Regardless of your specific purpose, knowing how to use a flow cytometry fixation buffer is an important skill to have as a scientist. Properly preserving cells in solution with the right chemical balance will make the difference between analysis-ready samples and those that need to be thrown out because of damage during the cell preservation process. This article will explain why it’s so important to use the right flow cytometry fixation buffer and how to select the best one for your own purposes as well as give you some tips on how to get the most out of your lab’s fixative.
When do you need to fix cells?
There are two main reasons to fix cells: First, if you want to do DNA or RNA extraction. Second, if you want to analyze protein expression. When deciding whether it’s necessary, here are two questions you should ask yourself: (1) Are you interested in extracting or amplifying nucleic acids? and (2) Do your antibodies detect intracellular proteins? If either of these is true, then fixing your cells first is essential. The flow cytometry fixation buffer contains formaldehyde, which denatures cell proteins so that they can be easily seen by antibodies and other detection methods. You should also consider what type of downstream analysis you need to perform on your sample: will it be a Western blot or ELISA? The solution may not need fixed before proceeding with this type of analysis.
What is a flow cytometry fixation buffer?
Flow cytometry can be used to analyze individual cells. However, when cells are separated from other types of matter, they are exposed to buffer. To ensure your flow cytometry experiment gives you accurate results, it is crucial that you choose a flow cytometry fixation buffer that is compatible with your specific cell type. This post will help you learn more about flow cytometers and how fixatives differ based on cell type.
How does it work?
The flow cytometry fixation buffer uses sodium azide as a fixative, which reduces nucleic acids by forming cross-links with phosphate groups in RNA and DNA. It may take several hours to days for all degradation products to be degraded into inert materials, depending on sample type and fixation time. Although it is best used when cells are freshly harvested from tissue or whole organisms, it can also be used on stored samples.
Is there any downside?
Fixatives are used by cytotechnologists to preserve cells and cell components (like organelles). In other words, fixatives stop cells from continuing to live and maintain their current state. Therefore, flow cytometers can’t analyze dead cells very well. The primary function of a fixation buffer is to prevent damage caused by fixation. While there are some variations depending on what you want your sample for, you’ll almost always use one of these three buffers: Bouin solution, Carnoy solution or Zenker solution. Each type provides its own benefits and drawbacks, but they all serve one purpose—to maintain your samples in their current condition until you use them in your experiment.
Which Flow Cytometry Fixation Buffer should I use?
There are various types of flow cytometry fixation buffers that can be used depending on what you are trying to accomplish. Try out each buffer, and make notes about how your samples reacted. This will help you identify which buffer works best for your needs and ensure future success with flow cytometry. Here is an example of a table you could use to keep track of different solutions
