biotinylated anti hamster ab  (Vector Laboratories)

Journal: Bone

Article Title: Novel approaches for two and three dimensional multiplexed imaging of osteocytes

doi: 10.1016/j.bone.2015.02.011

Figure Lengend Snippet: Multiplexed imaging of osteocytes and their extracellular matrix – (A) Sum Z projected confocal image from a thick 50 µm frozen section of a 10 week old GFP-collagen mouse femur (10 planes with 0.13 µm Z plane separation, 100× oil lens, 1.7× digital zoom, with compensation). Note that the GFP-collagen reveals considerable substructure within the bone ECM and also note the rings of GFP fluorescence around osteocyte lacunae. Inset shows a high magnification view of an osteocyte lacuna with perilacunar GFP-collagen fluorescence. (B) Maximal Z projected 4-color confocal image from the same field of view (100 planes with 0.13 µm Z plane separation, 100× oil lens, 1.7× digital zoom). The GFP-collagen is pseudocolored gray, actin cytoskeleton is shown with Alexa Fluor 633 phalloidin [pseudocolored green], lacunocanalicular system is shown with fixable Texas Red labeled dextran [red] and nuclei are stained with DAPI [blue]. (C) 3D rendered image of the same field in (B) (see online Supplementary Movie 5 for animation of 3D rendered image). (D) Maximal Z projected image of an undecalcified whole mount neonatal mouse calvarium stained with alizarin red and (E) merged image of same field showing alizarin red staining combined with immunostaining for E11/gp38 (biotinylated anti-hamster antibody/streptavidin FITC [green]) (74 planes with 0.43 µm Z plane separation, 40× lens). (F) 3D isosurface rendered image of an enlarged area from (E) [imaging – Leica TCS Sp5 II (A–C) or Zeiss LSM 710 confocal microscope (D–F); image processing – ImageJ; 3D rendering – 3D viewer in ImageJ]. Bar = 20 µm.

Article Snippet: Detection antibodies and reagents used for immunostaining included Cy3 conjugated anti-hamster and anti-goat antibodies and a biotinylated anti-hamster antibody used in conjunction with FITC-streptavidin (Jackson Immunoresearch, Westgrove, PA).

Techniques: Imaging, Fluorescence, Labeling, Staining, Immunostaining, Microscopy