Buffer HEPES

FIGURE 4.35 Elution profiles of adenovirus and vesicular scomacicus virus on TSK-GEL G6000PW and G50000PW. Column TSK-GEL G6000PW + G5000PW in series. Sample (A) adenovirus and (B) vesicular stomatitus virus. Elution 145 mA1 NaCI in 10 mM Na-HEPES buffer, pH 7.4. Flow rate ... 

Fig. 3.1.5 Effects of salt concentration on the activity of Cypridina luciferase (solid lines) and quantum yield (dotted lines). In the activity measurement, Cypridina luciferin (1 pg/ml) was luminesced with a trace amount of luciferase in 2.5 mM HEPES buffer, pH 7.5, containing a salt to be tested, at 20°C. In the measurement of quantum yield, luciferin (1 pg/ml) was luminesced with luciferase (20 pg/ml) in 20 mM sodium phosphate buffer (for the NaCl data) or MES buffer (for the CaCl2 data), pH 6.7.

Figure 4. Effect of hyperosmolality on lysosomal enzyme release from rabbit neutrophils. Cells were preincubated 10 min at 37 C in either regular HEPES buffer at 320 mosmol/kg ( ) or in HEPES buffer with 0.3-M sucrose at 680 mosmol/kg ( ), 5 Mg/mL cyto-chalasin B was added, cells were stimulated with FMLP, and p-glucuronidase was released into the medium during a 6-min period measured.

Fig. 1. EPR spectrum of the dithionite-reduced Fepr protein fromD. vulgaris [from (7)]. The protein was 272 ftmol dm" in 25 mmol dm Hepes buffer, pH 7.5, and was reduced under argon with 10 mmol dm sodium dithionite for 3 min at ambient temperature. EPR conditions microwave frequency, 9331 3 MHz modulation frequency, 100 kHz modulation amplitude, 0.63 mT microwave power, 200 mW temperature (relative gain) 16 K (6.3X).

The stoichiometry of the recharged DNA/PLL/SPLL particles was studied using sucrose-gradient ultracentrifugation of fluorescently labeled polyion complexes in 25 mM HEPES buffer. Rhodamine-labeled DNA (Rh-DNA) and either fluorescein-labeled PLL (Fl-PLL) or SPLL (Fl-SPLL) were used to determine their relative amounts within DNA... 

Hepes, buffer for ion-exchange chromatography, 3 830t Heptacesium dioxide, 5 700 Heptadecanoic acid, physical properties, 5 29t... 

The reconstituted system consisted of Cytochrome P-488 (0.2 nmol), NADPH-Cytochrome c reductase (1500 units) and sodium cholate (1.25 mg). It was preincubated for 30 min at 31°. The final reaction mixture (which was incubated at 31° for 20 min) com tained the preincubated system described above, excess NADPH and t4C-BP (100 nmol 4.1 mCi/mmol) in a final volume of 1 mL 0.5M HEPES buffer, pH 7.6. Rate of BP metabolism was 665 pmol/min/nmol Cytochrome P-488. Abbreviations used for metabolites are described in legend to Figure 2. 

In brief, the rats are anesthetized, followed by an injection of 0.2 mL of the test solution into the common carotid artery. The injection solution consists of a HEPES buffered Ringer s solution (containing 141 mM NaCl, 4 mM KC1, 2.8 mM CaCl2, and 10 mM HEPES, pH 7.4) which contains both the test substrate (e.g., a [3H]-labeled compound, about 10 /xCi) and a reference compound, which is highly extracted by the tissue (e.g., 0.1 /xCi [14C]n-butanol) in the presence or absence of transport inhibitors. If a [14C]-labeled compound is used as a test substrate, [3H]H20 can be selected as a reference compound. Rats are decapitated at 15 s after injection and the retina is removed. The retina is dissolved in 2 N NaOH and subsequently neutralized with 2 N HC1. The radioactivity is measured by liquid scintillation spectrometry. The RUI value, an index of the retinal distribution characteristics of the [3H] test substrate, is estimated using the following relationship ... 

Aspirate supernatants, taking care not to disturb the cell pellets, and leaving about 30-50 fl in each tube. Add about 1.3 ml of a suitable buffer, such as phosphate-buffered saline or HEPES-buffered saline. 

Using pyranine (8-hydroxy-1,3,6-pyrene trisulfonate) as intraliposome pH indicator, the liposomes were prepared as above (as in section Preparation of 100 nm SSL Loaded with DOX via Transmembrane AS Gradient ) with the exception that pyranine (0.5 mM) was included in the hydration solution. Removal of untrapped pyranine was achieved by gel filtration on a Sephadex G-50 column, preequilibrated with either NaCl, KCl, sucrose or AS solution (according to need). All these solutions also contained lOmM Hepes buffer at the desired pH (usually pH 7.5). 

DSPC/Chol (55 45) LUVs (diameter = 100 nm) are prepared as described in section Preparation of Sphingomyelin/Cholesterol (55 45) Large Unilamellar Vesicle by Extrusion [(Lipid) = 20 mM, volume = 5mL], using 350 mM citrate pH 4.0 as the hydration buffer, and 20 mM HEPES 1.50 mM NaCl pH 7.5 (HEPES-buffered saline) as the external buffer. In this case, the pH gradient is formed during the final dialysis step. It would also be possible to omit the final dialysis step and form the pH gradient by one of two common column methods. This could be desirable if the LUV... 

