Colorimetric Phospholipid Determination with Erythrosin B

Andree, Hans; Soedjak, Helena S.

doi:10.1006/abio.1994.1518

Cited by 6 publications

(6 citation statements)

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“…Shift of the absorbance maxima of erythrosine dye at pH 4.25 after complexing to all the cationic liposomes in the present study is similar to that with anionic and neutral phospholipids (Andree and Soedjak, 1994). There is a measurable difference between absorbance of free dye to that of dye complexed to liposomes at 549 nm.…”

Section: Discussionsupporting

confidence: 76%

“…An alternative and relatively easy method is herein described for the quantitation of cationic liposomes. This method is an extension of previous work by Andree and Soedjak (1994) who developed it for detection of anionic and neutral phospholipids using erythrosine dye. The method is simple, requires minimal preparation, uses nonsophisticated equipment, is nonhazardous , and is time-ef cient.…”

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confidence: 93%

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Liposomes Containing Cationic Dimethyl Dioctadecyl Ammonium Bromide: Formulation, Quality Control, and Lipofection Efficiency

2002

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This article describes a novel, simple, and relatively inexpensive method to prepare cationic liposomes using an ethanol injection/pressure extrusion method. The study also demonstrated that binding erythrosine dye to cationic liposomes results in a shift of the absorption maximum of the dye from 528 nm to 549 nm at pH 4.25, allowing quantification and visualization of these vesicles. In addition, a relatively simple Ficoll-based gradient centrifugation method for separation of lipoplexes from unbound molecules is presented. Laboratory-formulated dimethyl dioctadecyl ammonium bromide (DDAB) containing liposomes were just as efficient in complexing nucleic acids as commercially available types, and binding increased as the positive to neutral lipid ratio was increased. Transfection efficiency of the DDAB-containing liposomes increased as the ratio of cationic to neutral lipid was increased from 1:1 to 4:1 with either PtdChol or DOPE as the neutral lipid. A concomitant increase in cytotoxicity of CSU-SA1 cancer cells was noted as the ratio of positive to neutral lipid of the liposomes was increased. Nevertheless, our present study showed that the 2:1 liposome is a good choice since it delivers functional plasmids at a comparable rate to commercial liposome formulations, has similar toxicities to the less harmful commercial liposomes, and is at least 1000-fold more economical to prepare inhouse, a major factor to be considered in preclinical and clinical studies with these carriers.

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Section: Discussionsupporting

confidence: 76%

mentioning

confidence: 93%

Liposomes Containing Cationic Dimethyl Dioctadecyl Ammonium Bromide: Formulation, Quality Control, and Lipofection Efficiency

2002

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“…From experience, these vesicles should also be able to deliver nucleic acid constructs into other cell types including plant cells. Shift of the absorbance maxima of erythrosine dye at pH 4.25 after complexing to all the cationic liposomes in the present study is similar to that with anionic and neutral phospholipids (Andree and Soedjak, 1994). There is a measurable difference between absorbance of free dye to that of dye complexed to liposomes at 549nm.…”

Section: Discussionsupporting

confidence: 76%

“…A relatively easy method is herein described for the quantitation of cationic liposomes. This method is an extension of previous work by Andree and Soedjak (1994) who developed it for detection of anionic and neutral phospholipids using erythrosine dye. The method is simple, requires minimal preparation, uses nonsophisticated equipment, is non-hazardous, and is timeefficient.…”

Section: Introductionmentioning

confidence: 93%

Formulation and quality control of cationic liposomes

Dass¹

2001

S. Pac. J. Nat. App. Sci.

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Cationic liposomes are traditionally used for delivering macromolecules such as nucleic acids to mammalian and plant cells. This paper describes a novel simple and relatively inexpensive method for preparation of cationic liposomes using an ethanol injection/pressure extrusion method. The study also evaluated the utility of a colorimetric method for quantification of cationic liposomes. Binding of erythrosine dye to cationic liposomes resulted in a shift of the absorption maximum of the dye from 528nm to 549nm in a buffer at pH 4.25, allowing quantification of these vesicles. Colour development was completed in 5 to 10 minutes at room temperature, with only 10% decrease in absorbance observed in the following 2 hours. Divergent values were noted in the presence of interfering agents such as detergents and salts. The erythrosine method is sensitive down to 0.20 �g/mL of cationic lipid and is linear to 3.13 �g/mL. The erythrosine dye method for quantitation of cationic liposomes is valuable for the field of liposome technology. In addition, a relatively simple method for separation of nucleic acids complexed to cationic liposomes from unbound molecules is presented. This method utilises a Ficoll-based gradient centrifugation method. Laboratory-formulated liposomes were just as efficient in binding nucleic acids as commercially available types.

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“…Eluates were dialyzed and control liposomes collected by centrifugation. Lipid concentrations were measured by the method of Andree and Soedjak (13). Other routine methods can be found in Figler et al (11).…”

Section: Methodsmentioning

confidence: 99%

A Novel Electron Paramagnetic Resonance Approach to Determine the Mechanism of Drug Transport by P-glycoprotein

Omote

Al‐Shawi

2002

Journal of Biological Chemistry

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ATP-driven pumping of a variety of drugs out of cells by the human P-glycoprotein poses a serious problem to medical therapy. High level heterologous expression of human P-glycoprotein, in the yeast Saccharomyces cerevisiae, has facilitated biophysical studies in purified proteoliposome preparations. Membrane permeability of transported drugs and consequent lack of an experimentally defined drug position have made resolution of the transport mechanism difficult by classical techniques. To overcome these obstacles we devised a novel EPR spin-labeled verapamil for use as a transport substrate. Spin-labeled verapamil was an excellent transport substrate with apparent turnover number, K m and K i values of 5.8 s ؊1 , 4 M, and 210 M, respectively, at pH 7.4 and 37°C. The apparent affinities were ϳ10-fold higher than for unlabeled verapamil. Spin-labeled verapamil stimulated ATPase activity ϳ5-fold, was relatively hydrophilic, and had a very low flip-flop rate, making it an ideal transport substrate. The K m for MgATP activation of transport was 0.8 mM. By measuring the mobility of spin-labeled verapamil during transport experiments, we were able to resolve the location of the drug in proteoliposome suspensions. Steady state gradients of spin-labeled verapamil within the range of K i /K m ratios were observed.

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Colorimetric Phospholipid Determination with Erythrosin B

Cited by 6 publications

References 0 publications

Liposomes Containing Cationic Dimethyl Dioctadecyl Ammonium Bromide: Formulation, Quality Control, and Lipofection Efficiency

Liposomes Containing Cationic Dimethyl Dioctadecyl Ammonium Bromide: Formulation, Quality Control, and Lipofection Efficiency

Formulation and quality control of cationic liposomes

A Novel Electron Paramagnetic Resonance Approach to Determine the Mechanism of Drug Transport by P-glycoprotein

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