Tumor-Associated Schwann Cell Remodeling under Metabolic Stress via Lactate Sensing Orchestrates

Biotin pull-down assay
The Magic Dynabeads MyOne Streptavidin T1 were pre-incubated with free biotin or biotinylated lactate at room temperature for 30 minutes to saturate the bead surface with biotin. Subsequently, the pre-treated beads were mixed with the cell lysate of interest and incubated overnight at 4°C with rotation. The beads were then washed four times with PBS buffer to remove non-specifically bound molecules. After washing, the proteins were separated from the beads, and the presence and expression levels of the target protein were detected using techniques such as Western blotting or proteomics.

生物素下拉试验（Biotin pull-down assay）
将Dynabeads MyOne链霉亲和素T1磁珠（Magic Dynabeads MyOne Streptavidin T1）与游离生物素或生物素修饰的乳酸盐于室温下预孵育30分钟，以使磁珠表面被生物素充分饱和。随后，将预处理后的磁珠与目标细胞裂解液混合，于4℃条件下旋转孵育过夜。之后使用PBS缓冲液洗涤磁珠四次，以去除非特异性结合的分子。洗涤完成后，将结合的蛋白从磁珠上解离分离，并通过蛋白质印迹（Western blotting）或蛋白质组学等技术检测目标蛋白的存在情况与表达水平。