Backgrounds Systemic amyloidosis is classified according to the deposited amyloid fibril protein (AFP), which determines its best therapeutic scheme. The most common type of AFP found are immunoglobulin light chains. The laser microdissection combined with mass spectrometry (LMD-MS) technique is a promising approach for precise typing of amyloidosis, however, the major difficulty in interpreting the MS data is how to accurately identify the precipitated AFP from background. Objectives The objective of the present study is to establish a complete data interpretation procedure for LMD-MS based amyloidosis typing. Methods Formalin-fixed paraffin-embedded specimens from patients with renal amyloidosis and non-amyloid nephropathies (including diabetic nephropathy, fibrillary glomerulonephritis, IgA nephropathy, lupus nephritis, membranous nephropathy, and normal tissue adjacent to tumors) were analyzed by LMD-MS. Forty-two specimens were used to train the data interpretation procedure, which was validated by another 50 validation specimens. Area under receiver operating curve (AUROC) analysis of amyloid accompanying proteins (AAPs, including apolipoprotein A-IV, apolipoprotein E and serum amyloid P-component) for discriminating amyloidosis from non-amyloid nephropathies was performed. Results A stepwise data interpretation procedure that includes or excludes the types of amyloidosis group by group was established. The involvement of AFPs other than immunoglobulin was determined by P-score, as well as immunoglobulin light chain by variable of λ-κ, and immunoglobulin heavy chain by H-score. This achieved a total of 88% accuracy in 50 validation specimens. The AAPs showed significantly different expression levels between amyloidosis specimens and non-amyloid nephropathies. Each of the single AAP had a AUROC value more than 0.9 for diagnosis of amyloidosis from non-amyloid control, and the averaged level of the three AAPs showed the highest AUROC (0.966), which might be an alternative indicator for amyloidosis diagnosis. Conclusions The proteomic data interpretation procedure for LMD-MS based amyloidosis typing was established successfully that has a high practicability in clinical application.
Backgrounds: Systemic amyloidosis is classified according to the deposited amyloid protein, which determines its best therapeutic scheme. The laser microdissection combined with mass spectrometry (LMD-MS) technique is a promising approach for precise subtyping of amyloidosis, however, is hampered by how to interpret the MS data.Objectives: The objective of the present study is to establish a complete data interpretation procedure for LMD-MS based amyloidosis subtyping.Methods: Formalin fixed paraffin-embedded specimens from patients with renal amyloidosis were analyzed by LMD-MS for proteome quantification. Forty-two specimens were used for training the data interpretation procedure, which was validated by another 50 validation specimens. Area under receiver operating curve (AUROC) analysis of amyloid accompanying proteins (APOE, APOA4 and SAMP) for discriminating amyloidosis from non-amyloid nephropathies was performed.Results: A stepwise data interpretation procedure that include or exclude the subtypes group by group was established, in which, involvement of non-immunoglobulin amyloid protein is determined by P-score, involvement of immunoglobulin light chain is determined by variable of λ-κ, and immunoglobulin heavy chain’s participation is judged by H-score. This data interpretation method achieved a 88% accuracy in 50 validation specimens. The amyloid accompanying proteins showed significant quantitative differences between amyloidosis specimens and non-amyloid nephropathies. Each of the single accompanying protein had a AUROC value more than 0.9 for diagnosis of amyloidosis from non-amyloid control, and the averaged value of spectral count of the three accompanying proteins showed the highest AUROC (0.966), indicating it might be an alternative indicator for amyloidosis diagnosis.Conclusions: The proteomic data interpretation procedure for amyloidosis subtyping based on LMD-MS was established successfully, which has high clinical application value.
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