在转录调控、信号转导与染色质生物学研究中，获取高纯度、可重复的胞浆与细胞核组分是实验成败的分水岭。然而，把一块冷冻或纤维化的实体组织干净利落地拆成“胞质”和“胞核”两个组分，远比处理细胞样品困难得多——核膜易碎、基质黏连、细胞异质、冷冻损伤，每一步都可能让核蛋白“串场”或让胞浆蛋白“越界”。
1.结构脆弱——核孔复合体像一张精密的滤网，一旦机械力稍大或去垢剂稍强，核膜即碎；
2.物理纠缠——细胞骨架、核纤层层层包裹，不撕碎骨架就抽不出完整核，撕得太狠又连核一起破；
3.生物多样——不同细胞对渗透压、酶解、去垢剂的耐受差异巨大，没有“万能”裂解液；
4.动态交换——核-质之间持续进行能量依赖的主动运输，裂解时间一长，核内蛋白、RNA早已“漏网”到胞质。
培养的细胞多数具有完整的细胞膜，且为均匀的悬浮液。
而实体组织（尤其冷冻后）：
1.容易出现膜结构破裂，导致胞浆蛋白失控释放；
2.含有大量胞外基质和细胞碎片阻碍裂解并污染组分；
3.由于细胞异质性致使操作重复性差。
因此，用于培养细胞分离胞质胞核试剂盒应用于冷冻组织会导致严重的污染、较差的核完整性和不可靠的下游数据。
为了解决组织样品胞质胞核分离的难点，Invent用一套“膜选择性缓冲液 + 温和机械均质方法 + 离心柱纯化”的组合拳，给出了可规模化的解决方案（Minute™新鲜/冷冻组织胞质胞核分离试剂盒 Cat#NT-032 ），显著提高了冷冻或纤维化组织中的分离纯度与重复性，并可兼容多种下游实验。
核心优势
1.针对组织学特点：机械均质+去污剂缓冲液相结合，可在不破坏核膜前提下裂解细胞膜。即使在纤维组织或富含基质的组织中，也能有效。
2.冷冻样本专用：缓冲体系考虑了冷冻导致的膜完整性下降，显著减少交叉污染。
3.高纯度/高得率：离心柱+选择性裂解，经多组织验证，核/浆交叉污染极低。
4.广泛兼容下游应用：获得的组分适用于Western blot、蛋白质组学、RNA 分析、ChIP、酶活检测、qPCR 等。
5.可扩展与重复：支持从微小活检到大量组织的并行处理，人工操作差异小。
跨样品类型下游应用案例
神经退行性疾病和脑损伤模型通常依赖于评估转录因子和应激调节因子的核转位。Li等人[1]在大鼠卒中模型中证实了核Nrf 2激活和p62-Keap 1调节，而Yu等人[7]在亨廷顿病中观察到α-突触核蛋白介导的转录失调。类似地，Ryan等人[20]分析了脊髓损伤中的隔室特异性细胞因子信号传导。这些研究中的冷冻脑组织胞质胞核分离提成为了重大挑战，Invent NT-032大限度地减少了细胞质对核蛋白的污染，提高了下游分析的准确性，如Western印迹和RNA-seq。
Fig1.Nuclear/cytoplasmic fractionation of striatum samples from 12-month-oldWT, zQ175 and SNCAKO mice.图片来源：10.1186/s40478-022-01379-8
2.肝脏和代谢组织
肝脏在代谢和应激反应中的核心作用需要分离高纯度细胞核来研究转录因子，如SREBP-1c和Nrf 2。Zhou等人[4]和Weng等人[8]强调了NASH和缺血-再灌注模型中的隔室特异性NF-κB活化，而Zhu等人[15]和Schmidt等人[16]需要分离干净的核来研究肝脂肪变性和肝细胞癌。NT-032优化的裂解缓冲液和温和的组织匀浆步骤可有效处理纤维化肝组织和冷冻活检组织，为ChIP、蛋白质组学和免疫测定保留细胞核完整性。
Fig2.c-MYC is present in both the nucleus and cytosol of Sirt2+/+ HCC mice. 图片来源：10.3390/ijms241612618
心脏和肾脏组织是纤维性和异质的，使得核质分离困难。Bao 等人[5]和Luo 等人[6] 分别研究了左心室肥大和心脏毒性中信号蛋白的核转位，而Wang等人[9] 研究了肾纤维化中的IL-1β/TGF-β信号传导。NT-032有效地将细胞核与纤维化基质分离，从而能够准确评估核蛋白表达和翻译后修饰。
Fig3.The protein content of NF-κB in cytosolic and nuclear fractions was detected
by immunoblotting. GAPDH and Lamin B1 were used as loading controls.
