as AA amyloidosis also exhibited staining for light chains. One case had strong (3+) signal for kappa and moderate (2+) for lambda light chain, while the other showed weaker staining. Immunofluorescence staining for immunoglobin light chains on renal biopsy, as the first step to differentiate between AL and AA amyloidosis, may sometimes be inconclusive or even misleading. Applying amyloid A immunostain on a routine basis and detailed clinical history are essential to avoid misclassification.

Classification of amyloidosis: misdiagnosing by way of incomplete immunohistochemistry and how to prevent it

Classification of every individual case of amyloid disease is necessary in order to recognize its origin and its possible pathogenesis for therapeutic consideration. Classification of the amyloids can be performed in different ways. One method primarily exploits serum proteins-but these are risk factors only, and therefore render only ancillary information. In principle, one cannot establish the diagnosis alone through their use. Another approach analyzes the origin of the deposited amyloids, either by extracting the amyloid proteins followed by immunochemical or chemical analysis, or by using immunohistochemistry. Based on chemical analysis of prototypes of amyloid fibril proteins, we have developed a profile of antibodies over the years that specifically identify amyloid in tissue sections. These antibodies have been used for years as a routine service for clinicians and pathologists in immunohistochemically classifying amyloid found in formalin-fixed tissue sections. The typing is always controlled by established amyloid classes. In several cases, we have been asked for a second opinion on a diagnosed amyloid class. Our own immunohistochemical data were then compared with those submitted. These submitted immunohistochemical results represented misdiagnoses of amyloid classes in most patients, since the technique performed was usually incomplete. It is the purpose of this report to analyze such cases and to document some of the typical mistakes. Here, we show how to avoid common pitfalls and how one can arrive at a correct diagnosis using immunohistochemistry appropriately.

Biochemical Micro-Techniques in the Diagnosis and Classification of Amyloidosis

Biochemical Micro-Techniques in the Diagnosis and Classification of Amyloidosis

The analysis of gastrointestinal amyloidosis in 78 patients with chronic renal failure

Systemic amyloidosis is not a single disease, but the product of a variety of diseases. Amyloid proteins are insoluble fibrils that are deposited extracellularly in many organ tissues. They stain with Congo red and appear apple green under polarized light. Definitive diagnosis and classification of amyloidosis requires histologic examination of tissue samples. Gastrointestinal tract involvement is common, and all parts of the system can be affected. Immuno–histochemical studies have shown that amyloid deposited in the gastrointestinal system is most often of the AA, A kappa, or A lambda types. Another type of amyloid protein, beta–2 microglobulin (β2M), predominantly affects the musculoskeletal system, and is usually seen in patients who have been on long–term hemodialysis. Mixed systemic amyloidosis (β2M and AA) is seen only rarely in these patients. In this study, we attempted to answer why this is so, and examined whether or not mixed amyloidosis is related to amyloidogenesis.We studied gastrointestinal tissues from 78 chronic renal failure patients who had systemic amyloidosis with gastrointestinal involvement. A total of 115 endoscopic samples and 1 jejunal resection specimen were analysed immunohistochemically. Immunohistochemical testing using a panel of antisera directed against two major amyloid fibril proteins (AA–Monoclonal, Dako–, and β2 MPolyclonal, Dako–) showed that all samples contained AA amyloid, but not β2M type protein. These findings can be explained by the patients' relatively short average duration of hemodialysis and the predominance of endoscopic biopsy samples in our study.

Amyloid protein and amyloidosis.1.Classification of amyloidosis.

Amyloid protein and amyloidosis.1.Classification of amyloidosis.

Demonstration and classification of amyloidosis in needle biopsies of the kidneys, with special reference to amyloidosis of the AA-type.

To examine whether sequence-specific antibodies directed against serum amyloid A were useful in the demonstration and classification of amyloidosis, needle biopsy specimens from the kidneys of 152 cases with renal disorders were investigated using the avidin-biotin-peroxidase complex technique of immunohistochemistry. A distinct immunoreactivity of protein AA was seen in biopsies from all 42 individuals who were clinically classified as having the AA-type of amyloidosis. The stained areas coincided with deposits stained by Congo red. Four of these cases demonstrated immunoreactivity of both protein AA and light immunoglobulin chains and all biopsies except one showed immunoreactivity for the amyloid P-component. After treatment with potassium permanganate, the amyloid deposits in the biopsies of all 42 cases lost their affinity for Congo red. Ten patients with clinical and laboratory findings compatible with the AL-type of amyloidosis were also investigated. All their biopsies demonstrated Congophilic amyloid deposits but none of them showed any immunoreactivity of protein AA. Amyloid deposits of lambda light immunoglobulin chains-but not kappa-were demonstrated in biopsies from four patients. The amyloid P-component was found in biopsies from six individuals and positive Congo red staining after treatment with potassium permanganate was seen in biopsies from four of the cases. Biopsies of 100 patients suffering from non-amyloid renal disorders were also examined. None of them displayed any immunoreactive deposits of protein AA. The investigation shows that amyloid deposits of the AA-type can be identified in needle biopsies when sequence-specific antibodies against serum amyloid A are used in the avidin-biotin-peroxidase complex technique. Both the diagnostic sensitivity (42 of 42) and specificity (110 of 110) of the assay were optimal (1.0). The method was found to be superior to other investigated techniques and useful for classifying amyloidosis in formalin-fixed renal biopsies.

