Detection of tdTomato (TD) and IGF1-TD fusion protein in human neuronal progenitor cells (hNP).
(A) Expression of TD protein in transfected hNPs by their red fluorescence. (B) TD protein can also be detected by using immunohistochemistry (FITC, green). (C) Expression of IGF-TD fusion protein in hNPs by their red fluorescence. (D) Detection the IGF-1 moiety of the IGF-TD fusion protein in transfected cells by immunostaining (FITC, green). (E) Expression of IGF-1 component of the IGF-TD mRNA in hNPIGF-TD cells by qRT-PCR; IGF-1 mRNA is undetectable in untransfected hNP and hNPTD cells. (F) Western blot analysis of IGF-1 component of the IGF-TD protein in hNPIGF-TD cell lysates confirms the expected molecular weight of approximately 60 kD. IGF-1 was not detected in untransfected hNP and hNPTD cells. (G) IGF-1 levels are higher in the medium of cells transfected with the IGF-TD fusion protein (red triangles). Data are presented as mean ± SE for the day 0 wells (B; blank wells). Symbols correspond to the fitted means ± the 95% confidence limits of the measurements conducted in conditioned medium from the difference hNPs (open symbols; empty vector, yellow symbols; TD-transfected cells, green symbols; IGF-TD—transfected cells). Analysis was conducted by fitting a two way linear model for treatments and time points on the inverse of the data. Multiple comparisons were conducted on the transformed scale by the Tukey test. Asterisks indicate statistically significant differences versus all groups at the same time point. Abbreviations: hNP, neuronal progenitor cells; TD, tdTomato; IGF-TD, IGF-1-tdTomato. Scale bar in A—D: 50 μm.