or labeling of the specimen is not necessary, making preparation easy.
On the second day, after several washes with phosphate buffered saline, the sections were incubated with biotinylated rabbit anti-rat secondary antibody and avidin-biotin horseradish peroxidase (Vector Elite Avidin Biotin Complex system, Vector Laboratories, Burlingame, CA) for 30 minutes, respectively, visualized with 3,3-diaminobenzidine (DAB kit, Vector Laboratories) followed by counterstaining
with 10% Mayer's hematoxylin, and finally, mounted in Permamount (Vector Laboratories, Burlingame, CA), and examined by light microscopy.
All nuclei were visualized by propidium iodide (PI) counterstaining
Slides of formalin-fixed, paraffin embedded tumors were immunohistochemically stained involving rehydration, antigen retrieval, primary, antibody application, secondary antibody application, label application, staining, counterstaining
by PI allows the discrimination of apoptotic from necrotic cells.
2]) as substrate, followed by counterstaining
Binding was detected by using monoclonal antibody 2G3 against RHDV, biotinylated anti-mouse IgG, horseradish peroxidase-conjugated avidin, and 3-amino-9-ethylcarbazole substrate with hemalum counterstaining
, as described (2).
By employing the multiplexing capabilities of this imaging system, cell enumeration was carried out by counterstaining
Hematoxylin and eosin counterstaining
was used to visualize cellular structures.
with hematoxylin, the slides were dehydrated with grade series of alcohol, cleared in xylene, and mounted with a coverslip.
We used the methods outlined by Kent (1988) by staining with hematoxylin and then counterstaining
with eosin to enhance the contrast between the lines on the hinge.
the nuclei with diamidinophenylindole (DAPI; Molecular Probes), sections were examined with a Zeiss LSM 510 laser scanning confocal microscope system (Carl Zeiss Inc.
Y-specific immunofluorescent stain with histiocyte counterstaining