Bioluminescent Imaging of Breast Tumor Progression in a Female Mouse Model

Begin with a female mouse injected with luciferase-expressing breast tumor cells.

The tumor cells proliferate at the injection site and develop into a primary tumor.

Place the mouse in an imaging chamber with an anesthetic flow to anesthetize the mouse.

Then, inject luciferin, a substrate for luciferase, into the anesthetized mouse.

Luciferin circulates and enters the tumor cells, reacting with luciferase to emit bioluminescence.

Using the imaging system, capture the bioluminescence signal from the mouse.

Over time, the tumor progresses at the primary site and metastasizes to the lungs.

Acquire the mouse image at regular intervals and define regions of interest.

Based on the bioluminescence intensities, construct tumor growth kinetics for both sites.

Over time, a steady increase in signal at the primary tumor site indicates continuous tumor proliferation.

However, a sharp rise in lung bioluminescence at later time points suggests tumor cell metastasis.

To begin, maintain the 4T1 cell line in the supplemented Dulbecco's Modified Eagle Medium or DMEM. Incubate the cells at 37 degrees Celsius in a humidified atmosphere containing 5 percent carbon dioxide.

Transfect the 4T1 cell line with the Luc2 gene to express luciferase, a bioluminescent enzyme that is constitutively produced. Incubate the cells to keep them ready for mouse injection.

Five minutes after anesthetizing and prepping a Balb-C female mouse, inject 2 times 10 to the power five luciferase-expressing 4T1 cells subcutaneously into the left dorsum using a 1-milliliter insulin syringe. House the animal for 4 weeks after the injection.

For imaging, place the anesthetized mouse in a dorsal recumbent position within the designated area of the bioluminescent imaging equipment. Inject the mouse intraperitoneally with 100 microliters of D-luciferin solution at a concentration of 15 milligrams per milliliter.

To adjust the imaging parameters, set the emission filter open without excitation, binning to 8, F-Stop to 2, and field of view type to D. Capture 20 bioluminescent images over 20 minutes for each animal at each imaging endpoint, ensuring the images represent a 60-second signal integration.

Using the equipment's software, define specific regions of interest to distinguish between the primary tumor and lung metastasis signals. Identify bioluminescent signals at the injection site and in the thoracic region where metastasis occurs. Calculate tumor progression based on bioluminescent signal intensities.

Using images collected weekly, construct a growth kinetics profile to represent signal intensity changes over time at both primary tumor and metastatic sites. Compare signal intensity trends to analyze tumor progression and metastasis.