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. Author manuscript; available in PMC: 2011 Aug 1.
Published in final edited form as: Mol Cancer Res. 2010 Jul 27;8(8):1105–1115. doi: 10.1158/1541-7786.MCR-09-0383

Figure 6. Relative p-Glycoprotein mRNA prior and after CPT treatment, and activity measurements of p-Glycoprotein in C6 cells permanently transfected with GFP or GFP-CD133.

Figure 6

(A) Quantitative real-time PCR curves of C6-GFP(left) and C6-GFPCD133 (right) treated with DMSO vehicle or 10 μM CPT. Treatment was achieved in 3 trials and 3 PCR reactions were accomplished for each trial. Corresponding β-actin PCR curves were used to normalize input cDNAs by subtracting actin Ct values from p-Glycoprotein Ct values. (B) Relative p-Glycoprotein mRNA abundance based on DMSO treated C6-GFP with the lowest level, which is compared to the abundance of mRNA with CPT-treated C6-GFP and C6-GFPCD133 cells as well as DMSO treated C6-GFPCD133 cells (Student's t-test; *CPT-treated GFP versus DMSO-treated GFP cells, p= 7.33 × 10−5; **CPT-treated GFPCD133 cells versus DMSO-treated GFPCD133, p= 0.000237; ***DMSO-treated GFP cells versus DMSO-treated GFPCD133 cells, p= 0.0469; ****CPT-treated GFP cells versus CPT-treated GFPCD133 cells, p= 0.552) (C) Hydrolysis of calcein AM from 0 to 24 min. (Values represent the mean ± SD for twelve wells. SD shown as calculated average percentage of SD/mean for each line. *Student's t-test revealed statistical significance starting at 4 min with a p= 0.0046. Duplicate wells of C6-GFP and C6-GFPCD133 without calcein AM were measured to determine background emission from green fluorescent protein. Background fluorescence from C6 cells lines did not change during the 24 min assay. Therefore GFP did not contribute to rising calcein signal detection.