Heparin acrylic

The transcriptional effects of FGF signaling in the expression of Uhrf genes were studied by analyzing the effects of the local administration of FGF2 (diluted at 0.5 mg/ml, Peprotech), and the FGF inhibitor SU5402 (diluted at 4 mg/ml, Calbiochem), using heparin acrylic (Sigma) or ion exchange (AG1-X2, Bio-Rad) microbeads as described^{21 (link)}.

Bead implantations were performed according to published methods6 (link) with the following modifications: Heparin acrylic (Sigma) and Affigel-Blue (Bio-Rad) beads were size-selected at 250 μm. Affigel-Blue beads were soaked in SHH-N protein (human origin, R&D Systems) diluted to 1 mg ml⁻¹ in PBS with 0.1% BSA. Heparin beads were soaked in cyclopamine (Toronto Research Chemicals) diluted to 4 mg ml⁻¹ in DMSO. Control experiments were performed using the respective vehicle-only beads. Stage 30 embryos were dissected from their egg cases, anaesthetized and the bead was surgically implanted into the posterior region of the pelvic fin using tungsten needles. For all bead implants, L. erinacea embryos were sorted by sex and then randomly allocated into either control or experimental treatment groups. After manipulation, animals were returned to their tank for 24-h before tissue-harvesting for qRT–PCR, or they were allowed to develop for an additional ∼10 weeks for phenotypic analysis.