pn-Fect™

Designed for use with Primary Neurons including our:

• E18 and E20 Rat Primary Neuronal Tissue -NEURON CULTURES
• E18 Rat Primary Neuronal Tissue - ASTROCYTE CULTURES
• E18 Mouse Neuronal Tissue -NEURON CULTURES
• E18 Mouse Neuronal Tissue -ASTROCYTE CULTURES
• hN2 Human Neurons Discovery Kit-New
• Frozen Primary Rat Cortical Neurons

pn-Fect Customer Publications

Images: IKAP regulates neuronal differentiation in the DRG. Reduction in IKAP leads to increased numbers of neurons in the immature DRG (A–C). Embryos at St. 12 were transfected with either control shRNAs or IKBKAP shRNAs and analyzed at St 24/25. Embryos were sectioned, and immunolabled with the neuronal markers Tuj-1 or Ben and the percentage of GFP+ neurons determined. Significantly more IKBKAP shRNA transfected DRG precursor cells differentiated into neurons (arrows in B; IKBKAP shRNA 7.4; n = 3 embryos; p = 0.002; IKBKAP shRNA 1.6 & 4.5, n = 3 embryos; p = 0.001) than Control shRNA transfected DRG precursor cells (n = 5 embryos). (D–H) IKBKAP shRNA-transfected DRG precursor cells (n = 3 embryos; 252 cleaved-Caspase 3+ cells counted) were also more likely to die by apoptosis (compare D & E to F & G) than control shRNA-electroporated cells (n = 3 embryos; 117 cleaved caspase-3+ cells counted). (H) The number of cleaved Caspase 3+ cells was quantified in DRG on both the transfected side of the embryo and the non-transfected side of the embryo and a ratio determined. Significantly more cleaved-Caspase 3+ cells were present in the transfected DRG of IKBKAP shRNA transfected embryos than in the transfected DRG in embryos transfected with control shRNAs; p = 0.006. (I–N). Over-expression of the c-terminus of IKAP prevents neural crest cells from coalescing with the DRG (I, K, L; p = 0.004) but the few that do join, tend not to differentiate into neurons (p = 0.007; J, M, N). Embryos were transfected with either a construct driving expression of the c-terminus of chicken IKAP with a His tag (CT-IKAP-His; L,N) or a control, His-tagged construct (K, M) and analyzed at St. 21. The location and neuronal identity of transfected cells was determined in 3 embryos for each treatment; Control His-plasmid: n = 343 transfected cells counted; CT-IKAP-His: n = 278 transfected cells counted. Statistical analysis by Student t-test. doi:10.1371/journal.pone.0032050.g006.