Mouse on mouse immunodetection kit bmk 2202

Eyes were fixed in 4% paraformaldehyde, embedded in optical coherence tomography (OCT) compound (Sakura Finetek), and sectioned using a cryostat (CM3050; Leica Microsystems). The section was blocked with 5% donkey serum for 30 min, incubated with goat anti-Brn3a antibody (sc-31984, 1:100; Santa Cruz Biotechnology), mouse anti-chop antibody (#2895, 1:500; Cell Signaling Technology), or rabbit anti-p53 antibody (sc-6243, 1:100; Santa Cruz Biotechnology) for 1 hr and then stained with an appropriate secondary antibody (anti-goat Alexa Fluor 488, anti-rabbit Alexa Fluor 488, or anti-mouse Alexa Fluor 488; Thermo Fisher Scientific) and DAPI (Vector Labs) for an additional 45 min. For primary antibodies raised in mice, frozen sections were treated in using the Mouse on Mouse Immunodetection kit (BMK-2202, Vector Labs) according to the manufacturer’s instructions. Cells in the GCL were counted in a 1 mm² area on each side of the optic nerve under a 20× objective in five 60-μm-spaced sections from each retina.

Immunostaining of brain sections and quantification of Iba1⁺ cells and synaptophysin⁺ puncta was performed as described previously (22 (link)). For p-TDP43 staining, a Mouse on Mouse Immunodetection Kit (BMK-2202; Vector Laboratories) was used to reduce background. Images were acquired on a Nikon C2 confocal microscope with a 60× objective. For analysis of lipofuscinosis, DAPI-stained brain sections were imaged and total autofluorescence intensity per field in the green channel was quantified using ImageJ.