Immunofluorescence staining was performed as described previously (28 (link),29 (link)). Testicular sections were deparaffinized and rehydrated. Endogenous peroxidase activity was blocked by incubation in 3% hydrogen peroxide for 15 min at 37 °C. After antigen retrieval in sodium citrate buffer, sections were blocked and incubated overnight at 4 °C with primary antibodies (Table S1 ). After three washes with phosphate-buffered saline (PBS), sections were incubated with Alexa Fluor series secondary antibodies (Thermo Scientific, Waltham, USA) and images were captured on a Zeiss laser confocal microscope (LSM800, Carl Zeiss).
Alexa fluor series secondary antibodies
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Alexa Fluor series secondary antibodies are a range of fluorescently labeled antibodies used for detection in various immunoassay techniques. These antibodies are designed to bind to the Fc region of primary antibodies, enabling visualization and quantification of target proteins or cells. The Alexa Fluor dyes used provide a wide selection of excitation and emission spectra, allowing for flexibility in experimental design.
Automatically generated - may contain errors
2 protocols using alexa fluor series secondary antibodies
1
Immunofluorescence Staining of Testicular Sections
2
Histological Evaluation of Colonic Inflammation
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We euthanised mice and excised theirs colons, which were cut open longitudinally and rinsed in 1 × PBS to remove faeces, then fixed in a Swiss roll shape in 10% Formalin Neutral Buffer Solution (Wako). Paraffin-embedded sections were stained with haematoxylin and eosin (H&E) for histological assessment. The disease severity was evaluated according to a previously described scoring system31 (link). The total score was given by the sum of the scores for the proximal, middle, and distal regions of the colon. For immunohistochemical analysis, fresh colon tissues were embedded in OCT compound (Sakura Finetek). Frozen sections were fixed in 4% paraformaldehyde (PFA), blocked in Blocking One (Nacalai Tesque), and incubated with rat anti-CD68 antibody (FA-11, AbD Serotec) or rat anti-B220 antibody (RA3-6B2, BD Pharmingen), followed by incubation with Alexa Fluor® series secondary antibodies (Thermo Fisher Scientific). Samples were examined using a fluorescence microscope (Fluoview FV10i, Olympus).
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