title: Analysis of microvessels in pancreatic cancer: by light microscopy, confocal laser scan microscopy, and electron microscopy.
abstract: OBJECTIVE
  The microvessel density (MVD) of most malignant tumors is considered to be strongly related to metastasis and prognosis. Weidner's "hot spot method" for determining MVD is in general use, but it is possible that cells other than endothelial cells will also be stained. In our previous study, no correlations were observed between MVD determined by the "hot spot method" and prognosis/metastasis. But, using the "lumen method," we found a correlation with the number of vessel structures only. In the present study, we analyzed the staining of microvessels in pancreatic cancer, using light microscopy, confocal laser scan microscopy (CLSM), and transmission electron microscopy (TEM).
  
  
  METHODS
  Microvessel staining of pancreatic cancer with CD34, factor VIII, and CD45 antibodies was examined in consecutive slices by light microscopy. For CLSM, freshly resected specimens were immunostained with factor VIII and fluorescein isothiocyanate. For TEM, specimens were fixed with 2.5% glutaraldehyde, treated with 1% osmium tetroxide, and embedded in epoxy resin.
  
  
  RESULTS
  Staining of vessels with CD34 and factor VIII antibodies appeared similar under light microscopy. However, CD34-stained consecutive slices were judged not to reveal vessel structures, and some cells stained with CD45 antibody were similar in appearance to CD34-stained cells. Under CLSM, irregular arrangements of neovascularization, consisting of many branches, were observed, but many positively stained cells not identified as vessels were also seen. Microvessels were distinctly identified under TEM, but the types of individual cells could not be determined.
  
  
  CONCLUSIONS
  An integrated, reproducible method for the measurement of MVD is vital. For pancreatic cancer, the "lumen method" is recommended.
authors.affiliation: Department of Surgery, Teikyo University School of Medicine, 2-11-1 Kaga-cho, Itabashi-ku, Tokyo 173-8606, Japan.
authors.forename: Kenji
authors.initials: K
authors.lastname: Takagi
authors.forename: Tadahiro
authors.initials: T
authors.lastname: Takada
authors.forename: Hodaka
authors.initials: H
authors.lastname: Amano
authors.forename: Masahiro
authors.initials: M
authors.lastname: Yoshida
authors.forename: Humihiko
authors.initials: H
authors.lastname: Miura
authors.forename: Naoyuki
authors.initials: N
authors.lastname: Toyota
authors.forename: Keita
authors.initials: K
authors.lastname: Wada
authors.forename: Ichirou
authors.initials: I
authors.lastname: Takahashi
mesh_terms: D021441:Carcinoma, Pancreatic Ductal / Q000098:blood supply* / Q000378:metabolism / Q000473:pathology; D006801:Humans; D007150:Immunohistochemistry; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D055806:Microvessels / Q000033:anatomy & histology*; D009389:Neovascularization, Pathologic / Q000473:pathology*; D010190:Pancreatic Neoplasms / Q000098:blood supply* / Q000378:metabolism / Q000473:pathology