[Clinical applications of MALDI imaging using sliced sections of formalin-fixed paraffin-embedded tissues and longitudinal sliced hairs].

MALDI-imaging MS (IMS) with MSMS analysis is a new powerful tool for the identification of not only disease-related proteins in formalin-fixed paraffin-embedded (FFPE) tissue sections but also protein/peptides/drugs/medicine in fresh-frozen tissues. IMS is used to reveal the mass profiles and spatial distribution of proteins in tissue sections and/or digested peptides derived from deposited protein in pathologic organs and then MSMS analysis identifies the amino acid sequence of the detected proteins in the tissue section. Moreover, on-tissue digestion combined with the MALDI-IM-TOF-IMS approach allows a proteomics "bottom-up" strategy with clinical samples, especially perioperative isolated tissues and FFPE tissues conserved for a long time in a clinical sample bank. The mass barcode-like image (MBI) on a longitudinal sliced hair by IMS is used in the selected reaction monitoring mode for serially chronological monitoring and traceability every few hours after drug and medicine intake. The advances of quantitative MBI for sliced sections of hair allow a new universal standardized assessment of drugs and medicines throughout the drug history.

[In a disaster, what should medical technologists at a university hospital do? What do they have?].

"Disaster medicine" involves "cases that cannot be supported by the normal medical service system of the hospital because many injuries have occurred by an explosion/chemical pollution/radioactive pollution or a pandemic caused deliberately as well as natural disasters". In "disaster medicine", university hospitals should become the base for advanced medical services and wide area transportation. Also, the clinical laboratory/medical technologists of the university hospital should offer high quality laboratory analysis. On the other hand, one of the advantages of a university hospital is an experimental medicine laboratory (integrated universities also have a Department of Science and Department of Engineering). Development of testing equipment to cover all the functions necessary for disaster medicine may be a possibility. A system to conduct simple testing is expected at the actual location of the emergency, and an identification system must be established. "Emergency medicine" and "disaster medicine" have different aspects, but the essence is the same, and medical technologists must have knowledge/techniques to report and interpret results quickly. Because a university hospital is the core of logistical support, daily training is important.

[The values of multi-laboratories serum values evaluation].

The members of 23 laboratories, ten clinical laboratory centers and thirteen hospital laboratories in the Kinki District participated in share their clinical laboratory data. In this joint work, we cross-checked twenty-seven serum values, and all data from the 23 laboratories well accorded; however, several values, such as urea nitrogen, calcium, and albumin needed to be standardized to share the laboratory data.

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

BACKGROUND: 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. METHODS: 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. RESULTS: 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. CONCLUSIONS: 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.

Detection of citrullinated proteins in synovial fluids derived from patients with rheumatoid arthritis by proteomics-based analysis.

BACKGROUND: Rheumatoid arthritis (RA) is the most common inflammatory joint disease. The aetiology of RA remains unknown, but autoimmune responses are considered to play an important role in the disease pathophysiology. Currently available data suggests that the process of diagnosing RA may benefit from testing for anticyclic citrullinated peptides. Identification of the presence of citrullinated proteins in rheumatoid synovial fluids is important for the elucidation of the aetiology of RA as well as in the differential diagnosis of rheumatic-related diseases. METHODS: A proteomics-based approach using electrophoresis/mass spectrometry was applied to identify the citrullinated proteins in synovial fluids from patients with RA. Synovial fluids from patients with RA were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot analysis to detect the citrullinated proteins. Identification bands were then subjected to mass spectrometry. RESULTS: Three proteins - citrullinated fibrinogen, citrullinated fibronectin and citrullinated vimentin - in synovial fluids from RA patients were identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. CONCLUSIONS: Proteomics-based analysis can be used to detect citrullinated proteins in synovial fluids from RA patients.

Detection of eight antibodies in cancer patients' sera against proteins derived from the adenocarcinoma A549 cell line using proteomics-based analysis.

To screen cancer for specific autoantibodies, we applied the approach established by Brichory et al., who reported annexins I and II as specific antigens. Solubilized proteins from a cancer cell line (A549) were separated using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), followed by Western blotting (WB) analysis, in which the sera of individual patients were tested for primary antibodies. We found 11 positive spots on PVDF membrane using a WB/enhanced chemiluminescence detection Kit, and identified eight proteins, such as alpha-enolase, inosine-5'-monophosphate dehydrogenase, aldehyde dehydrogenase, 3-phosphoglycerate dehydrogenase, 3-oxoacid CoA transferase, chaperonin, peroxiredoxin 6 and triosephosphate isomerase, that reacted with these antibodies in patients' sera using MALDI-TOF/TOF. All eight antibodies were not detected in the sera derived from lung tuberculosis and healthy controls.

[Proteome analysis of autoantibodies in sera of patients with cancer].

Circulating autoantibodies are useful diagnostic markers of cancers and autoimmune diseases. Research over the past decade has resulted in some reports on the presence of autoantibodies against disease-related proteins such as annexin-I & II, recoverin and protein gene product 9.5 in the sera of patients with lung cancer, and also against calreticulin and alpha-enolase in autoimmune diseases. In this study, we first identified the a-enolase autoantibody in the sera of patients with lung adenocarcinoma by proteomics-based analysis. The comparison of two-dimensional polyacrylamide gel electrophoresis (2D-PAGE)/western blot (WB)/ECL detection revealed distinct distributions of antibodies in the sera of lung adenocarcinoma, tuberculosis and healthy subjects which reacted with soluble proteins derived from the adenocarcinoma A549 cell line. We found 16 spots in patients with adenocarcinoma by 2D-PAGE/WB/ECL detection and identified alpha-enolase, chaperonin, and other autoantibodies in the adenocarcinoma patients' sera. The specificities of an antibody against alpha-enolase was preliminarily observed in sera from 3 of 5 patients with adenocarcinoma, 0 of 10 patients with tuberculosis and 0 of 10 healthy subjects. In conclusion, we first identified alpha-enolase autoantibody in sera of lung adenocarcinoma and the autoantibody was seemed to be a specific marker of the lung adenocarcinoma. In addition, we also identified various autoantibodies in esophageal cancer, hepatocellular carcinoma, and non-Hodgkin's lymphoma. Moreover, we tried to identify the corresponding antigen of an unknown anti-cytoplasmic autoantibody, and an anti-red blood cell antibody by proteomics-based analysis. These antibodies might become new diagnosis markers.