请教淋巴细胞活化的抗体配制

fangming0411

请教：做淋巴细胞活化和共培养用，请问CD40L和anti CD3 mAb的怎么配制？用什么溶剂？母液及工作液如何保存？

niwanmao

抗体买来就是液体的啊，为什么还要另外配制呢？
你只要按照所需的抗体浓度，加入相应的量，使最终浓度达到你需求即可：http://www.ebioscience.com/media ... vation-in-vitro.pdf

此外，我发现还有专门的CD3/CD28 共刺激beads，Life公司出的，这里有一篇有关两种刺激方法之间的对比文章：http://www.translational-medicine.com/content/8/1/104

fangming0411

50ug一次用不完，不是说反复冻融不好吗？部分母液冻存，部分配成工作液现取用不是更方便吗？

niwanmao

fangming0411 发表于 2014-3-20 10:52
50ug一次用不完，不是说反复冻融不好吗？部分母液冻存，部分配成工作液现取用不是更方便吗？ ...

那么其实就是指抗体的保存了，关于这方面，AvesLab网页上有一段非常清楚的指导，在此转来与朋友们共享：

Antibody Storage Recommendations

Overview

Since antibodies are designed to retain biological activities in vivo over periods of many weeks at body temperatures in the 41-42˚C range, they are quite thermostable. Other issues are:

Freezing. The biggest issue with regards to antibody stability is freezing. All antibodies are labile to one degree or another by freezing conditions, presumably due to the damage incurred during ice crystal formation. In this regard, chicken antibodies, like rabbit and mouse antibodies, will lose some biological activity (typically about 50%) upon freezing. Therefore, it is essential that multiple freeze-thaw cycles be avoided at all cost.

Light. All antibodies (and most proteins) are also labile to light. It is essential, therefore, to keep them in the dark when storing antibodies for long periods of time.

Contamination. Similarly, all proteins, including antibodies, are labile to degradative enzymes release by bacteria and other microorganisms, especially proteinases. It is essential, therefore, to keep aliquots of antibodies biologically sterile.

Short-term Storage of Antibodies

For short-term (up to 1 year), we recommend storage at –4˚C in the dark. When removing aliquots, it is essential that sterile techniques be followed.

Long-term Storage of Antibodies

For longer-term storage (from 1-5 years), we recommend diluting the antibodies 1:1 with glycerol, re-filter sterilizing them, and then storing them at -20°C in the dark. Again when removing aliquots, it is essential that sterile techniques be followed.

Very Long-term Storage of Antibodies

For very long-term storage (more than 5 years), we recommend storage at –80˚C. Although there will be some loss of biological activity accompanying the freezing (typically about 50%), the remaining biological activity will be stable indefinitely.

Sodium Azide

We routinely add sodium azide as an antimicrobial agent to all preparations of antibody in order to discourage the growth of bacteria and fungi. If you do NOT want us to add sodium azide to your preparations for any reason, please let us know ahead of time. We can substitute thimerosal or not add any antimicrobial agents at all (afterall, the antibodies will be filter-sterilized). If you want to get rid of sodium azide, we recommend simple dialysis.