The fluorescence imaging of proteins in vivo is possible using the iBox® Scientia animal imaging system from Analytik-Jena. The imaging system is designed for imaging a wide variety of different animals and multiple label studies are possible using the wide variety of different fluorescent proteins with different emission spectra.
As the proteins used alongside the system are inherently fluorescent substrates and cofactors are not required. In addition the fluorescent behavior of reporter proteins is constantly being improved upon, with a range of bright red fluorescent proteins becoming recently available. Both commercial and academic institutions can purchase the genes for these proteins.
Expressed Fluorescent Proteins
The iBox® Scientia, alongside Analytik-Jena’s f1.2 optics can be used for rapid exposures, allowing accurate images, which are unaffected by the breathing of the animal, to be captured. Using Analytik-Jena’s VisionWorks software with strongly fluorescent proteins the margins of tumors can be identified and the size of the tumor can be quantified.
Following research conducted by Zhao et al., the specimen should be injected daily with a Salmonella typhimurium mutant. S. typhimurium accumulates in tumors and reduces (or completely stops) the metastatis of PC-3 prostate tumor cells in humans. This research used a red fluorescent protein (RFP), U87, to express human glioma cells.
Figure 1. iBox Scientia small animal imaging system
Materials and Methods
The iBox Scientia in vivo imaging system from Analytik-Jena was set up as follows:
Analytik-Jena’s iBox Scientia in vivo imaging system was configured with the following features:
- Motorized sample platen
- BioLite™ - Automated excitation light source
- BioChemi 500 camera - High resolution (4.2 mpx)
- f1.2 – Large area, rapid biolens
- CFP, YFP, RFP and GFP emission and excitation filter sets
- Analytik-Jena Heater – set to 370 °C
- VisionWorks®LS – Analysis software
A simple imaging method was used. A small volume of U87 cells were introduced via subcutaneous injection into nude mice. Day 0 was designated at the point at which the tumor was observed to have grown to a size of 1 mm2. The treatment group received an injection of 5x107/100µl Salmonella typhimurium A1-R via the tail on a weekly basis.
Figure 2. RFP orthotopic tumors in nude mice imaged over a period of two weeks using Analytik-Jena iBox® Scientia small animal imaging system.
The excitation and emission filter for RFP was then used with the Analytik-Jena iBox to take white light and fluorescence images. These images were then combined via blend control (with the source and destination set at 75% and 25% respectively. The fluorescent image was colored red to indicate where RFP was present. The exposure times used for RFP and white light images were 1 second and 0.01 seconds respectively.
Comparing the treated group and untreated control after a week and two weeks showed that tumor growth had been reduced in the treated group. This suggests that weekly injections of Salmonella typhimurium can be used to reduce tumor growth.
Research into tumor imaging using fluorescent imaging such as this shows the importance of a correct process in time course research and the importance of fluorescent protein imaging when studying cancers in vivo. The iBox® Scientia combined with Analytik-Jena’s f1.2 optics can be used to provide sharp images which are unaffected by the breathing of the subject being imaged.
- Hoffman, R.M. and Yang, M. Whole-body imaging with fluorescent proteins. Nat. Protoc. 2006, 1, 1429-1438.
- Hoffman, R.M. and Yang, M. Color-coded fluorescence imaging of tumor-host interactions. Nat. Protoc. 2006, 1, 928-935.
- Zhao, M.; Geller, J.; Ma, H.; Yang, M.; Penman, S.; Hoffman, R.M. Monotherapy with a tumor-targeting mutant of Salmonella typhimurium cures orthotopic metastatic mouse models of human prostate cancer. Proc. Natl. Acad. Sci. U. S. A. 2007, 104, 10170-10174.
- Zhao, M.; Yang, M.; Ma, H.; Li, X.; Tan, X.; Li, S.; Yang, Z.; Hoffman, R.M. Targeted therapy with a Salmonella typhimurium leucine-arginine auxotroph cures orthotopic human breast tumors in nude mice. Cancer Res. 2006, 66, 7647-7652.
This information has been sourced, reviewed and adapted from materials provided by Analytik Jena US.
For more information on this source, please visit Analytik Jena US.