Identification of L-plastin autoantibody in plasma of patients with non-Hodgkin's lymphoma using a proteomics-based analysis

Abstract

The diagnosis of malignant lymphoma (ML) such as non-Hodgkin's lymphoma (NHL) and Hodgkin's lymphoma (HL) was mainly performed by morphological examination and gene analysis. There are only a few serum/plasma biomarkers such as lactate dehydrogenase and soluble interleukin-2 receptor alpha to diagnose ML. The classifications are various, and therefore the cell surface markers using flow cytometry or lymph node biopsy have been examined. It is difficult, however, to distinguish the two diseases, NHL and HL, from each other. In order to identify the haematological malignancy-associated autoimmunoreactivity (autoantibodies) in patients' plasma, a novel proteomics-based approach using electrophoresis/mass spectrometry was applied. Solubilized proteins from a Burkitt's lymphoma cell line (Raji) were separated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and Western blotting analysis, in which the plasma of individual patients with haematological malignancies was tested for primary antibodies, followed by visualization with anti-IgG antibody conjugated with horseradish peroxidase. Two proteins, L-plastin and alpha-enolase, capable of reacting with the antibodies in plasma of patients with NHL, were detected using matrix-assisted laser desorption ionization/time-of-flight mass spectrometry and tandem mass spectrometry. The rates of the detections of an anti L-plastin autoantibody were significantly higher: 0.84 (21/25) in patients with NHL; 0.00 (0/4) in HL; 0.38 (5/13) in autoimmune diseases; 0.20 (2/10) in leukaemia; and 0.13 (1/8) in healthy controls. In contrast, those of anti alpha-enolase antibody were not specific to NHL. We first identified autoantibody against L-plastin in plasma of patients with NHL, suggesting that the autoantibody can be a new diagnostic biomarker for NHL.