Translatome Hypoxia-Acidosis, TMT-pulse SILAC

Cells were grown in light (R0K0) SILAC media (AthenaES) for 7 days, they were then incubated in light media for 24 hours in respective condition (normoxia neutral (NN) pH: 7.4; hypoxia neutral (HN)1% O2, pH: 7.4; hypoxia acidosis (HA), 1% O2, pH=6) and pulsed with heavy (R10K8) SILAC media (AthenaES) for 16 hr (TMT-pSILAC) following treatment.

将细胞于轻同位素（R0K0）稳定同位素标记细胞培养（Stable Isotope Labeling by Amino acids in Cell culture, SILAC）培养基（AthenaES）中培养7天；随后将细胞置于对应条件的轻同位素培养基中孵育24小时，各组条件分别为：常氧中性（normoxia neutral, NN）组pH值7.4；低氧中性（hypoxia neutral, HN）组为1% O₂、pH 7.4；低氧酸中毒（hypoxia acidosis, HA）组为1% O₂、pH=6。处理完成后，使用重同位素（R10K8）SILAC培养基（AthenaES）对细胞进行16小时的脉冲标记，该实验采用串联质量标签-稳定同位素标记细胞培养联用技术（TMT-pSILAC）策略。