破膜剂亦称通透液，通常可以通过使用有机溶剂例如甲醇、丙酮等，或者使用去垢剂例如Triton X-100、Saponin等来实现对于细胞膜的通透。
甲醇或丙酮等有机溶剂，一方面可以溶解细胞膜和核膜从而充分暴露细胞浆和细胞核中的目的蛋白等，另一方面也可以使细胞内的蛋白变性起到固定的作用。其优点是操作比较简单，一步处理可以同时实现固定和通透，缺点是膜蛋白也会被溶解掉并且有些蛋白变性后不利于后续的检测，因此有机溶剂使用相对较少，仅用于一些要求比较粗糙的检测。Triton X-100是常用的通透试剂，可以通透细胞膜和核膜，作用原理是非特异性地溶解细胞膜，缺点是不利于膜蛋白的检测，但当使用多聚甲醛进行交联和固定后，相当一部分膜蛋白会被交联固定，从而不会被Triton X-100溶解，因此后续仍然能检测到。Saponin可以特异性地溶解细胞膜中的胆固醇，从而实现在细胞膜上选择性地打孔，其优点是适合于细胞膜蛋白的检测，特别适合通过流式细胞仪检测细胞膜上的标志性蛋白，缺点是对于一些胆固醇含量低的细胞通透效果差，通透效果弱于Triton X-100和有机溶剂，并且不能通透胆固醇含量很低的核膜和线粒体膜。对于凝集素 (lectin)的检测，含Triton X-100等非特异性去垢剂的通透液的效果显著优于主要含Saponin的通透液。
PBS-Triton溶液 (破膜剂, 0.1%)亦称为免疫染色通透液，主要由PBS磷酸盐缓冲液 (0.01M, pH 7.4)、0.1%Triton X-100组成，可以用于免疫染色等多种原位检测时细胞样品、冰冻或石蜡切片的通透处理，有助于暴露抗原、核酸等作用靶点，使抗体、探针或标记物等更容易进入细胞内，从而确保染色等的检测效果。本产品通透能力强，推荐用于各种常规的免疫荧光、免疫组化、免疫细胞化学、流式检测等中细胞的通透。本产品为即用型工作液，可以直接使用，无需稀释。该试剂仅用于科研领域，不适用于临床诊断或其他用途。
操作步骤(仅供参考)：
1、对于切片，在固定、洗涤结束后，每样滴50-100μl PBS-Triton溶液，也可以在染色缸中完全浸没进行通透。
对于细胞样品，在固定、洗涤结束后，按照六孔板每孔加入1毫升PBS-Triton溶液，其它多孔板参考该比例加入。
3、对于其它样品，加入PBS-Triton溶液的量以充分盖住样品为准。
4、通常在PBS-Triton溶液中室温孵育5-10分钟，即可完成通透。对于较难通透的样品或者要求通透特别充分的情况，可以在室温孵育10-30分钟。
注意事项：
1、对于通透效果要求不高的情况，也可以使用PBS-Triton溶液 (破膜剂, 0.05%)或免疫染色洗涤液。
2、对于需要检测细胞膜蛋白的情况推荐使用PBS-Saponin溶液 (破膜剂, 0.5%)。
3、对于通透要求比较高的情况，可以使用PBS-Triton溶液 (破膜剂, 0.3%)或PBS-Triton溶液 (破膜剂, 0.5%)。
4、对于凝集素 (lectin)的检测推荐使用本产品或PBS-Triton溶液 (破膜剂, 0.3%)。
5、为了您的安全和健康，请穿实验服并戴一次性手套操作。
Permeabilization Reagent, also known as permeabilization solution, usually achieves cell membrane permeabilization by using organic solvents such as methanol and acetone, or detergents such as Triton X-100 and Saponin.
Organic solvents like methanol or acetone can, on the one hand, dissolve cell membranes and nuclear membranes to fully expose target proteins in the cytoplasm and nucleus; on the other hand, they can denature intracellular proteins to play a fixation role. Their advantage is simple operation-one-step treatment can achieve both fixation and permeabilization simultaneously. However, their disadvantages are that membrane proteins will also be dissolved, and the denaturation of some proteins is not conducive to subsequent detection. Therefore, organic solvents are relatively less used and only applicable to some assays with rough requirements. Triton X-100 is a commonly used permeabilization reagent that can permeabilize cell membranes and nuclear membranes through the mechanism of non-specifically dissolving cell membranes. A drawback is that it is not conducive to membrane protein detection. Nevertheless, after cross-linking and fixation with paraformaldehyde, a considerable portion of membrane proteins will be cross-linked and immobilized, thus avoiding dissolution by Triton X-100 and enabling subsequent detection. Saponin can specifically dissolve cholesterol in cell membranes, thereby selectively creating pores on the cell membrane. Its advantages are suitability for membrane protein detection, especially for the detection of cell membrane marker proteins via flow cytometry. The disadvantages are poor permeabilization effect on cells with low cholesterol content, weaker permeabilization capability compared with Triton X-100 and organic solvents, and inability to permeabilize nuclear membranes and mitochondrial membranes with extremely low cholesterol levels. For lectin detection, permeabilization solutions containing non-specific detergents such as Triton X-100 exhibit significantly better performance than those mainly composed of Saponin.
PBS-Triton Solution (Permeabilization Reagent, 0.1%), also referred to as immunostaining permeabilization solution, is mainly composed of 0.01M PBS (phosphate-buffered saline, pH 7.4) and 0.1% Triton X-100. It can be used for the permeabilization treatment of cell samples, frozen or paraffin sections in various in situ detection methods such as immunostaining. It helps expose action targets including antigens and nucleic acids, facilitating the entry of antibodies, probes or labels into cells, thereby ensuring the detection effectiveness of staining and other assays. This product has strong permeabilization ability and is recommended for cell permeabilization in various routine assays such as immunofluorescence, immunohistochemistry, immunocytochemistry and flow cytometry. This product is a ready-to-use working solution and can be applied directly without dilution. The reagent is for research use only and not intended for clinical diagnosis or other purposes.
Operating Procedures (for reference only)
1. For sections: After completion of fixation and washing, add 50-100 μl of PBS-Triton Solution dropwise to each sample. Alternatively, the sections can be completely immersed in a staining jar for permeabilization.
2. For cell samples: After completion of fixation and washing, add 1 ml of PBS-Triton Solution to each well of a 6-well plate. For other multi-well plates, add the solution at a proportional volume.
3. For other samples: Add an amount of PBS-Triton Solution sufficient to completely cover the samples.
4. Generally, incubate the samples in PBS-Triton Solution at room temperature for 5-10 minutes to complete permeabilization. For samples that are difficult to permeabilize or when thorough permeabilization is required, the incubation time can be extended to 10-30 minutes at room temperature.
Precautions
1. For applications with low requirements for permeabilization effect, PBS-Triton Solution (Permeabilization Reagent, 0.05%) or immunostaining washing solution can also be used.
2. For membrane protein detection, PBS-Saponin Solution (Permeabilization Reagent, 0.5%) is recommended.
3. For applications requiring high permeabilization efficiency, PBS-Triton Solution (Permeabilization Reagent, 0.3%) or PBS-Triton Solution (Permeabilization Reagent, 0.5%) can be selected.
4. For lectin detection, this product or PBS-Triton Solution (Permeabilization Reagent, 0.3%) is recommended.
5. For your safety and health, please wear a lab coat and disposable gloves during operation.
ZJ26F0737882
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