Figure 1: Localizing luciferase labeled cells

Luciferase: Cell tracker

Luciferase is a series of enzymes that can catalyze substrates to produce bioluminescence. Different sources of luciferase have their own characteristics, and different luciferase can catalyze substrates to emit different colors of light. Firefly luciferase became the most commonly used mammalian cell reporter among these enzymes because of its high sensitivity and wide detection linear range (up to 7 to 8 orders of magnitude).The effect is that specific cells can be tracked and detected at any time in subsequent experiments by simply inserting the reporter once.

Figure 2: The principle of luciferase and luciferin potassium salt reaction luminescence

The advantages of luciferase imaging methods

Radiation-free and virtually harmless to living organisms.

Imaging via bioluminescence rather than excitation light source.

High sensitivity: The number of cells detected can be as low as hundreds.

Good penetration, the fluorescence signal can still be detected even through 3-4cm of tissue.

High signal-to-noise ratio, strong fluorescent signal, good anti-interference.

Application scenario

Monitoring of tumor growth

Real-time observation of tumor growth into tumor in nude mice in vivo, the tumor body without separation.

Monitoring of tumor drug function

To detect the effect of drugs on tumor growth or tumor metastasis in vivo. The fluorescein substrate can be completely eliminated in 3 hours so it won't interfere with the drug.

Cell localization

The localization and distribution of foreign cells in animals were detected.

Gene expression regulation

The target gene or target gene promoter fuses with the luciferase gene to detect changes in gene expression during drug treatment or disease progression.

Stem cell research

Monitoring the transplantation, survival and proliferation of stem cells; Trace the distribution and migration of stem cells in vivo.

Experimental Results

Figure 3: In vivo imaging detection of CAR-MUC1 T/CAR-MUC1-IL22 T cells for tumor formation by subcutaneous injection of HN4 cells in mice

Figure 5: In vivo imaging of the ability of mesenchymal stem cells (MSC) to migrate to the burn site. Mesenchymal stem cells (MSC/FLuc) were injected intravenously into a mouse back burn model. Bioluminescent signals appeared at the injury site of the burn wound 4 days after injection, and then gradually decreased (red arrow indicates the burn site) [3].

FAQ

Q1: What are the advantages of bioluminescent in vivo imaging methods compared to other similar methods?

A: Compared with other types of technology, bioluminescence in vivo imaging method is more sensitive than traditional methods in the study of tumor metastasis, gene therapy, epidemiological pathogenesis, stem cell tracer, leukemia related research, etc., and can also quickly and intuitively carry out the pathogenesis and drug screening research of related diseases through a series of transgenic animal disease models.

Q2: How to label stem cells with luciferase gene?

A: By markers of sexual expression of genes to the preparation of transgenic mice, which stem cells were marked. Hematopoietic stem cells were extracted from the bone marrow of such transgenic mice and transplanted into the bone marrow of another mouse to track the proliferation and differentiation of hematopoietic stem cells in vivo and the process of migration to the whole body. Or you can label stem cells with lentiviruses.

Q3: What is the appropriate detection time after injection of fluorescein and how long does the luminescence last?

A: After intraperitoneal injection, the fluorescence signal generally reached the strongest stable period after 10-15min, began to decay after 20-30min, and eliminated fluorescein after 3h.

Q4: What is the available injection method of luciferase reagent for mouse experiments? What are the differences between different injection methods?

A: Fluorescein can be injected into mice by intraperitoneal injection or intravenously at the tail. It can spread to the whole body of mice in about 1 min. Most cases use fluorescein concentrations of 150 mg/kg. About 3 mg of fluorescein is sufficient for 20 g mice. For intraperitoneal injection, diffusion is slower, the start of light is slower, and the duration of light is longer. For the tail intravenous injection of fluorescein, diffusion is fast and the luminescence begins quickly, but the luminescence duration is short.

Product information

References

[1]. Mei Z, Zhang K, Lam AK, Huang J, Qiu F, Qiao B, Zhang Y. MUC1 as a target for CAR-T therapy in head and neck squamous cell carinoma. Cancer Med. 2020 Jan;9(2):640-652. doi: 10.1002/cam4.2733. Epub 2019 Dec 4. PMID: 31800160; PMCID: PMC6970025.

[2]. Chen G, Fan XY, Zheng XP, Jin YL, Liu Y, Liu SC. Human umbilical cord-derived mesenchymal stem cells ameliorate insulin resistance via PTEN-mediated crosstalk between the PI3K/Akt and Erk/MAPKs signaling pathways in the skeletal muscles of db/db mice. Stem Cell Res Ther. 2020 Sep 16;11(1):401. doi: 10.1186/s13287-020-01865-7. PMID: 32938466; PMCID: PMC7493876.

[3]. Oh EJ, Lee HW, Kalimuthu S, Kim TJ, Kim HM, Baek SH, Zhu L, Oh JM, Son SH, Chung HY, Ahn BC. In vivo migration of mesenchymal stem cells to burn injury sites and their therapeutic effects in a living mouse model. J Control Release. 2018 Jun 10;279:79-88. doi: 10.1016/j.jconrel.2018.04.020. Epub 2018 Apr 12. PMID: 29655989.