Serum ethanol

By extraction of animal lymph glands, parotid glands, pancreas, liver, milt and blood serum with diluted acetic acid-ethanol-mixtures upon removal of fat and proteins. 

Red blood cells also contain sufficient acid phenylphospha-tase for mild hemolysis to cause false elevations. Therefore, inhibitors such as ethanol, formaldehyde, copper sulfate> and 1-tartrate have been used to inhibit selectively the enzyme of one or more tissues and enhance the specificity of the test (101). Ethanol is unsuitable because it inhibits the enzyme from erythrocytes and prostate simultaneously, and because it yields serum activities which correlate poorly with prostatic disease. Formaldehyde inhibits the erythrocytic enzyme and has been said to yield clinically satisfactory results. The copoper resistant acid phosphatase of serum is elevated by metastatic carcinoma of the breast, as well as by other metastatic cancers, and is also elevated by a wide variety of non-cancerous diseases. 

Solvents or their metabolites are commonly determined by GC (Tokunaga et al. 1974) or GC-MS. In spite of the high importance of exposure to solvents, and the great number of determinations performed worldwide, reference materials for solvents in serum or urine are virtually non-existent. There are a number of reference materials used in occupational hygiene, for example the ethanol in water standard from NIST (SRM 1828a) is commonly used in the clinical laboratory. 

The risk of gout increases as the serum uric acid concentration increases, and approximately 30% of patients with levels greater than 10 mg/dL (greater than 595 pmol/L) develop symptoms of gout within 5 years. However, most patients with hyperuricemia are asymptomatic. Other risk factors for gout include obesity, ethanol use, and dyslipidemia. Gout is seen frequently in patients with type 2 diabetes mellitus and coronary artery disease, but a causal relationship has not been established. 

Vasopressin is a peptide hormone produced by the hypothalamus and secreted by the posterior pituitary in response to stimulation. Normal stimuli for vasopressin release are hyperosmolarity and hypovolemia, with thresholds for secretion of greater than 280 mOsm/kg and greater than 20% plasma volume depletion. A number of other stimuli, such as pain, nausea, epinephrine, and numerous drugs, induce release of vasopressin. Vasopressin release is inhibited by volume expansion, ethanol, and norepinephrine. The physiological effect of vasopressin is to promote free water clearence by altering the permeability of the renal collecting duct to water. In addition, it has a direct vasoconstrictor effect. Consequently, vasopressin results in water retention and volume restoration. In patients with septic shock, vasopressin is appropriately secreted in response to hypovolemia and to elevated serum osmolarity (R14). 

Cells are maintained in suitable medium and starved of serum, if required. Agonist/inhibitors are added for 30 min first, then [35S]Met (5 iCi/ml medium) is added for 60 to 120 min. Cells are then lysed and cleared lysates (equal amounts of proteins) are loaded onto 3 MM papers. After the samples have been allowed to soak into the paper, the papers are boiled in 5% (w/v) TCA three times (2 min each time), rinsed with 100% ethanol, and oven dried. [35S]methionine incorporation is measured in 3 ml of scintillation fluid. 

Cohn, E.J. et al. (1947) Preparation and properties of serum and plasma proteins. XIII. Crystallization of serum albumins from ethanol-water mixtures./. Am. Chem. Soc. 69, 1753-1761. 

The sample materials from which proteins for proteomics studies may be extracted include fresh or snap-frozen cells from varied sources such as biological fluids, (serum, urine, plasma) and solid tissues such as biopsy specimens. Moreover, proteins isolated from ethanol-fixed paraffin-embedded tissues can be utilized for MS analysis.2 Protocols for the identification of proteins from formalin-fixed paraffin-embedded (FFPE) tissues have been recently developed.3 4 FFPE materials are the most common forms of biopsy archives utilized worldwide, and represent an important advancement for the large-scale interrogation of proteins in archival patient-derived materials. Finally, laser capture microdissected tissues have been successfully used for MS analysis.45... 

Prepare treatment medium containing various concentrations of test compound 19.7 ml of Eagle s medium (without serum) plus 300 pi of stock concentration of compound in a preferred solvent (e.g., water, ethanol, DMSO, etc.). The final concentration of solvent other than water should not exceed 1% v/v. Normally a range of 0-5000 pg ml-1 (final concentration) is covered. For a sparingly soluble compound, the highest concentration will be the lowest at which visible precipitation occurs. Similarly, if a compound has a marked effect on osmolality, concentrations should not be used that exceed 500 milliosmoles (mosm) per kg. In addition, a pH range of 6.5-7.5 should be maintained. 

X 10 2 M Tris HC1 buffer, pH 7.5, 2.4 X 10 3 M glucose 6-phosphate (G-6-P), 1.6 units glucose 6-phosphate dehydrogenase (G-6-P dH), 5.1 X 10"5 M NADP, and 2.7 X 10 3 M KC1. In addition, the following chemicals were included in the final concentration indicated 5.1 X 10-3 M NADH, 1% (W/V) bovine serum albumin (BSA), and 1.0 mg aldrin in 0.1 ml ethanol. Whole body homogenate experiments included all of the above chemicals unless otherwise noted. Reaction mixtures were incubated with swirling in test tubes at 30 - 1°C. Reactions in Steps 1-4 of the experimental sequence were stopped after 1 hr and Steps 6-8 after 15 min, by the addition of 2 ml 5% TCA. 

Figure 2.10. The dependence of the position of the fluorescence spectrum maximum on excitation wavelength for 2,6-TNS in model media (a) and in complexes with proteins (b). (a) 2,6-TNS (3 x 10-s) M in glucose glass at 20°C (1), glycerol at +1°C (2), and 80% aqueous ethanol at 20°C (3). Excitation spectra are for glycerol (4) and 80% ethanol (5). (b) 2,6-TNS in complexes with / -lactoglobulin (1), tetrameric melittin (2), human serum albumin (3), and lysozyme (4) at 20°C. Excitation spectrum (5) is for human serum albumin.

Protein concentration was determined using the Bradford assay at 595 nm. 100 pL of the sample were introduced into a cuvette containing 5 mL of Bradford solution (100 mg of Coomassie blue, 50 mL of ethanol and 100 mL of 85 % phosphoric acid dissolved in 850 mL of H2O). The solutions were incubated for 5 min at room temperature. The absorbance was measured at 595 nm. The protein concentration in the sample was determined using a calibration curve plotted with serum albumin (1 mg mL ) as a standard.)... 

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