I'm confused and therefore write here with the hope that someone might provide useful input. Maybe it's too obvious and I just happen to miss the forest for the trees.
I've stably transfected HEK293 cells using HD Fugene. Once I transfected them with a EGFP-only-vector, once I used a vector with my gene of interest before EGFP. Afterwards I selected positive cell clones using G418/Geneticin (mock- and untransfected HEK293 cells died while doing so).
From flow cytometry I can clearly see that all of the colonies picked (doesn't matter which construct) are EGFP-Positive, some more, some less. Yet when using fluoresence microscopy only the colonies transfected with the GOI-EGFP-vector show GFP-fluorescence, while none of the EGFP-transfected HEK293 cells seem to exhibit GFP-fluorescence. If that matters: Cells are put on slides using cytospin centrifugation for fluorescence microscopy.
Does that sound familiar to anyone or has anyone else encountered this problem before?
Great thanks in advance!
Edited by DoraTheExplorer, 14 June 2015 - 11:59 AM.