Hi!
I am trying to measure fluorescence intensity (calcein self-quenching) as a way to measure changes in liposome volume as a response to hyperosmotic shock.
My liposomes are made from PG and cholesterol (5:1 ratio) and encapsulate calcein at a concentration of 50mM. The liposomes are made by the thin hydration method, the free dye is separated from the vesicles using centrifugation size exclusion columns, and the suspension is extruded through a 100nm polycarbonate membrane (mean diameter of 157 nm).
I am using a black 96 well plate, and mix 100ul of the liposomes with 100 ul of the hyperosmotic solution (150 mOsm gradient). I did see some reaction, but I am not sure its self-quenching. Attached is one of the curves I obtained.
I am thinking of increasing the concentration of calcein to 70 mM, but other than that, any advice on changes I should make to my protocol?
Thanks!
Hi,
A basic explanation with respect of your query can be found in the following text and attached Figure (from a 2010 publication cited below):
... Any of the purification technique serves to separate nanoliposome encapsulated materials from those that remain in the suspension medium. Therefore, they can also be used to monitor the storage stability of nanoliposomes in terms of leakage or the effect of various disruptive conditions on the retention of encapsulants. In the latter case, total lysis can be induced by the addition of a surfactant such as Triton X100. Retention and leakage of the encapsulated material depend on the type of the vesicles, their lipid composition and Tc, among other parameters. It has been reported that SUV and MLV type liposomes are less sensitive than LUV liposomes to temperature- induced leakage (Fig. 6). This property of liposomes and nanoliposomes can be used in the formulation of temperature-sensitive vesicles (55). ...
From:
Nanoliposomes: preparation and analysis.
M.R. Mozafari (2010);
Liposomes, 29-50.
- Badges
- Science topic
- Similar topics
- Engineering
- Chemical Engineering
- Membranes