EDTA细胞消化液 (0.02%, 含酚红)由PBS、EDTA (乙二胺四乙酸)、酚红、去离子水等配制而成，经无菌处理，不含胰蛋白酶，在细胞消化中的作用原理主要基于其螯合作用，通过选择性结合细胞间和细胞-基质连接处的二价阳离子 (如Ca²+、Mg²+)，破坏细胞黏附系统的稳定性，对细胞损伤较小，能有效地使贴壁细胞与培养瓶或培养皿表面脱离而达到分离细胞目的。酚红可动态监测pH变化，直观判断消化状态；调控氧化应激，增强部分细胞贴壁率。该试剂仅用于科研领域，不适用于临床诊断或其他用途。
其特点是：
1、作用温和；
2、对细胞的损伤和破坏极小，不影响细胞生物学特性，是肿瘤细胞的极好细胞脱壁方法；
3、可以在血清存在的情况下进行消化。消化后的细胞可进行传代培养，亦可用于提取核蛋白和胞浆蛋白、Western Blot、免疫共沉淀等实验，通常室温下3-10min即可消化下大多数贴壁细胞，推荐用于原代干细胞，而不建议用于胚胎干细胞的消化。
自备材料：
PBS、培养液、倒置显微镜、离心机
操作步骤 (仅供参考)：
贴壁细胞的消化
1、吸除培养液，用无菌PBS、培养液洗涤细胞1次。
2、加入少量EDTA细胞消化液 (0.02%, 含酚红)，略盖过细胞即可 (一般按细胞的有效体积的10倍添加)。
3、室温放置2-10min，如置于37℃脱壁反应会加速，直到细胞完全脱壁，不同的细胞消化时间有所不同；亦可显微镜下观察，细胞明显收缩，并且肉眼观察培养器皿底部发现细胞的形态发生明显的变化或者用枪吹打细胞发现细胞刚好可以被吹打下来，吸除消化液。
4、加入细胞培养液或5倍体积PBS缓冲液终止反应。如果发现消化不足，则加入本品重新消化。
5、1000-2000g离心3-5min，沉淀细胞，弃上清，尽量去除细胞消化液，加入含血清的完全培养液重新悬浮细胞，即可用于后续实验。
注意事项：
1、尽量减少反复冻融的次数，以免失效。
2、试剂开封后请尽快使用，以防影响后续实验效果。
3、在使用细胞消化液的过程中，要特别注意避免消化液被细菌污染。
4、细胞消化液消化细胞时间不宜过长，否则细胞铺板后生长状况会较差。
5、37℃时作用速度比4℃快6-8倍，但细胞死亡率增加。
6、为了您的安全和健康，请穿实验服并戴一次性手套操作。
EDTA Cell Detachment Solution (0.02%, Phenol Red-Containing) is formulated with PBS, EDTA (Ethylenediaminetetraacetic acid), phenol red, and deionized water. It undergoes sterile processing and contains no trypsin. Its mechanism of action in cell dissociation is mainly based on chelation: by selectively binding divalent cations (e.g., Ca²⁺, Mg²⁺) at intercellular and cell-matrix junctions, it disrupts the stability of the cell adhesion system. This solution causes minimal cell damage and effectively detaches adherent cells from the surface of culture flasks or dishes to achieve cell separation. Phenol red can dynamically monitor pH changes for intuitive judgment of digestion status; it also regulates oxidative stress and enhances the adherence rate of certain cell types. For research use only. Not intended for clinical diagnosis or other purposes.
Features:
1. Mild in action;
2. Causes minimal damage and disruption to cells without affecting their biological properties, making it an excellent method for detaching tumor cells;
3. Can perform cell detachment in the presence of serum. Digested cells can be used for subculture, and also for experiments such as nuclear and cytoplasmic protein extraction, Western Blot, and Co-Immunoprecipitation (Co-IP). Under room temperature, most adherent cells can be detached within 3-10 minutes. This solution is recommended for primary stem cells but not for embryonic stem cell dissociation.
Materials to Be Prepared by Users
PBS, Culture medium, Inverted microscope, Centrifuge
Operating Procedures (For Reference Only)
Detachment of Adherent Cells
1. Aspirate the culture medium and wash the cells once with sterile PBS or culture medium.
2. Add a small volume of EDTA Cell Detachment Solution (0.02%, Phenol Red-Containing), just enough to cover the cells (generally, add 10 times the effective volume of the cells).
3. Incubate at room temperature for 2-10 minutes; incubation at 37°C will accelerate the detachment reaction. Continue the process until the cells are completely detached. Note that the digestion time varies for different cell types. Alternatively, observe the cells under a microscope: once the cells shrink significantly, or the cell morphology at the bottom of the culture vessel changes obviously by visual inspection, or the cells can be just blown off by pipetting, aspirate the detachment solution immediately.
4. Add cell culture medium or 5 volumes of PBS buffer to terminate the reaction. If insufficient detachment is observed, add this solution again for re-digestion.
5. Centrifuge at 1000-2000×g for 3-5 minutes to pellet the cells. Discard the supernatant, remove as much of the cell detachment solution as possible, and resuspend the cells in complete medium containing serum for subsequent experiments.
Precautions
1. Minimize the number of repeated freeze-thaw cycles to avoid inactivation of the reagent.
2. Use the reagent as soon as possible after opening to prevent adverse effects on subsequent experimental results.
3. During the use of the cell detachment solution, take special care to avoid bacterial contamination of the solution.
4. Do not prolong the cell digestion time excessively, otherwise the cell growth status will be poor after plating.
5. The reaction rate at 37°C is 6-8 times faster than that at 4°C, but the cell mortality rate will increase accordingly.
6. For your safety and health, wear a lab coat and disposable gloves during operation.
ZJ26F0433940
危险品化学品经营许可证（带存储）