両側頬部にみられた限局性アミロイドーシスの１例

Amyloidosis is a lesion which amyloid substance, fibrous protein, is deposited in all organs and tissues of the body.In the oral region, tongues are most affected. Lips, cheeks, palates and submandibular salivary glands may also be affected, but few cases were reported so far.Clinical classification of amyloidosis is generally as follows;(1) primary amyloidosis, (2) amyloidosis with multiple myeloma, (3) secondary amyloidosis, (4) hereditary amyloidosis, (5) organ limited amyloidosis.Recently, we have experienced a case of amyloidosis, which had occured in both cheeks of a 45-year old man.The patient complained of a tumor formations of both cheeks.These tumors were excised intraorally and amyloid fibrils were detected in them by light and electron microscopy.Furthermore, by several systemic examinations this case was diagnosed as organ limited amyloidosis.

Isolation and Characterization of a x Amyloid Fibril Protein

The fibril in primary amyloidosis (AL) is composed of a monoclonal light chain or portions thereof. No unique primary structure has been identified that predisposes certain light chains to form amyloid fibrils. Currently, classification of amyloidosis is based on the biochemistry of the amyloid fibril. We determined the NH2-terminal sequence of an amyloid fibril and found it to be of the kappa I immunoglobulin subgroup. No structural alterations were detected to account for the conversion of the light-chain fragment to an amyloid fibril. Antiserum produced to the fibril protein did not react in immunodiffusion with purified LEP or MAG antigens, which are kappa I proteins. This antiserum may be directed to antigenic sites unique to the immunizing protein and is unable to recognize homologous proteins, rendering it unsuitable for immunochemical identification of amyloid deposits of light-chain origin. PAG represents the 10th reported variable kappa I amyloid fibril protein subjected to partial sequence analysis. Antisera that recognize antigenic determinants present in all members of an immunoglobulin subgroup need further development.

Primary localized amyloidosis of the bladder: A case report with review of the literature and criteria for classification of amyloidosis

Primary localized amyloidosis of the bladder: A case report with review of the literature and criteria for classification of amyloidosis

Demonstration of amyloid in murine and human secondary amyloidosis by the immunoperoxidase technique.

A specific rabbit anti-murine amyloid A protein (anti-AA) has been used in the immunoperoxidase (IP) technique for the detection of amyloid in formalin-fixed, paraffin-embedded, tissue obtained from amyloidotic mice and a post-mortem kidney specimen from a patient with amyloidosis secondary to osteomyelitis. Amyloidosis was induced in five out of 20 white mice by weekly intraperitoneal injections of complete Freund's adjuvant. Amyloid deposits were clearly demonstrated both by the Congo Red and by the IP techniques. The antibody against murine AA was found to be cross-reactive with human AA. These preliminary results suggest that the IP technique with the use of specific antibodies against the different amyloid proteins might be extremely useful for the histological dianosis and classification of amyloidosis.

Current Concepts of Amyloid

This chapter discusses the structure, chemistry, immunology, cellular origin, metabolism, and possibly the pathogenesis of amyloid. The simplest and most widely used classification of amyloidosis is based on the four major categories introduced by Reimann. Primary amyloid occurs with no known antecedent or coexistent disease and usually involves mesodermal tissues, such as smooth or skeletal muscle, or the cardiovascular system. Generally there is variability in staining of the deposits with Congo red, iodine, and metachromatic dyes. Secondary amyloid is usually associated with chronic diseases, such as infections, neoplasms, neurological disorders, or connective tissue diseases—especially rheumatoid arthritis. It generally involves spleen, liver, kidney, intestines, and adrenals and tends to have reproducible and characteristic staining properties with the above mentioned dyes. Amyloid associated with myeloma tends to resemble the primary type but is invariably associated with a neoplasm involving plasma cells or lymphocytes—such as multiple myeloma, macroglobulinemia, or heavy chain disease. Tumor-forming amyloid is characterized by the small masses of amyloid in the skin, eye, bladder, urethra, respiratory tract, or other organs and is generally unassociated with any underlying disease.

Classification of amyloidosis

Classification of amyloidosis

CLASSIFICATION OF AMYLOIDOSIS WITH SPECIAL REGARD TO THE GENETIC TYPES.

CLASSIFICATION OF AMYLOIDOSIS WITH SPECIAL REGARD TO THE GENETIC TYPES.