Figure 1

1A and 1B: Macrophages can be separated from carcinoma cells after microneedle collection. Cells collected from a MMTV-PyMTprimary tumor in a microneedle filled with matrigel and 25 nM EGF were magnetically separated using Mac-1 coated iron beads. Approximately 80% of the macrophages were removed from the sample, leaving only a 6% contamination of the carcinoma cell population by macrophages after a single separation step.

Figure 1C: Quantitative real-time PCR shows a decrease in the amount of Mac-1 mRNA in cells separated after collection from the primary tumor. Cells were collected and separated as described in Materials and Methods. After RNA purification from cells before and after separation, real-time PCR was run for the Mac-1 (macrophage marker) and keratin 18 (carcinoma cell marker) primers. Normalization was done using β-actin as reference gene. A decrease in expression of Mac-1 is seen in the cell-separated sample compared to that of the sample containing both cell types. This further confirms our ability to reduce the contaminating cells from our collected sample, so as to have a pure carcinoma cell population for gene discovery. The level of Keratin 18 expression shows a slight increase after separation.