图片来源：10.3390/ijms241612618
Kawauchi等人[28]评价了胺碘酮诱导的肠损伤期间的核反应，而Yang等人[10]需要完整的核部分来研究肠屏障应激模型中的转录调控。NT-032可有效地从肠道组织中去除细胞碎片，提升实验结果的可重复性，如核转录物的qPCR。
Fig4.Western blotting analysis of the nuclear and cytoplasmic β-catenin protein levels. 图片来源：10.1038/s41598-025-04680-3
冷冻组织对核分离提出了更大的挑战，细胞膜在冷冻过程中经常破裂，导致细胞质蛋白不受控制地释放和污染细胞核组分。Wahl等人[14]、Ju等人[27]和Mei等人[26]的研究分别利用存档组织研究衰老、分娩和肝内胆管癌中的核蛋白定位。NT-032专门设计了保持冷冻样品匀浆中的核完整性方案，从而实现可重复的下游测定，如质谱、染色质免疫沉淀和转录组学分析。
Fig5.Western blot analysis showing the abundance of S100A9 and p-S100A9 in the cytosolic and nuclear fractions of hAFs isolated from TL and TNL. 𝛼-tubulin and Lamin A/C were used as internal controls for cytoplasmic and nuclear protein. 图片来源：10.1002/advs.202414682
细胞核与细胞质的分离本质是在“完整性”与“纯度”间寻求平衡，Invent用“温和而坚定”的方式，把冷冻或纤维化组织里zui脆弱的核膜和zui坚固的基质拆开，让研究者不再在“纯度”与“完整性”之间做二选一。
1. Li Y, Wu J, Yu S, Zhu J, Zhou Y, Wang P, et al. Sestrin2 promotes angiogenesis to alleviate brain injury by activating Nrf2 through regulating the interaction between p62 and Keap1 following photothrombotic stroke in rats. Brain Res. 2020;146948.
2. Wang B, Huang X, Pan X, Zhang T, Hou C, Su WJ, et al. Minocycline prevents the depressive-like behavior through inhibiting the release of HMGB1 from microglia and neurons. Brain Behav Immun. 2020.
3. Song T, Guan X, Wang X, Qu S, Zhang S, Hui W, et al. Dynamic modulation of gut microbiota improves post‐myocardial infarct tissue repair in rats via butyric acid‐mediated histone deacetylase inhibition. FASEB J. 2021;35(3):e21385.
4. Zhou J, Zhao Y, Guo YJ, Zhao YS, Liu H, Ren J, et al. A rapid juvenile murine model of nonalcoholic steatohepatitis (NASH): Chronic intermittent hypoxia exacerbates Western diet-induced NASH. Life Sci. 2021;119403.
5. Bao J, Lu Y, She QY, Dou W, Tang R, Xu X, et al. MicroRNA-30 regulates left ventricular hypertrophy in chronic kidney disease. JCI Insight. 2021.
6. Luo LF, Qin LY, Wang JX, Guan P, Wang N, Ji ES. Astragaloside IV attenuates the myocardial injury caused by Adriamycin by inhibiting autophagy. Front Pharmacol. 2021;12:669782.
7. Yu D, Zarate N, Cuccu F, Yue JS, Brown TG, Nanclares TNM, et al. CK2 alpha prime and alpha-synuclein pathogenic functional interaction mediates inflammation and transcriptional dysregulation in Huntington’s disease. Neurodegener Dis. 2021;70:71.
8. Weng J, Wang X, Xu B, Li W. Augmenter of liver regeneration ameliorates ischemia -reperfusion injury in steatotic liver via inhibition of the TLR4/NF-κB pathway. Exp Ther Med. 2021;22(2):1–8.
9. Wang M, Wang L, Zhou Y, Feng X, Ye C, Wang C. Icariin attenuates renal fibrosis in chronic kidney disease by inhibiting interleukin‐1β/transforming growth factor‐β‐mediated activation of renal fibroblasts. Phytother Res. 2021.
10. Yang C, Luo P, Chen SJ, Deng ZC, Fu XL, Xu DN, et al. Resveratrol sustains intestinal barrier integrity, improves antioxidant capacity, and alleviates inflammation in the jejunum of ducks exposed to acute heat stress. Poult Sci. 2021;101459.
11. Imai S, Ohama M, Suzuki M, Katayanagi Y, Kayashima Y, Tezuka H. Short alkyl chain-length resorcinol olivetol protects against obesity with mitochondrial activation by Pgc-1α deacetylation. SSRN 3924601. 2021.
12. Wang D, Wang T, Li Z, Guo Y, Granato D. Green tea polyphenols upregulate the Nrf2 signaling pathway and suppress oxidative stress and inflammation markers in D-galactose-induced liver aging in mice. Front Nutr. 2022;130.
