[Classification of amyloidosis by laser micro-dissection and mass spectrometry based proteomic analysis]

Abstract

Objective: To establish a novel method to determine specific type of amyloidosis through laser microdissection and mass spectrometry (LMD/MS) based proteomic analysis.

Methods: There were 138 formalin-fixed and paraffin-embedded (FFPE) biopsy samples of patients who were diagnosed as systemic amyloidosis used in this study. For each case, a 10 μm section stained with congo-red and positive amyloid deposits were identified under fluorescent light, followed by micro-dissection and mass spectrometry analysis. The amyloidosis subtype was confirmed based on the most abundant amyloid protein.

Results: The tissue types of 138 specimens were as following: subcutaneous abdominal fat accounted for 26%, tongue for 19%, gingiva for 11%, kidney for 9%, intestine for 9%, heart for 6% and others for 20%. Specific types of amyloid were accurately detected in 121 cases, including 106 (87.6%) amyloid light chain (AL) type, 7 (5.8%) amyloid trans-thy-retin (ATTR), 2 (1.7%) amyloidogenic protein A (AA), 2 (1.7%) amyloid heavy chain (AH)/AL+AH, 2 (1.7%) fibrinogen alpha chain (AFib), 1(0.8%) amyloid apolipoprotein A-type II (AApoA-II) and one (0.8%) amyloid lysozyme (ALys). Diagnosis of amyloidosis was excluded in 5 cases. The types of twelve cases were indeterminate by LMD/MS. On the whole, LMD/MS reached 91.3% accuracy rate in amyloid typing. Commonly involved organs (for example, heart, kidney and liver) turned out to be suitable sources of FFPE samples with typing success rate of almost 100%. In contrast, MS analysis was successful in only 83.3% of subcutaneous abdominal fat samples.

Conclusion: LMD/MS method provided a more direct technique for accurate typing of amyloidosis in a single procedure.

图1. 激光显微切割舌体标本系统性淀粉样变性病变区域流程图（刚果红染色，×150）

A：光镜下可见鳞状上皮下均质粉染物（箭头所示）；B：与周边组织相比，病变部位在荧光显微镜下呈现亮红色（箭头所示）；C：选取病变区域，开始显微切割；D：切割区域组织已脱落到Eppendorf 管内