Temperature molecular probes

It is necessary to note that fluorescence characteristics demonstrate remarkable sensitivity to variations of physicochemical parameters of the environment. Therefore, such parameters as polarity, viscosity, temperature, electric potential, local electric field, pressure, pH, etc., can be registered successfully using the modem sensitive apparatus for fluorescence detection [1, 4—12]. As a consequence, fluorescent molecules are used successfully as molecular probes to study the local characteristics of physicochemical, biochemical and biological systems. 

Dissolve 7-AAD (Molecular Probes Inc.) in buffer at a concentration of 100 pg/ml. The solution should be stored as small aliquots at 4°C or -20° C, and brought to room temperature before use. 

Figure 5. Fluorescence of collagen IV conjugated with Oregon Green 488 (Molecular Probes, Eugene, OR, USA.) spontaneously adsorbed to unmodified polyethylene foils (A) or polyethylene modified with 1014 0+ ions/cm at the energy of 30 keV (B). Collagen was diluted in phosphate-buffered saline to the concentration of 0.02 mg/ml (10 pg/cm ) and incubated with the foils for 24 h at room temperature. For auto fluorescence control.

Baker and coworkers [16] reported on a self-referencing luminescent thermometer designed around the temperature-dependent intramolecular excimer formation/dissociation of the molecular probe l,3-Ws(l-pyrenyl)pro-pane (BPP) dissolved in 1-butyl-l-methylpyrrolidinium bjs(trifluoromethyl-sulfonyl)imide, [C4Cipyr][Tf2N]. Upon an increase in temperature, and hence a decrease in the IL s bulk viscosity, the excimer-to-monomer fluorescence... 

The peaks in s j dipolar (which are usually close to the peaks in e") can be used to determine the time or point in the cure process when the mean dipolar relaxation time has attained a specific value, i = 1/co, where to = 2nf is the frequency of measurement. The dipolar mobility as measured by the mean relaxation time t can be used as a molecular probe of the buildup in Tg. The time of occurrence of a given dipolar relaxation time as measured by a peak in a particular high frequency value of e"(co) can be quantitatively related to the attainment of a specific value of the resin s glass transition temperature. 

Carboxyfluorescein-A-hydroxysuccinimide ester (Roche Diagnostics, Lewes, UK Molecular Probes, Eugene, OR) and 5(6)-carboxytramethylrhodamine-iV-hydroxysuccinimide ester (Molecular Probes, Eugene, OR) are alternative reagents. They can be conjugated to IgG in PBS at room temperature in about 1 h. 

In Sec. IV we discuss another TPM dye, malachite green (MG), which was used as a molecular probe for glass transition of alcohols and polymers [11,12], Analysis of the temperature dependence of nonradiative relaxation in MG shed light on the understanding of the mechanism of glass transition. Novel experimental observations are divided into two classes. (1) The critical temperature (Tc) predicted by the mode-coupling theory (MCT) was undoubtedly... 

Immersion of dry samples in liquids of different molecular size This method is designed to take advantage of molecular sieving. The basic data are simply in the form of a curve of the specific energy of immersion versus the molecular size of the immersion liquid. This provides immediate information on the micropore size distribution. For room-temperature experiments one can use the liquids listed in Table 8.1, which are well suited for the study of carbons. Because of the various ways of expressing the critical dimension of a molecular probe or its molecular size , one must be careful to use a consistent set of data (hence the two separate lists in Table 8.1). Again, one can process the microcalori-metric data to compare either the micropore volumes accessible to the various molecules (see Stoeckli et a ., 1996), or the micropore surface areas, as illustrated in Figure 8.5. 

Finally, dynamic structure-based pharmacophore models can be derived through a method first described by Carlson et al that uses multiple conformations of the target protein, which are obtained either by molecular dynamics simulation or by the use of multiple experimentally determined conformations. The binding sites of the respective snapshots are flooded with small molecular probes (e.g., methanol for hydrogen-bond interactions and benzene for aromatic hydrophobic interactions) and while the protein structure is held rigid the probe molecules are subjected to a low-temperature Monte Carlo minimization where they undergo multiple, simultaneous gas-phase... 

Table 3.10 Summary of the characteristics of protonic zeolites and on the diffusion of molecular probes in their cavities as measured by IR spectroscopy at room temperature. 

Kmilarly, Sung et al. recently reported the use of azo chromophoric labels as a molecular probe of physical aging in amorphous polymers . By measuring the kinetics of trans cis photoisomerization of azo duomophores covalently bonded to amorphous polyurethanes, they defined a parameter a which corresponds to the fraction of the free volume above a critical size at a given temperature and time of aging. 

The success of the method prompted the design of an Automatic Molecular Probe apparatus for the collection of retention data 82). At a preset cycle time a mixture of solute and noninteracting marker was iiqected into the carrier gas and the output of the GC detector was fed to an electronic peak detector. The temperature in the oven was programmed linearly and recorded with a thermocouple. Upon conversion into digital form, a printout of net retention time and temperature was obtained. Retention diagrams have been obtained with this apparatus for high and low density polyethylene 82, 83). 

Ricin A-chain (Inland Labs) at 3 mg/mL in PBS was reduced for 30 min with 30 mM DTT at 30°C. Excess DTT was removed by Sephadex G-25 gel filtration in 5 mM sodium acetate buffer, pH 4.7, containing 50 mM NaCl and 0.5 mM EDTA. The thiol content was assessed by Ellman s assay (Ellman, 1959). MIANS (Molecular Probes) was added at a ratio of 0.9 mole MIANS 1.0 mole thiol. The reaction mixture was incubated at ambient temperature for 15 min after the pH was raised to 7.0 using 1 M Tris.HCl buffer, pH 7.4. The MIANS was quenched by addition of 1 mole equivalent of freshly prepared cysteine.HCl/mole MIANS and incubation for an additional 15 min. Any remaining thiol groups were alkylated by adding a 5-fold molar excess of iodoaceteimide over total thiol and after an additional 30 min, the protein (MIANS-ricin A-chain) was dialyzed against 0.1 M Tris.HCl buffer, pH 8.5. 

IR spectra were recorded on a Pcrkin-Elmer FT-IR spectrometer of double window type and with resolution of 4 cm L In-situ IR cells suitable to the chamber were made of quartz. Cap2 windows. The gas pressure in the cell could be varied from 0.1 MPa to L3 mPa. Room temperature CO adsorption was used as a molecular probe. XPS XRD, and TPR measurements were implemented as described in refs. 9 and 10. 

Synchrotron-based EXAFS can be used to study most chemical elements in solid, liquid, or gas phases at concentrations as low as millimoles per cubic meter. The high intensity of synchrotron radiation allows the study of very small or dilute samples under conditions of varying temperature or pressure and in controlled environments, including the presence of liquid water. Thus, the method is noninvasive and can be used with in situ molecular probes. 

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