Does anyone know how long a fixed sample can be stored in the dark at 4 degrees C without significant loss of quality? My samples are single cell suspensions from rat skeletal muscle and spleen. They are stained with 6 fluorochromes, fixed and permeabilized. Thanks in advance for your input.
For fixed samples, I would perform the analysis within a 2 week time frame, always stored in the dark at 4 ºC. As a rule of thumb, perform the analysis as soon as possible to be significant and representative.
Keep in mind that the most advisable thing would be to compare the differences between your fixed and unfixed cells, with your own stains.
Agree with Pablo completely. for fixed samples, it can store up to 2 weeks but analyze sooner the better.
As an addition to Pablo's reply: If you have used tandem fluorophores (e.g. Pe-Cy7, APC-780, PerCp-Cy5.5 etc), I would strongly advise reading no more than 48 hours after fixation. Once Cy7 starts to detach from Pe, etc., you will begin to collect false positive events. Best of luck!
Thank you all for sharing your expertise. I am using two tandem fluorochromes. I will do some experimentation to see how the flow data changes with age of the sample.
Fixed cell can be stored in complete dark for two days without affecting the result as per my experience
As per my experience, 48 hrs time is max for capturing the fixed samples, but you will see population of dead cells as well, so in my opinion, it is affecting our results. So its always best to capture the samples as soon as they are ready. It gives best results as i have checked the results with the age of cells.
I would rather ask one question here with all of you. Please suggest the facs buffer you use for washing the cells and for storage of cells.
- Badges
- Science topic
- Similar topics
- Biological Science
- Immunology