Calorimetric investigation of the phase partitioning of the fluorescent carbocyanine probes in phosphatidylcholine bilayers

Ethier, Michael F.; Wolf, David E.; Melchior, Donald L.

doi:10.1021/bi00274a029

Cited by 25 publications

(16 citation statements)

References 20 publications

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“…different effective viscosities in both liposomes (Wu and McConnell, 1975;Shimshick and McConnell, 1973;Johnson and Garland, 1983) and phospholipid monolayers (Seul et al, 1985;McConnell et al, 1984) of pure and mixed lipid compositions. However, the extent to which diI partitions between fluidlike and solidlike phases may depend on a number of factors (Ethier et al, 1983;Packard and Wolf, 1985). The third possibility agrees with previous reports of the formation of diI aggregates in model membranes (Ethier et al, 1983;Klausner and Wolf, 1980).…”

supporting

confidence: 88%

“…However, the extent to which diI partitions between fluidlike and solidlike phases may depend on a number of factors (Ethier et al, 1983;Packard and Wolf, 1985). The third possibility agrees with previous reports of the formation of diI aggregates in model membranes (Ethier et al, 1983;Klausner and Wolf, 1980). None of the measured parameters for the rotational motion of diI in DSPC monolayers and DSPC/DNP-DOPE monolayers untreated and treated with unlabeled ANO2 antibodies were statistically different according to two-tailed t-tests with a 95% confidence limit (Table 4).…”

supporting

confidence: 88%

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Slow rotational mobilities of antibodies and lipids associated with substrate-supported phospholipid monolayers as measured by polarized fluorescence photobleaching recovery

Timbs¹,

Thompson²

1990

Biophysical Journal

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Polarized fluorescence photobleaching recovery has been used to monitor slow rotational motions of a fluorescently-labeled anti-dinitrophenyl mouse IgGl monoclonal antibody (ANO2) specifically bound to substrate-supported monolayers composed of a mixture of distearoylphosphatidylcholine (DSPC) and dinitrophenyldioleoylphosphatidylethanolamine (DNP-DOPE). ANO2 antibodies were labeled with a new bifunctional carbocyanine fluorophore that has two amino-reactive groups; steady-state fluorescence anisotropy data confirmed the expected result that the ANO2-conjugated bifunctional probe had less independent flexibility than ANO2-conjugated unifunctional fluorescence labels. Rotational mobilities were also measured for the fluorescent lipid 1,1'-dioctadecyl 3,3,3',3'-tetramethylindocarbocyanine (dil) in DSPC and in mixed DSPC/DNP-DOPE monolayers in the presence and absence of unlabeled ANO2 antibodies. The apparent rotational correlation time and fractional mobility of ANO2 on supported monolayers were approximately 70 and approximately 0.3 s, respectively. These measured parameters of rotational mobility did not depend on the ANO2 surface density or on kinetic factors, but addition of unlabeled polyclonal anti-(mouse IgG) antibodies significantly decreased the apparent mobile fraction. The measured fluorescence recovery curves for dil were consistent with two fluorophore populations with rotational correlation times of approximately 4 and approximately 100 s and a population of immobile fluorescent lipid. No difference in fluorescence recovery and decay curves was measured for dil in DSPC monolayers, DSPC/DNP-DOPE monolayers, and DSPC/DNP-DOPE monolayers treated with unlabeled ANO2 antibodies.

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supporting

confidence: 88%

supporting

confidence: 88%

Slow rotational mobilities of antibodies and lipids associated with substrate-supported phospholipid monolayers as measured by polarized fluorescence photobleaching recovery

Timbs¹,

Thompson²

1990

Biophysical Journal

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“…If the second component did indeed represent membrane flow, the C12NFlu probe should participate in this exchange process and one should see two recoveries with this probe. On the other hand, in the presence of immiscible gel and fluid lipid domains, previous reports have indicated that such a small probe should partition primarily into the fluid phase (4,8,27 Moreover, the percent recovery for C12NFlu is equivalent to the total mobile fraction of the other lipid probes (see Table 1), suggesting that it samples the same compartments. Jacobson et al (24)-have detected two diffusing species in plasma membrane lipid multibilayers below 10°C and have pointed out that multiple phase equilibrium is strongly suggested by the detection of two diffusing components.…”

Section: Discussionmentioning

confidence: 90%

Lateral diffusion of phospholipids in the plasma membrane of soybean protoplasts: Evidence for membrane lipid domains

Metcalf

Wang²,

Schindler³

1986

Proc. Natl. Acad. Sci. U.S.A.

