**5.5 Application in analytical fields**

*Liposomes - Advances and Perspectives*

**5.3 Application in vaccine delivery**

**5.4 Application in diagnosis**

cost effectiveness, possibility of monitoring toxicity, biodegradability, biocompatibility and lack of dangerous immunogenicity. However, they are suffering from poor transfection efficiency due to their limited endosomal escaping ability, which exposes the genetic materials to enzymatic and acid degradation in lysosomes. This has been improved by incorporating fusogenic lipids, such as DOPE, in the liposomal formulation. These helper lipids facilitate endosomal escaping by membrane fusion with endosomes, leading to early cytoplasmic release of the gene. Though cationic liposomes have shown some dose-dependent toxicity, successful results have been obtained in cancer therapy when delivering genes encoding for tumor suppression proteins [66, 67]. He et al. have recently developed folate receptor alpha-targeted lipoplexes with therapeutic gene expression regulated by an hTERT promoter. These liposomes have shown some promise for the treatment of ovarian cancer [68].

The use of specific lipids or molecules such as phosphatidylserine, DOTAP, fatty acids and monophosphoryl lipids can produce liposomes with attractive immune-stimulating activities. Liposomes carrying antigenic materials, either encapsulated in the aqueous core, grafted or coated on the surface; can stimulate immune responses on macrophage uptake. Following endolysosomal degradation, macrophages present the antigen to T-lymphocytes that initiate the production of cytokines and specific antibodies via activation of B-lymphocytes [4]. The immune response produced by liposomes can be influenced by their composition, lamellarity, size and surface charge [69]. Liposomes containing a glycolipid, trehalose 6,6'-dibehenate, and a cationic lipid, dimethyldioctadecylammonium, in a 1:5 mass ratio have demonstrated efficient delivery of the TB vaccine Ag85B-ESAT-6. This vaccine has shown prolonged immune response without any toxic effects [70]. Although a liposomal vaccine (namely Stimuvax®) targeting the major histocompatibility class I complex for lung carcinoma was not successfully implemented, some other vaccines such as Epaxal®, Inflexal® V and Mosquirix® have been

clinically established for the treatment of hepatitis A, influenza virus infections and malaria, respectively. These vaccines, classified as virosomes, are liposomes generally made of reconstituted viral envelop supplemented with phosphatidylcholine. They offer the advantage of undergoing membrane fusion either with the cells or

The use of liposomes for diagnostic purposes is one of current topics of great interest in biomedical applications. Liposomes with magnetic properties, also called magnetoliposomes, are made by entrapping superparamagnetic iron-based nanoparticles or iron oxides or gadolinium (III) chelates for magnetic resonance imaging. Functionalized liposomes labeled with radioisotopes can also be used as molecular probes in nuclear imaging. For instance, liposomes labeled with 64Cu, 18F, 89Zr or 52Mn have been reported in positron emission tomography while 99mTC, 111In or 67Ga labeled liposomes were applied in single photon emission computed tomography. Acoustic liposomes which are liposomes made of perfluoropropane gas can be used as contrast agents in ultrasound imaging technique. The encapsulation of quantum dots and fluorescent dyes into liposomes has also led to the development of attractive liposomal platforms for diagnosis. Additionally, liposomes have shown great potential for simultaneous accommodation of drugs and diagnostic agents such as radionuclides, magnetic or contrast substances. This can be achieved by encapsulation in the inner core, embedding in the lipid bilayer, chemically grafting

the endosomes, and thus leading to efficient cytosolic delivery [2, 9].

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In analytical domains, liposome-based formulations can be usefully involved in immunoassays, biosensors analysis and liquid chromatography. Liposomes can be used to encapsulate, embed or conjugate the analytical entities with high loading capacity and huge surface area that can enhance the intensity of analytical signal. In a direct enzyme-linked immunosorbent assay (ELISA) like method, fluorophoreconjugated liposomes carrying a substantial amount of secondary antibody quickly bind to the antigen that has been fixed on the primary antibody. The addition of a colorimetric substrate leads to color development and allows for analytical estimation of the antigenic analyte. The use of liposomes has been deemed to lower the limits of detection of analytes and increase the sensitivity of immunological analysis technique [73]. In chromatography, the conjugation of liposomes to the stationary phase for gel permeation chromatography is useful for separation of drugs and proteins and for exploring possible molecular interactions on phospholipid membranes. Additionally, the cell-like appearance of liposomes makes them appropriate simulated cell models for studying and predicting the interactions between biologically active compounds and cell membranes [4].
