These data confirm that genomic in situ hybridization with Alu probes is a specific method to identify human cells, and efficiently stains both cell lineages and low-passage, heterogeneous primary cell populations.
Heterogeneity is a hallmark of cell biology, and is strongly evident in primary cell populations isolated from the same tissue [2].
Currently, we are routinely using primary cell populations to validate new questions and reagents that subsequently will be addressed in organ culture systems.
As culture conditions become better defined, either to support the specific expansion of homogeneous primary cell populations or to support the survival and expansion of complex primary cell mixtures derived from disrupted mucosal surfaces, so the utility of primary cell populations should increase.
The experiments with primary cell populations are at least one step further removed from reality than tissue pieces in organ culture but primary cells do provide biological value relative to standard immortalized epithelial cell lines.
This method allows for highly efficient transfection of DNA or siRNA into isolated primary cell populations or within mixed cell suspensions as well as into other cell populations that are considered difficult to transfect.
In several cases, mixed fibroblast/epithelial primary cell populations expanded concurrently in adjacent but non-overlapping zones on the same transwell membrane and preferential attachment of C. albicans colonies to fibroblasts was consistently observed.
Methods describing the manipulation of tissue pieces on collagen supports and the preparation of primary cell populations on transwell membranes (Cell Culture Inserts [0.4 µm pore size, polyethylene tetraphthalate membranes without additional ECM coating] BD Biosciences, Bedford MA) have already been published [24], [42].
Furthermore, on a practical level, because not all tissue pieces are of sufficient quality for organ culture experiments we have found that tissue pieces with limited surface epithelium can still be put to good use in the derivation of primary cell populations [24], [42].
