Flow Cytometry Research Core: Sorting Guideline

SORTING GUIDELINES

1. If you have less than 30 million cells to sort, we can use the FacStar Instrument and the concentration of your cells should be 3-4 million cells/ml. This is the low speed sorter.

2. If you have more than 30 million cells to sort, we can use the MoFlo and the concentration of your cells should be 10- 20 million/ml. This is a high-speed sorter.

3. All your sorts need to have a negative control. Please let us know if the cells will need to be kept cold during the sort.

4. Allow 30 minutes additional time for cleaning the instrument for each sample to be sorted.

5. All sorts are charged an additional one (1) hour for instrument set-up time.

6. The sorts need to be scheduled at least one (1) week in advance.

7. Bring about 50 ml of media into which to sort the cells. This media needs to have antibiotics and antimycotics to help maintain sterility, and should contain 10-20% serum.

8.     If the cells need serum to be kept alive, the sample to be sorted should not exceed 2% serum. Serum can make the cells sticky and clog the instrument. The media into which the cells are to be sorted may have a higher concentration of serum.

9. All biohazardous sorts and protocols need to be pre-approved by Dr. Celsa Spina at ext. 7189.

10. Cloning dishes need to be pre-filled with 200 microliters of media per well.

11.  All UCSD and outside researchers need a PO # for sorting and cloning.

12.  If the amount or the number of samples to sort changes, please notify the Flow Lab ASAP so that we can try to arrange our schedule appropriately. Please be aware that it may not always be possible to do more cells or samples than what was originally scheduled.

13.  The final cell yield is dependent upon many factors: The starting Percent of the target population, the total number of live and dead cells in the sample, cell stickiness and the instrument’s efficiency. If you need a specific number of cells after the sort, it is recommended that you bring more of your starting sample to compensate for yield variability.