A new technical approach for the ultrastructural analysis of hematopoietic cells using cell monolayers adherent to electron-transparent melamine resins.
The melamine resin is a polymer of a hexamethylol melamine ether which can coat glass slides with an electron-transparent foil (approximately 80 nm thick) after polymerization by p-toluene sulphonic acid and warming. Provided that the cells had been resuspended in a serum-free medium, normal peripheral blood, or bone marrow cells, blasts of different acute leukemias, cells of B-cell chronic lymphocytic leukemia, and of the cell-lines K562, KG1a, and HL-60 became adherent to the melamine-resin-covered glass slides. The optimal sedimentation time and cell concentration was 45 min and 10(7) cells/ml, respectively. Moreover, in serum-free culture medium the cells could be maintained adherent for up to 96 h without a great loss in cell number and viability. For transmission electron microscopical (TEM) analysis, the monolayers could be embedded in situ in epon after routine fixation and staining procedures. Alternatively, the foils could be removed from the glass and mounted on grids for whole mount electron microscopic analysis (WMEM). Both methods could be combined with immunogold labelling for the detection of surface antigens. This technique permits ultrastructural in situ analysis of morphological and/or immunological changes of cells induced by in vitro stimulation.
Department of Internal Medicine III, University of Ulm, Germany.
G
G
Heil
J
J
Krauter
M
M
Hartl
E
E
Beyer-Johannböke
E
E
Gunsilius
A
A
Ganser
E
E
Kurrle
C
C
Westphal-Frösch
D002448:Cell Adhesion; D006412:Hematopoietic Stem Cells / Q000648:ultrastructure*; D006801:Humans; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000473:pathology; D015470:Leukemia, Myeloid, Acute / Q000473:pathology; D008854:Microscopy, Electron / Q000379:methods; D054198:Precursor Cell Lymphoblastic Leukemia-Lymphoma / Q000473:pathology; D012117:Resins, Synthetic; D014227:Triazines; D014407:Tumor Cells, Cultured / Q000648:ultrastructure
D012117:Resins, Synthetic; D014227:Triazines; C011907:melamine