Water blank

To prepare standards and to estimate the procedmal blank of the method, water is needed with a zero or very low carbon content. Several methods have been suggested by different workers to prepare carbon-free water, including multiple distillation with oxidants and UV treatment (see also Chapter 16). Several manufacturers now offer equipment producing water with very low carbon concentration (3-5 figfL). The difficulty then is to maintain such purity during transport from the water producing unit to the analyser. Carbon blanks of 20 pg/L are low enough for most purposes. 

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Filtration Test. The filtration test equipment consisted of a glass column, with a fritted disc 2 cm. diameter, fitted with a stopcock and a 25 ml graduated cylinder. After mixing the MCC sols with electrolyte solution, the mixture was immediately poured into the glass column with the stopcock closed. The time required to collect water (blank) and filtered sol between 5 to 25 mis in the graduated cylinder after 2 hrs. was determined. The fritted glass was backwashed with water thoroughly after each measurement and a new blank was established. The relative filtration time between sample and water was used to compare differences between samples. 

Blank Nolse/Drllt -Water Blank 1 1 NoiseDrift ... 

The first experiment of this set (row 1) is used to establish the noise and drift characteristics of the detector. 1 mL/min water (with 1% methanol) is pumped through the system. The system is allowed to equilibrate then, a 1 pL water blank is... 

The precision of MS assays is in the range typical of most clinical assays (i.e., under 5-15%). The best choice of internal standard is the stable-isotope-labeled form (preferably 13C) of the compound of interest (e.g., P-hydroxy myristic acid or muramic acid). Specific trace detection of chemical markers in complex matrices requires appropriate negative controls. Procedures are often described that do not employ the mass spectrometer and false positives are often reported. The mere analysis of blank filters or water blanks is not satisfactory since chemical noise contributed by the sample is much greater and is not accounted for with this form of control. 

Add 1 ml of DMB reagent to 100 pi of demineralized water (blank) or 100 pi of urine in a cuvette. In addition, 100 pi of each urine should be added to 1 ml of formiate buffer to assess the absorbance of the pure sample (sample blank). Each sample should be measured against pure formiate buffer (buffer blank, when measured separately). The color of the GAG-DMB complex is not stable over time, so that assay conditions have to be strictly standardized. Urine, standard, or water is added to all cuvettes first. The DMB reagent must then be added swiftly. All cuvettes are mixed with a spatula (10 x) and after 3 min samples are measured at 520 nm. When the absorbance exceeds the linear range, the urine has to be diluted. It is also recommended to measure all samples in duplicate using 50 and 100 pi of urine. This allows an evaluation of the plausibility of results. 

In general, most procedures are like the ones described above. For the modified method, an alternative DMB reagent is used and ten parts of this reagent are added to one part of Tris buffer to yield a reagent with a pH of 8.7-8.8. The reagent - Tris mixture is not stable and has to be prepared just prior to analysis. First, 20 pi of urine (sample) or water (blank) is added to the cuvettes or microtiter plates. The volume of standards, samples, and blanks are adjusted to 50 pi with demineralized water. Then, 275 pi of DMB-Tris reagent is added to each cuvette/well. Either all samples... 

All determinations are performed in duplicate. For total (free plus bound ) sialic acid take 50 pi sample (distilled water blank or standard or urine sample ), add 50 pi H2S04 and incubate for 60 min at 80°C in closed reaction tubes. For free sialic acid, pipette the same without incubation at 80°C. 

Dowex (2.5 g per column) is prewashed in a sintered glass funnel with distilled water. Disposable columns (5 ml) are filled with 2 ml bed volume of prewashed Dowex. Columns are washed with 6 ml ice-cold distilled water. A 60-pl aliquot of each standard (NeuAc-2, NeuAc-5, and NeuAc-10) and a water blank are applied to the column. For diagnostic samples, patients and controls apply 200 pi to the column. Wash all columns three times with 2-ml ice-cold ammonium acetate buffer. Elute NeuAc by applying 3 x 2 ml and 1 x 1 ml ice-cold ammonium formate buffer. Close the collection tubes (50-ml volume) with a pierced cap and centrifuge fast to spin down droplets from the wall of the tube. Freeze the eluates in an upright position in liquid nitrogen. 