13. Lv L, Shu H, Mo X, Tian Y, Guo H, Sun HY. Activation of the Nrf2 antioxidant pathway by Longjing green tea polyphenols in mice livers. Nat Prod Commun. 2022;17(12):1934578X221139409.
14. Wahl D, Smith ME, McEntee CM, Cavalier AN, Osburn SC, Burke SD, et al. The reverse transcriptase inhibitor 3TC protects against age‐related cognitive dysfunction.Aging Cell. 2023; e13798.
15. Zhu Y, Lei L, Wang X, Jiang Q, Loor JJ, Kong F, et al. Low abundance of insulin-induced gene 1 contributes to SREBP-1c processing and hepatic steatosis in dairy cows with severe fatty liver. J Dairy Sci. 2023.
16. Schmidt AV, Monga SP, Prochownik EV, Goetzman ES. A novel transgenic mouse model implicates Sirt2 as a promoter of hepatocellular carcinoma. Int J Mol Sci.2023;24(16):12618.
17. AlTamimi JZ, AlFaris NA, Alshammari GM, Alagal RI, Aljabryn DH, Yahya MA.Esculeoside A alleviates reproductive toxicity in streptozotocin-diabetic rats by activating Nrf2 signaling. Saudi J Biol Sci. 2023;103780.
18. AlTamimi JZ, AlFaris NA, Alshammari GM, Alagal RI, Aljabryn DH, Yahya MA.Esculeoside A decreases diabetic cardiomyopathy in streptozotocin-treated rats by attenuating oxidative stress, inflammation, fibrosis, and apoptosis: Impressive role of Nrf2. Medicina. 2023;59(10):1830.
19. Al Jadani JM, Albadr NA, Alshammari GM, Almasri SA, Alfayez FF, Yahya MA. Esculeogenin A, a glycan from tomato, alleviates nonalcoholic fatty liver disease in rats through hypolipidemic, antioxidant, and anti-inflammatory effects. Nutrients. 2023;15(22):4755.
20. Ryan F, Francos-Quijorna I, Hernández-Mir G, Aquino C, Schlapbach R, Bradbury EJ, David S. Tlr4 deletion modulates cytokine and extracellular matrix expression in chronic spinal cord injury, leading to improved secondary damage and functional recovery. J Neurosci. 2023.
21. Alsabaani NA, Amawi K, Eleawa SM, Ibrahim WN, Aldhaban W, Alaraj AM, et al. Nrf-2-dependent antioxidant and anti-inflammatory effects underlie the protective effect of esculeoside A against retinal damage in streptozotocin-induced diabetic rats. Biomed Pharmacother. 2024;173:116461.
22. Di Veroli B, Bentanachs R, Roglans N, Alegret M, Giona L, Profumo E, et al. Sex differences affect the NRF2 signaling pathway in the early phase of liver steatosis: A high-fat-diet-fed rat model supplemented with liquid fructose. Cells. 2024;13(15):1247.
23. Nasser AA, Amawi K, Eleawa SM, Nabeel Ibrahim W, Aldhaban W, Alaraj AM, et al.Nrf-2-dependent antioxidant and anti-inflammatory effects underlie the protective effect of esculeoside A against retinal damage in streptozotocin-induced diabetic rats. Biomed Pharmacother. 2024.
24. Jawharji MT, Alshammari GM, Binobead MA, Albanyan NM, Al-Harbi LN, Yahya MA. Comparative efficacy of low-carbohydrate and ketogenic diets on diabetic retinopathy and oxidative stress in high-fat diet-induced diabetic rats. Nutrients. 2024;16(18):3074.
25. Zhang Z, Zhang J, Shi R, Xu T, Wang S, Tian J. Esculetin attenuates cerebral ischemia-reperfusion injury and protects neurons through Nrf2 activation in rats. Neuropharmacology. 2024.
26. Mei Q, Zhang Y, Li H, Ma W, Huang W, Wu Z, et al. Hepatic factor MANF drives hepatocytes reprogramming by detaining cytosolic CK19 in intrahepatic cholangiocarcinoma. Cell Death Differ. 2025;1–19.
27. Zhang F, Li MD, Pan F, Lei WJ, Xi Y, Ling LJ, et al. Nuclear translocation of S100A9 triggers senescence of human amnion fibroblasts by de‐repressing LINE1 via heterochromatin erosion at parturition. Adv Sci. 2025;2414682.
28. Kawauchi S, Horibe S, Tanaka T, Sasaki N, Kunimasa J, Rikitake Y. Disruption of small intestinal mucosal homeostasis in mice with amiodarone induced steatohepatitis. Sci Rep. 2025;15(1):23132.
站内热点