Ht15 1kt kit

Muscles were harvested and used for cryosectioning and immunofluorescence staining as described earlier⁴⁴ . TA muscles transplanted with GFP positive MuSCs were fixed in 2% PFA for 3 h at room temperature (RT) and a sucrose gradient was applied for three consecutive nights (10% → 20% → 30% sucrose in ddH₂O) before freezing in liquid nitrogen. Sections were not additionally fixed for eMHC staining. Cross sections from transplanted muscles were additionally fixed in 2% PFA for 30 min before immunofluorescence staining. Afterwards, antigen retrieval was performed in a decloaking chamber (Biocare Medical) for 10 min at 95 °C using 1 x HIER antigen retrieval buffer (Abcam) before the staining was proceeded. Primary and secondary antibodies are listed in Table 1 and 2 in the supplemental information. Microscopic images were acquired on an upright Axio imager microscope from Zeiss with a 20x objective.
Masson Trichrome staining was performed with the HT15-1KT kit (Sigma Aldrich) according to the manufacturer’s protocol and sections were analyzed using Qupath 0.2.3^{45 (link)}.

Heart sections were fixed overnight in 10% formalin (Sigma-Aldrich, F5554) and then in ethanol (Fisher Scientific, 10437341, 70% v/v). The samples were embedded in paraffin, and staining was performed using a Masson’s trichrome kit (Sigma, HT15-1KTKIT) according to the manufacturer’s protocol. Images for analysis were acquired using a Zeiss Axio scan digital slide scanner. The total global collagen was quantified using the color threshold function in the Fiji software (version 2.0.0) to identify fibrotic regions and compared to the whole myocardial area. Perivascular fibrosis was assessed as the degree of fibrosis surrounding the vessels compared to the vessel size. The five largest vessels in each section were analyzed, and an average was taken for each animal.