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抗体保存注意事项（Handling Antibodies）![搜索无重复]
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首先要树立的一个观念是“抗体可一点儿都不脆弱，很强悍的”。 “Antibodies have a compact three-domain structure that has evolved to retain activity in most biological settings. One practical advantage of such compact and stable protein domain is that they are resistant to a broad range of mildly denaturing conditions, making long-term storage of antibodies relatively easy. Storage buffers seldom need to be supplemented with glycerol or other stabilizing compounds that are commonly added to help maintain the activity of proteins purified in the lab.” 经常使用的抗体可以4℃保存；长期保存可以置于-20℃。 “Working solutions of antibodies are conveniently stored at 4℃. At this temperature they are stable for months to years without loss of activity. For long-term storage, antibodies can be kept conveniently at -20℃ in the serum, tissue culture supernatant, or ascitic fluid in which they are collected. Lower temperatures will not hurt the antibody activities, but there is no significant advantage in lower-temperature storage.” 也有例外，不是所有抗体在4℃都稳定，有些得在常温保存。 “The only group of antibodies that should not be stored at 4℃ are the so-called cryoproteins, which are sensitive to low-temperature storage, where they precipitate. Some mouse antibodies of the IgG3 subclass may fit these characteristics. If antibodies precipitate at 4℃ over time, they should be kept at room temperature with addition of sodium azide.” 尽管抗体很稳定，还是要避免反复冻融，因为它可能造成抗体活力的丧失。原因是...... “Antibody solutions should not be frozen and thawed repeatedly, because this can lead to denaturation of antibodies that may lead to unwanted protein-protein aggregation. Aggregation of antibodies can cause the loss of activity due to steric interference of the antigen combining site or by generating insoluble material that is lost during centrifugation or filtration.” 抗体到底可以保存多长时间？什么？十年？ “Antibodies are remarkably stable. If tightly sealed to avoid evaporation, they can be stored at 4℃ for months with no loss of activity. Antibodies stored at -20℃ have been shown to keep their activity for decades.” 这么强悍！那，抗体到底怕什么？ “The only problem commonly encountered in storing antibodies is contamination of these solutions with bacteria or fungi. This can be prevented by addition of antimicrobial agents; for example, by adding sodium azide to 0.02% or, as an alternative, Merthiolate to 0.01%.” 何时需要添加叠氮化纳？什么时候又不能添加？ “Although useful and effective for most settings, sodium azide is not appropriate in some assays, because it blocks the cytochrome electron transport system and therefore is toxic to most organisms. In addition, the primary amine on the azide would attack the stability of the antibody's interaction with conjugate or matrix in coupling methods. If sodium azide is already present in a solution, it can be removed by dialysis or gel filtration.” 自己纯化的抗体，保存的时候要注意什么？ Purified preparations of antibodies are stable in most commonly used buffers. The pH should be kept near neutral; pHs between 7 and 8 show no deleterious effects, even over many years of storage. Salt concentrations between 0 and 150 mM are suitable for most applications, but little harm is caused with long-term storage in salt concentrations up to 500 mM. If no other indications are known, we recommend long-term storage in PBS or 50 mM Tris, pH8.0. If there is no reason to avoid the use of sodium azide, add it to 0.02%. Aliquot samples of the purified antibodies into convenient volumes and store frozen at -20℃. Solutions of purified antibodies should be stored relatively high concentrations(i.e., ≥ 1 mg/ml) at neutral pH. Concetrations up to 10 mg/ml are commonly used. Antibodies at lower concentrations should be concentrated prior to freezing. http://blog.bioon.com/user1/2081/archives/2009/231984.shtml [ Last edited by amisking on 2009-5-27 at 19:15 ] |
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