Vectastain elite abc rabbit kit

The ApopTag Plus Fluorescein In Situ Apoptosis Detection Kit (EMD Millipore, Bedford, MA, USA) was used as per manufacturer's instructions to quantify the number of apoptotic cells in the tibial GP of 7-week-old mice. Proliferating cell nuclear antigen (PCNA) immunohistochemistry (IHC) was performed using a 1:4000 dilution of an anti-PCNA antibody (Abcam, Cambridge, UK) followed by the Vectastain elite ABC rabbit kit (Vector Laboratories, Peterborough, UK) on paraffin-embedded sections from 3-week-old mice (when rates of proliferation were likely to be highest). Sections were counterstained by Haematoxylin and viewed using a Zeiss AxioImager brightfield microscope. The Haematoxylin-DAB colour deconvolution plugin was used in Fiji to calculate the percentage of PCNA+ve cells in the proliferating zone of the GP of each sample.

Proliferating cell nuclear antigen (PCNA) immunohistochemistry (IHC) was performed using a 1:4000 dilution of an anti-PCNA antibody (Abcam) followed by the Vectastain elite ABC rabbit kit (Vector Labs, Peterborough, UK) on paraffin-embedded sections of proximal right tibiae. Sections were counterstained by haematoxylin and viewed using a Zeiss AxioImager brightfield microscope. The haematoxylin–DAB colour deconvolution plugin was used within Fiji to calculate the percentage of PCNA +ve cells in the proliferating zone of the GP of each sample (Schindelin et al. 2012 (link)).