Resin Blank Artifacts Effect of 2 ppm of Residual Chlorine. Three basic types of blank experiments were performed. One was performed to identify any artifacts caused by the presence of 2 ppm of chlorine in blank water used in the separation-concentration procedure, and another was performed to identify any artifacts in the general resin procedure. In addition, a reagent blank was also concentrated and analyzed. The reagent blank was performed in a manner identical to the pure water blanks, except that no resin was included in the procedure. This reagent blank gave an indication of contaminants arising from sources other than the resin (i.e., glassware, water, solvents). 

Clean Water Blank Experiment. This experiment was performed for the purpose of identifying any artifacts that might arise from the resin in the course of normal resin experiments. Four replicate resin blanks were run by the normal resin separation-concentration procedure. Data from this experiment yielded the following conclusions ... 

Read light absorbance at 550 nm with a water blank. 

Dissolve 1 g of KI in the above solution. Add 10 mL of 1 4 H2S04. Allow the solution to stand in the dark for 5 min. Titrate this solution with standardized sodium thiosulfate or phenylarsine oxide solution, using starch indicator. Add the indicator when the solution becomes pale straw following the addition of Na2S203 or PAO. At the end point, the blue color disappears. Disregard any reappearance of blue color. Run a distilled water blank. 

Allow the solution to stand for 10 min for maximum color development. Read the absorbance at 630 nm after zeroing the spectrophotometer to reagent blank. Run a distilled water blank following the same procedure. Prior to the sample analysis, prepare a calibration standard curve following exactly the above procedure. Run one of the standards through the procedure with each batch of sample. 

Repeat the above steps exactly in the same manner and read absorbance for a distilled water blank and four S032- standards. The stock standard should be standardized by titrimetic method to determine the molarity of S032. Prepare a calibration curve plotting absorbance vs. microgram sulfite. Run at least one standard for each batch of samples. 

Fig. 6.5 Diagram of system set-up for postcolumn infusion test for matrix effect. The analyte in the mobile phase was infused by a syringe pump at about 10 pL/min. The blank matrix extract or the test control (mobile phase or water blank extract) was injected into the analytical column. The effluent from the analytical chromatographic column was mixed...

A standard solution containing 40 mg P F1 prepared by dissolving potassium dihydrogen phosphate (0.1757 g) in about 900 ml water and adjusting to 11. This solution is then diluted to give standards of 5,10,25,50,75,100 pg P H, and water blanks are also prepared. 

Identification Mix 333 mg of sample with 44 mL of water, 0.15 mL of a 1 10 sodium hydroxide solution, and 0.2 mL of ammonium hydroxide, warm to dissolve, and dilute to volume with water in a 500-mL volumetric flask. Pipet 10.0 mL of this solution into a 250-mL volumetric flask, dilute to volume with water, and mix. The resulting solution exhibits absorption maxima at 520 nm and 550 nm when determined in a 1-cm cell with a suitable spectrophotometer against a water blank, and the absorbance at 520 nm is not less than 0.30. Assay Not less than 50.0% of carminic acid (C22H2o013), calculated on the dried basis. 

Calibration Determine the absorbance of each Standard Solution in a 1-cm pathlength cell at 490 nm against the water blank. Calculate the slope of the curve obtained by plotting absorbance versus micrograms per milliliter of lactose. The slope of the curve is the absorptivity (a) of the lactose-reagent product. 

Determine a Water Blank as well as a Cold Blank by the same procedure, but for the Cold Blank, allow the flask containing the sample solution to stand at room temperature for 10 min rather than placing it in the boiling water bath. Calculate the percent of invert sugar by the formula... 

Prepare a water blank by pipetting 10 mL of equilibrated water into the wide arm of the viscometer. Determine the time (Tw) in seconds required for the meniscus to fall between the two marks. Use an average of five determinations for (7V). 

Titrate another 20-ml H20 sample (a water blank containing no unknown) with 2 N H2S04 1 drop at a time for the first 10 drops, 2 drops at a time for the next 10 drops, and finally 4 drops at a time until the solution reaches pH 1.0. Record the pH and volume of acid delivered after each addition. 

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