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Tissue biopsies were stained using a tissue gram stain and CFU was quantitated.
Biopsies were stained using hematoxylin eosin and evaluated using the latest criteria for histological diagnosis of aGVHD, which are indicated in Table 1 in the Results section.
Tumour biopsies were stained using immunofluorescence techniques and dual labelling for cytokeratin (cocktail of Dako monoclonal antibodies to AE1/AE3, HMWK, CK7, CK20 or Dako rabbit polyclonal antikeratin wide spectrum screening) was usually carried out to help with the identification of epithelial tumour cells.
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Sections of the brain biopsy were stained using AstV-specific antibody as used previously [ 16], kindly provided by Professor Ian Lipkin.
For cell death detection staining, paraffin-embedded liver biopsy sections (5 μm) were stained using DeadEnd Fluorimetric TUNEL system (Promega, Madison, WI, USA), in accordance with the manual instructions.
Microtubules were stained using antibodies.
Nuclei were stained using TOPRO3.
Formalin fixed, paraffin embedded biopsies were stained by using proCol11a1 monoclonal antibody clone 1E8.33 (ONCOMATRYX, Bilbao, SPAIN) as previously described [ 24, 25].
Formalin-fixed, paraffin-embedded tumour biopsies were stained with anti-factor VIII using immunohistochemistry. Vascularity was scored using the intra-tumour microvessel density (IMD), or 'hot-spot', technique.
Following incubation (1 week), biopsies were stained with Lugol's iodine and examined using a light microscope for the presence of enlarged, black stained parasite cells.
Cross-sections (7 µm) of isopentane-frozen TA biopsies were stained with hematoxylin and eosin and analyzed using a Leica microscope coupled with a camera (Media Cybernetics).
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