Hi, did anyone use the Clodrosome® standard macrophage depletion kit (containing Clodronate liposome and control liposomes), and know how to prepare it before injection? Could you kindly provide a SOP, if it is possible?
Macrophage depletion has been used extensively to study autoimmune disease and more recently in tumor models. The clodronate-containing liposomes will be recognized as foreign particles and get engulfed by macrophages upon dosing into the animal by the chosen routes. Consequently, macrophages that have engulfed liposomes will all be destroyed by the liposomal. In the protocol presented here the clodronate-containing liposomes were used to systemically deplete macrophages in mice.
Liposomes suspended in sterile phosphate buffered saline (see Recipes)
Equipment
Syringe (10 ml and 1 ml) (BD, catalog numbers: 309604 and 305111)
26 G 1/2 needles (BD, catalog number: 309659)
Procedure
Twenty, 8-week-old mice were randomly divided into two groups.
One group of mice were intravenously injected t using 1-ml syringe with 26 G 1/2 needles, twice per week with clodronate-loaded liposomes in 150 µl for the first month and 100 µl following the first month’s injection to reduce possible side effects, such as infection, live toxicity et al. The mice were treated for 3 months totally.
The second group was treated with PBS-liposomes.
At different time points (such as 2, 4, 8 weeks after treatments) of the experiment, one or two mice from each group will be euthanized to examine inflammation and other interesting analysis. Based on our experiments, the FACS analysis detected around 90% of macrophage reduction in the group that treated with the clodronate-loaded liposomes versus the control group after 2 weeks treatment, and this reduction kept to the end of the experiments.
During the whole treatment period, the treated mice need to be closely monitored for the possible signs and symptoms such as weight loss, yellow, bloody, or tar-colored stool, dark urine color, diarrhea, fatigue etc.
At the end of the experiment, (depend on the experiment), the mice were euthanized and the lung tissues were collected for various analyses.
Recipes
Notes:
Based on the instructions from company.
The clodronate-loaded liposomes and PBS-containing liposomes are ready for use, but the solutions that usually kept in 4 °C need to be warmed up to room temperature before injections.
Liposomes
Liposomes are artificial lipid spherules made of:Phosphatidyl choline
Cholesterol
Phosphate buffered saline (PBS)10 mM Na2HPO4
10 mM NaH2PO4
140 mM NaCI
References
Xiao, Z., Jiang, Q., Willette-Brown, J., Xi, S., Zhu, F., Burkett, S., Back, T., Song, N. Y., Datla, M., Sun, Z., Goldszmid, R., Lin, F., Cohoon, T., Pike, K., Wu, X., Schrump, D. S., Wong, K. K., Young, H. A., Trinchieri, G., Wiltrout, R. H. and Hu, Y. (2013). The pivotal role of IKKalpha in the development of spontaneous lung squamous cell carcinomas. Cancer Cell 23(4): 527-540.
Thanks a lot, Rafik. We bought the clodornate liposome from Clodrosome® . I have been told that it is ready-to-use product. However, I am not 100% sure about it. Just wondering, do you know anything about it?