Ab96901

To assess the number of SM cells positive for phosphorylated p38 or phosphorylated JNK, paraffin-embedded sections (2.5-mm-thick) of penile tissue were incubated overnight with primary antibodies against α-SMA (a SM cell marker, 1:100, M0851, Dako, Glostrup, Denmark; red) and phosphorylated p38 (1:20, #4511, Cell-Signaling Technology, Danvers, MA, USA; green) or phosphorylated JNK (1:20, #4668S, Cell-Signaling Technology, Danvers, MA, USA; green). After washing in PBS, the sections were incubated with two secondary antibodies (goat anti-mouse IgG 488 [1:400, A-11001, Invitrogen, Camarillo, CA, USA] and goat anti-rabbit IgG 594 [1:200, ab96901, Abcam, Cambridge, CB4 0FL, UK]) in 1% bovine serum albumin at room temperature for 1 h. Digital images were acquired using a confocal microscope (Leica TCS SP8, Leica Microsystems, Wetzlar, Germany). Ten rats from each group (two sections per animal) were analyzed. Under confocal microscopy, the number of SM cells positive for phosphorylated p38 or phosphorylated JNK (yellow) was quantified from five randomly selected high-power fields (white arrow). The slides were evaluated by three independent observers in a blinded fashion.