Abstract:Stretched polyethylene (PE) films have been used to orient small molecules for decades by depositing solutions on their surface and allowing the solvent to evaporate leaving the analyte absorbed on the polymer film. However, the non-polar hydrophobic nature of PE is an obstacle to aligning polar molecules and biological samples. In this work PE film was treated with oxygen plasma in order to increase surface hydrophilicity. Different treatment conditions were evaluated using contact angle measurement and X-ray… Show more
“…The film was treated using an Emitech (France) K1050X Plasma Asher with parameters: O 2 at 10 mmHg, 50 W RF power level, 1 min ashing time to increase the hydrophilicity of the surface. 2 …”
Section: Methodsmentioning
confidence: 99%
“…It has been shown in general that it does not matter whether the sample is added to PE films before or after stretching, 1 though different stretching amounts do affect the PE crystallite alignment and 20 population which does affect the spectrum observed. 2 However, when a dye is prone to assembling into higher order structures, different films prepared in nominally the same way can give rise to different spectra, so repeat experiments are required.…”
Section: Methodsmentioning
confidence: 99%
“…In the case of anthracene, stretching the film on which anthracene had been 70 deposited also led into a more monomeric LD spectrum. 2 In this paper we report film LD measurements on PE OX to determine the transition polarizations of some dyes that can be used as spectroscopic probes on biomolecules.…”
mentioning
confidence: 99%
“…7 Following work by Tissington, et al, 8 we found that oxygen plasma treatment formed oxygen containing groups on the surface of the polyethylene film thus introducing polar surface 60 groups. 2 We adopted less aggressive conditions than Tissington et al which did not obviously affect the surface morphology. This creates a film, which we denote PE OX , that can be used even for extremely hydrophilic molecules in the same manner as PE films can be used for hydrophobic molecules.…”
5Xanthene dyes are commonly used to label proteins in order to probe their location and activity using fluorescence spectroscopy and microscopy. However, fundamental properties such as the polarizations of transitions for many of the dyes have not been available. In this paper we report the use of recently developed oxidised polyethylene (PE OX ) stretched film linear dichroism to determine the transition polarizations of xanthene, 9-methyl-2,3,7-trihydroxy-6-fluorone, pyronin Y, pyronin B, fluorescein, and 10 rhodamine 6G. The effect of the formation of higher order structures is also discussed when they occur. The dyes (except xanthene) all have an intense long-axis polarized transition in the region of 500 nm. They also have long-axis (//) polarized transitions from about 280 nm downwards in wavelength. There are suggestions of a weak short axis (⊥) polarized transition in the region of 320-350 nm in each case.
“…The film was treated using an Emitech (France) K1050X Plasma Asher with parameters: O 2 at 10 mmHg, 50 W RF power level, 1 min ashing time to increase the hydrophilicity of the surface. 2 …”
Section: Methodsmentioning
confidence: 99%
“…It has been shown in general that it does not matter whether the sample is added to PE films before or after stretching, 1 though different stretching amounts do affect the PE crystallite alignment and 20 population which does affect the spectrum observed. 2 However, when a dye is prone to assembling into higher order structures, different films prepared in nominally the same way can give rise to different spectra, so repeat experiments are required.…”
Section: Methodsmentioning
confidence: 99%
“…In the case of anthracene, stretching the film on which anthracene had been 70 deposited also led into a more monomeric LD spectrum. 2 In this paper we report film LD measurements on PE OX to determine the transition polarizations of some dyes that can be used as spectroscopic probes on biomolecules.…”
mentioning
confidence: 99%
“…7 Following work by Tissington, et al, 8 we found that oxygen plasma treatment formed oxygen containing groups on the surface of the polyethylene film thus introducing polar surface 60 groups. 2 We adopted less aggressive conditions than Tissington et al which did not obviously affect the surface morphology. This creates a film, which we denote PE OX , that can be used even for extremely hydrophilic molecules in the same manner as PE films can be used for hydrophobic molecules.…”
5Xanthene dyes are commonly used to label proteins in order to probe their location and activity using fluorescence spectroscopy and microscopy. However, fundamental properties such as the polarizations of transitions for many of the dyes have not been available. In this paper we report the use of recently developed oxidised polyethylene (PE OX ) stretched film linear dichroism to determine the transition polarizations of xanthene, 9-methyl-2,3,7-trihydroxy-6-fluorone, pyronin Y, pyronin B, fluorescein, and 10 rhodamine 6G. The effect of the formation of higher order structures is also discussed when they occur. The dyes (except xanthene) all have an intense long-axis polarized transition in the region of 500 nm. They also have long-axis (//) polarized transitions from about 280 nm downwards in wavelength. There are suggestions of a weak short axis (⊥) polarized transition in the region of 320-350 nm in each case.
“… Fig. 9 Molecular structure of the fluorescent chromophore DPH and its stretched film LD spectrum (DPH was dropped from a concentrated solution in CHCl 3 onto prestretched polyethylene film) (Razmkhah et al 2014 ) …”
Section: Application Of Eq (
7
) To Remove Scattementioning
The interpretation of data from absorbance spectroscopy experiments of liposomes in flow systems is often complicated by the fact that there is currently no easy way to account for scattering artefacts. This has proved particularly problematic for linear dichroism (LD) spectroscopy, which may be used to determine binding modes of small molecules, peptides and proteins to liposomes if we can extract the absorbance signal from the combined absorbance/scattering experiment. Equations for a modified Rayleigh-Gans-Debye (RGD) approximation to the turbidity (scattering) LD spectrum are available in the literature though have not been implemented. This review summarises the literature and shows how it can be implemented. The implementation proceeds by first determining volume loss that occurs when a spherical liposome is subjected to flow. Calcein fluorescence can be used for this purpose since at high concentrations (> 60 mM) it has low intensity fluorescence with maxima at 525 and 563 nm whereas at low concentrations (<1 mM) the fluorescence intensity is enhanced and the band shifts to 536 nm. The scattering calculation process yields the average axis ratios of the distorted liposome ellipsoids and extent of orientation of the liposomes in flow. The scattering calculations require methods to estimate liposome integrity, volume loss, and orientation when subjected to shear stresses under flow.
Circular dichroism (CD) is the difference in absorption, A, of left and right circularly polarized light, Equation (1):
For randomly oriented systems such as solutions of molecules, only chiral molecules will show any CD intensity corresponding to their absorption bands. Chiral molecules are those molecules that cannot be superposed on their mirror images. Chiral is derived from the Greek word χϵιρ meaning hand, hence the alternate term for chirality, ‘handedness’. Two molecules that are mirror images of each other are often referred to as
enantiomers
, and equimolar mixtures of two enatiomers form a racemic mixture, which has no net CD intensity in solution. CD can be used to analyze chiral structures such as protein secondary structure in molecules and to probe interactions between chiral molecules and other molecules.
Linear dichroism (LD) is the difference in absorption of light linearly polarized parallel and perpendicular to an orientation axis, Equation (2):
LD can be used to provide orientation information about subunits of a molecular system such as small molecules absorbed onto stretched films, flow‐oriented DNAs and fibrous proteins, and lipid bilayer systems.
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