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Fluorescent lipid and phospholipid probes were incorporated at 40C into soybean protoplasts prepared from cultured soybean (SB-1) cells. Fluorescence microscopy showed that the plasma membrane as well as the nucleus were labeled. Fluorescence redistribution after photobleaching (FRAP) analysis was performed on these cells at 18°C to monitor the lateral mobility of the incorporated probes. After labeling at low concentrations (40 pg/ml) of phosphatidyl-N-(4-nitrobenzo-2-oxa-1,3-diazolyl)ethanolamine (NBD-PtdEtn), a single mobile component was observed with a diffusion coefficient (D) of w3 X 10-9 cm2/sec. After labeling at higher probe concentrations ('100 jig/ml), two diffusing species were observed, with diffusion coefficients of "'3 x 10-cm2/sec ("fast") and m5 x 10-10 cm2/sec ("slow"). Similar results were observed with fluorescent derivatives of phosphatidylcholine and fatty acids. In contrast to these results, parallel analysis of 3T3 fibroblasts, using the same probes and conditions, yielded only a single diffusion component. These results suggest that the soybean plasma membrane may contain two distinct lipid domains in terms oflipid mobility. Consistent with this idea, experiments with soybean protoplasts yielded a single diffusion component under the following conditions: (i) labeling with NBD-PtdEtn (100 ,ug/ml), FRAP analysis at 37C (D = 1.1 x 10-8 cm2/sec); (ii) labeling with NBD-PtdEtn (100 pg/ml), FRAP analysis at 180C in the presence of 2 mM EGTA (D = 4.2 x 10' cm2/sec); (iii) labeling with 5-(Ndodecanoyl)aminofluorescein (a short-chain lipid probe), FRAP analysis at 18'C or 3rC (D = 2.5 x 10-8 cm2/sec). (8,9). These measurements, however, failed to demonstrate any major differences in the diffusion coefficient (D) of a lipid probe in mammalian plasma membranes (10).We have reported diffusion coefficients for lectin-receptor complexes on the plasma membrane of soybean protoplasts derived from the SB-1 cell line (11). Here we extend the analysis of the lateral mobility of the components of the soybean cell plasma membrane by studying the diffusion of fluorescent lipid analogs. The data suggest that the soybean membrane may be a composite of large-scale, immiscible gel and fluid lipid domains. This suggestion is particularly intriguing in light of the higher content of negatively charged phospholipid species, particularly phosphatidylglycerol, in plant plasma membranes (12), as compared to mammalian plasma membranes (13). In this context, Verkleij et al. (14) and Ohnishi and Ito (15) have reported that phase separation and the formation of negatively charged phospholipid domains occurs after addition of Ca2+ to mixed phospholipid model systems. MATERIALS AND METHODSMaterials. The fluorescent probes used in this study were 1-acyl-2-[N-(4-nitrobenzo-2-oxa-1,3-diazoyl)amino]caproylsn-glycero-3-phosphocholine (NBD-PtdCho), phosphatidyl-N-(4-nitrobenzo-2-oxa-1,3-diazolyl)ethanolamine (NBDPtdEtn), 1,1'-dioctadecyl-3,3,3' ,3'-tetramethylindocarbocyanine iodide (diC18Icc), 1,1'-ditetradecyl-3,3-3',...

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“…A larger concentration of diD is necessary due to reduced camera sensitivity in the emission range of the probe. The fluorescent probes diI and diD have been shown to associate preferentially with DLPC and DPPC, respectively [41,43].…”

Section: Methodsmentioning

confidence: 99%

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D'Onofrio

Binns

Muth

et al. 2002

Journal of Biological Physics

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Local composition, structure, morphology, and phase are interrelated in lipid bilayer membranes. This gives us the opportunity to control one or more of these properties by manipulating others. We investigate theserelationships with combinations of simultaneous two-color widefield fluorescence imaging, three-dimensional rendering of vesicle domains, andmanipulation of the vesicle morphology via optical trapping and micropipetteaspiration. We describe methods to modulate, to measure, and to probe thelocal structure of model membranes through control of membrane curvature inliposomes.

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Calorimetric investigation of the phase partitioning of the fluorescent carbocyanine probes in phosphatidylcholine bilayers

Cited by 25 publications

References 20 publications

Slow rotational mobilities of antibodies and lipids associated with substrate-supported phospholipid monolayers as measured by polarized fluorescence photobleaching recovery

Slow rotational mobilities of antibodies and lipids associated with substrate-supported phospholipid monolayers as measured by polarized fluorescence photobleaching recovery

Lateral diffusion of phospholipids in the plasma membrane of soybean protoplasts: Evidence for membrane lipid domains

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