Anti mouse or anti rabbit igg hrp linked secondary antibodies

Mice were killed by cervical dislocation at 16 weeks of age and their brains were collected and stored at −80°C. The hippocampus was bilaterally dissected and separated into dorsal and ventral hippocampus (~50/50). Protein extraction and Western blot analysis were performed as previously described (Hill et al., 2014 (link)). Primary antibodies were rabbit anti-BDNF (1:200, Santa Cruz Biotechnology Inc, Santa Cruz, CA, USA or Almone Labs, Israel), rabbit anti-NT-4 (1:200, Santa Cruz), rabbit anti-pTrkB Y705 (1:1,000, Signalway Antibody LLC, Maryland, USA), rabbit anti-pTrkB Y515 (1:1,000, Abcam, Cambridge, MA, USA), rabbit anti-pTrkB Y816 (1:500, Millipore, CA, USA), rabbit anti-TrkB (1:1,000, Santa Cruz), rabbit anti-NMDAR subunit 1 (GluNR1, 1:1,000, Cell Signaling Technology Inc, Danvers, MA, USA), rabbit NMDAR subunit 2A (GluN2A, 1:1,000, Cell Signaling Technology), rabbit NMDAR subunit 2B (GluN2B, 1:1,000, Cell Signaling Technology), or mouse anti-β-actin (1:10,000, Sigma-Aldrich). Secondary antibodies included anti-mouse or anti-rabbit IgG HRP-linked secondary antibodies (1: 2,000; Cell Signaling Technology; Danvers, MA, USA).

Immunoblotting was carried out using 15 or 30 µg of total protein lysate as described previously. The following primary antibodies were used: anti-β-actin and anti-Laminin (Abcam, Cambridge, UK); anti-ERK1 K-23 and anti-FGFR3 B-9 (Santa Cruz Biotechnology); anti-Phospho ERK1/2, T202/Y204, anti-α-E-Catenin 23B2 and anti-Phospho-α-E-Catenin S652 (Cell Signalling, Hitchin, Hertfordshire, UK). Anti-mouse or anti-rabbit IgG HRP linked secondary antibodies (Cell Signalling, Hitchin, Hertfordshire, UK) were used.
The PathScan^® Intracellular Signalling Array Kit (Cell Signalling, Hitchin, Hertfordshire, UK) was used according to manufacturer’s instructions to explore the phosphorylation status of proteins fundamental to signal transduction. Slide images were captured using Odyssey Fc System (Li-Cor Biosciences).
Immunoprecipitation for both FGFR3 WT and FGFR3-TACC3 (RT112) fusion were performed as in [48 (link)]. Immunoprecipitation of Topoisomerase IIα used 100 ug of starting material and was precipitated using the anti-TOPO IIα antibody (Cell Signalling, Hitchin, Hertfordshire, UK). Eluted protein was probed via immunoblotting using TOPO IIα and pS 1469 TOPO IIα antibodies (Cell Signaling, Hitchin, Hertfordshire, UK). Additional information can be found in the Supplementary Materials and Methods.