Is it possible to enrich more than one cell type from the same sample sequentially? 

Yes, this is possible if you are using a positive selection strategy. To isolate several different cell types from one sample separately, do not add the different MACS MicroBeads at the same time. Instead, add one type first, perform the separation, and then add the second type of MicroBeads to the negative fraction, and so on.

The sequence in which you perform the separations is an important factor. Here are some general guidelines to follow:

  1. Isolate monocytes or other sticky cells first, because they tend to bind non-specifically to antibodies and negatively influence the purity of other separations.
  2. Start first with the least frequent cell type, and proceed to the most frequent cell type last. This order ensures the best recovery of the rare cell types, because some cells will be lost in each step.
  3. To ensure good purities, include an additional depletion step after 3 or 4 separations, to remove the cells that bound non-specifically to the MicroBeads, as well as any magnetically labeled cells that ran through the column during the first separations. Optimally, this depletion step would use an LD column, or a slower autoMACS Pro Separator program than the one used for the following separations.

Keep in mind that a sequential separation will become more difficult as the number of steps increases. It might be necessary to perform some tests to determine the optimal strategy for your particular need.