G-50 column and eluted in 2-(N-Morpholino) ethansulfonic acid hydrate (MES)/N-(2-Hydroxyethyl)piperazine-N-(2-ethane-sulfonic acid) (HEPES) buffer pH 7.2 (50 mM MES, 50 mM HEPES, 75mM NaCl) to remove unencapsulated BPs. Several formulations with different sizes were obtained (0.6, 0.4, 0.2, and 0.1 pm). Liposome size and morphology was determined by dynamic light scattering and cryo-TEM microscopy (Fig. 1). 

Figure 4 Zeta potential of liposomes with varying amounts of incorporated STPP. The zeta potential was determined at 2.5 V, 657 nm, 2.00 Hz and 25°C using the Zeta Potential Analyzer Version 3.26 from Brookhaven Instruments Corporation. For each measurement, 10 pL liposome solution (total lipid 25mg/mL STPP content varying between 0 and 25 mol%) were added into 2mL HBS, pH 7.4 and incubated until temperature equilibration was attained. Abbreviations STPP, stearyl triphenyl-phosphonium. HBS, HEPES-buffered saline. Source From Ref. 30.

DQAsome/DNA complexes ( DQAplexes ) can be prepared by simply mixing DNA with the appropriate amount of preformed DQAsomes in salt-free 5mM HEPES buffer at pH 7.4. To choose the correct ratio between DNA and DQAsomes, the DNA-binding capacity of each new batch of DQAsomes should be determined. The quantitative DQAsome-DNA-binding assay, which has been routinely used in our laboratory, employs SYBR Green I. The fiuorescence signal of this dye is greatly enhanced when bound to DNA. Displacement of the dye from DNA results in loss of fluorescence. 

The crystal structure (Strop et al. 2001) reveals a homodimer with the zinc atom ligated by the sulfur atoms of two cysteines (Cys 32 and Cys 90) and the nitrogen atom of a histidine (His 87), as is the case for the plant-type enzyme (Fig. 11.3). The active site contains an HEPES buffer molecule in a position that implicates involvement of Asp 34 in the transport of protons after ionization of the zinc-bound water. 

Figure 3 Molecular relaxivities of liposomes with different Gd-containing membranotropic chelators. Liposomes (egg lecithin cholesterol chelator = 72 25 3) were prepared by consecutive extrusion of lipid suspension in HEPES buffered saline, pH 7.4, through the set of polycarbonate filters with pore size of 0.6, 0.4, and 0.2 mm. Liposome final size was between 205 and 225 nm. Gd content determination was performed by Galbraith Laboratories, Inc. The relaxation parameters of all preparations were measured at room temperature using a 5-MHz RADX nuclear magnetic resonance proton spin analyzer. The relaxivity of liposomes with polymeric chelators is noticeably greater because of the larger number of Gd atoms bound to a single lipid residue [16].

Figure 4 Transverse scan of axillary and subscapular lymph nodes in a rabbit 5 min postinjection of Gd-containing liposomes. Liposomes (egg lecithin cholesterol Gd-poly-NGPE = 70 25 5, 20 mg total lipid) were injected subcutaneously into the forepaw of anesthesized rabbit in 0.5 mL of HEPES-buffered saUne. Images were acquired by using a 1.5 Tesla GE Signa MRl scanner operated at fat suppression mode and Tj-weighted pulse sequence [16].

The amorphous state of the lyophilized solids was verified by powder X-ray diffractometry (Rigaku Denki Co., Ltd., Danvers, MA), and their Ca/P04 ratios after dissolution in HCl were determined by atomic absorption (AAS, Perkin Elmer, Norwalk, CT) and UV spectrophotometric [6] (Varian Analytical Instruments, Palo Alto, CA) measurements of Ca + and PO4, respectively. Dissolution of the ACP fillers was studied by kinetically following the changes in Ca + and PO4 concentrations in continuously stirred HEPES-buffered (pH = 7.4) solutions adjusted to 240 mOsm/kg with NaCl at 37°C. All solutions initially contained 0.8 mg/mL of the ACP filler. 

Each individual composite disk specimen was immersed in a 100 mL NaCl solution [HEPES-buffered (pH = 7.4), 240 mOsm/kg, 37°C, continuous magnetic stirring] for up to 264 h. Aliquots were taken at regular time intervals, filtered (Millex GS filter assemblies Millipore, Bedford, MA), and the filtrates analyzed for Ca + (AAS) and PO4 (UV). Upon completion of the immersion tests, the disks were removed, dried, and again characterized by XRD. Variations in the total area of disk surface (A) exposed were taken into account and Ca + and P04 values normalized to an average surface area of 500 mm. ... 

Figure 2 X-ray diffraction patterns of (A) the exposed front surfaee and (B) the shielded back surface of a P2O7-ACP/R 2 eomposite slab molded in a Teflon holder and suspended in a HEPES-buffered saline solution for 167 h at 37°C. The relative intensity (RI) values for (A) ranged from 230 to 570 counts/sec and for (B) from 90 to 800 counts/sec. The standard uncertainty of these values is 1/(3/ /)where 3 is the measurement time constant (in seconds).

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