Vesicles, pore size

Figure 1 Freeze-fracture electron micrographs of egg phosphatidylcholine large unilamellar vesicles prepared by extrusion through polycarbonate filters with pore sizes of (A) 400 nm, (B) 200 run, (Q 100 nm, (D) 50 nm, and (E) 30 nm. The bar in panel (A) represents 150nm. Source From Ref. 7.

The superficial two to three cell layers of the corneal and conjunctival epithelium are the main barrier for the permeation of topically applied compounds. In this rate-limiting cell layer, the transcellular permeation is dictated by the lipophilicity of the cell membrane whereas the paracellular permeation is limited by the paracellular pore size and density. Vesicular penetration (e.g., receptor- or endocytosis-mediated) of macromolecules across surface epithelium is possible [33], However, the proposed mechanism is energy consuming (e.g., incorporation into pinocytotic vesicles and phagosomes) and thus more feasible in cell lines with abundant intracellular energy sources like corneal endothelium and RPE [34-37]. 

Shape selective reactions are typically carried out over zeolites, molecular sieves and other porous materials. There are three major classifications of shape selectivity including (1) reactant shape selectivity where reactants of sizes less than the pore size of the support are allowed to enter the pores to react over active sites, (2) product shape selectivity where products of sizes smaller than the pore dimensions can leave the catalyst and (3) transition state shape selectivity where sizes of pores can influence the types of transition states that may form. Other materials like porphyrins, vesicles, micelles, cryptands and cage complexes have been shown to control product selectivities by shape selective processes. 

In order to prepare liposomes, the lipid preparation is dried at low temperature under an inert gas atmosphere (protect the lipid from oxidation). The lipid film is swollen with water or buffered aqueous solution and several freeze-thaw cycles are carried out to get optimal rehydration of the lipid. The rehydrated lipid preparation is filtered using membrane filters with defined pore size. After repeated filtration steps (extrusion) an unilamellar liposome preparation with a defined size distribution is obtained. Large unilamellar vesicles (LUV) are produced in this way. LUV s are about 100 nm in size the thickness of the lipid bilayer is about 4 nm. Even smaller liposomes can be derived from sonication (sonication probe or ultra-sonication bath). Separation of the prepared liposomes... 

Amatore C, Arbault S, Bonifas I, Bouret Y, Erard M, et al. 2005. Correlation between vesicle quantal size and fusion pore chromaffin cell exocytosis. Biophys J 88 4411-4420. 

After sonication the samples are filtered by syringe filters with the pore size of 0.45 pm to separate large vesicles that may blockade the extruder membrane (see Note 3). 

Zhelev, D.V. and Needham, D. (1993) Tension-stabilized pores in giant vesicles-determination of pore-size and pore line tension. Biochimica et Biophysica Acta, 1147 (1), 89-104. 

Liposomes are formed by rehydration of a lipid film at temperatures below 4°C. The substances to be entrapped are added to the aqneons solntion consequently, formation of vesicles and entrapment occur simultaneously. After this preparation period, inhibitor molecules are added to the external medium that block all enzymes that have not been entrapped, before the suspension is extruded through filters with appropriate pore sizes. One main drawback of this strategy is that it can only be applied with lipids that have their main transition temperature below 0°C. This technique has basically been applied for the synthesis of poly(Phe) in liposomes. ... 

The resulting large unilamellar vesicles (LUVs) have a mean diameter of 100-140 nm as determined by quasielastic light scattering (QELS). This diameter can be modified by using membranes of different pore size. 

Adsorption on glass fiber filters is simple, inexpensive, and popular. The large internal surface of the filters adsorbs membrane vesicles via weak electrostatic and hydrophobic interactions. Bigger vesicles (mitochondriae, synaptosomes) are also filtered mechanically (pore size of glass fiber filters 1 to 3 pm 0 mitochondriae 1 pm). Molecules in solution largely... 

Oligolamellar vesicles and LUV Homogenization of MLV with liposome extruder and polycarbonate membranes of defined pore size Heterogeneous vesicle mixture requires specialized equipment... 

The advantage of such approach is the possible selective effects deriving from the molecular weight cut-off, set by the pore size. It is then possible that small molecules may penetrate and react into the vesicles, whereas large macromolecules (such as enzymes, nucleic acids) are retained inside. 

Size Control. Once the vesicles are formed, they will keep their size and size distribution. In order to reduce their size one has to disrupt the vesicle membrane, which is done by strong shear forces, eg by ultrasonic irradiation, ball milling, and high-pressure extrusion through membranes or slits ( French press ). Extrusion through polycarbonate membranes, which are available with pore diameters between 50 and 800 nm, allows a reduction and control in vesicle size and polydisper-sity to obtain small unilamellar vesicles with size distributions within 10-20%. Fast and direct preparation of small monodisperse vesicles in the size range of 50-200 nm is possible using inkjet printers (133). By appropriate choice of sample preparation method, it is thus possible to tailor the size of the vesicle between 50 nm and 10 fim with narrow size distribution. 

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