Fallesen Todd, Brumm A Sophie
Crick Advanced Light Microscopy, The Francis Crick Institute, London, UK.
Human Embryo and Stem Cell Laboratory, The Francis Crick Institute, NW1 1AT London, UK.
STAR Protoc. 2025 Jun 20;6(2):103849. doi: 10.1016/j.xpro.2025.103849. Epub 2025 May 28.
The transforming growth factor β (TGF-β) signaling superfamily includes NODAL and bone morphogenetic protein (BMP) signaling, which lead to the phosphorylation of different SMAD proteins and regulate key developmental events. Here, we present a protocol for immunofluorescence detection of phosphorylated SMAD proteins combined with other transcription factors in pre-implantation human embryos. We describe steps for segmenting the nuclei in human blastocysts and quantifying their immunofluorescence intensity. This protocol can be adapted to investigate TGF-β superfamily signaling activity in other mammalian embryos or in vitro models of their development. For complete details on the use and execution of this protocol, please refer to Brumm et al..
转化生长因子β（TGF-β）信号超家族包括NODAL和骨形态发生蛋白（BMP）信号，它们导致不同SMAD蛋白的磷酸化并调节关键的发育事件。在此，我们展示了一种用于在植入前人类胚胎中免疫荧光检测磷酸化SMAD蛋白并结合其他转录因子的方案。我们描述了对人类囊胚中的细胞核进行分割并量化其免疫荧光强度的步骤。该方案可用于研究其他哺乳动物胚胎中TGF-β超家族信号活性或其发育的体外模型。有关此方案的使用和执行的完整详细信息，请参考Brumm等人的研究。
