Optical density units

A concentrated Ca-D-pantothenate solution (1 mg/ml) is prepared in distilled water and dilutions made as needed. Refrigerated solutions are stable for 6 months. Pantothenate is added at 5, 10, 20, 40, 60, 80, and 100 mpg/ml final concentrations the control flask consists of basal medium alone for estimation of carry-over error—i.e., the pantothenate activity of the inoculum. The details of aseptic technique have been discussed elsewhere (H18, H19). Growth is measured in optical density units with a Welch Densichron, equipped with a red-sensitive probe to minimize blank readings due to the color of the medium. 

Growth measured in optical density units with a Welch Densichron. 6 Equal concentration of each total is twice the value given. 

One optical density unit change is equivalent to 311.5 fig/ml cellular dry weight. Initial and final pHs are given at the beginning and end of each curve. 

Two UV detectors are also available from Laboratory Data Control, the UV Monitor and the Duo Monitor. The UV Monitor (Fig.3.45) consists of an optical unit anda control unit. The optical unit contains the UV source (low-pressure mercury lamp), sample, reference cells and photodetector. The control unit is connected by cable to the optical unit and may be located at a distance of up to 25 ft. The dual quartz flow cells (path-length, 10 mm diameter, 1 mm) each have a capacity of 8 (i 1. Double-beam linear-absorbance measurements may be made at either 254 nm or 280 nm. The absorbance ranges vary from 0.01 to 0.64 optical density units full scale (ODFS). The minimum detectable absorbance (equivalent to the noise) is 0.001 optical density units (OD). The drift of the photometer is usually less than 0.002 OD/h. With this system, it is possible to monitor continuously and quantitatively the absorbance at 254 or 280 nm of one liquid stream or the differential absorbance between two streams. The absorbance readout is linear and is directly related to the concentration in accordance with Beer s law. In the 280 nm mode, the 254-nm light is converted by a phosphor into a band with a maximum at 280 nm. This light is then passed to a photodetector which is sensitized for a response at 280 nm. The Duo Monitor (Fig.3.46) is a dual-wavelength continuous-flow detector with which effluents can be monitored simultaneously at 254 nm and 280 nm. The system consists of two modules, and the principle of operation is based on a modification of the 280-nm conversion kit for the UV Monitor. Light of 254-nm wavelength from a low-pressure mercury lamp is partially converted by the phosphor into a band at 280 nm. 

Figure 2. Calibration curve for the Daresbury Optronics P-1000 microdensitometer by using Ilford Industrial G x-ray film. The figures above the data points indicate the nominal optical density of each measurement, assuming that a scanner reading of 255 corresponds to three optical density units.

Adsorption onto activated charcoal (Norit A) has been used for small oligonucleotides, up to tetranucleotides (Crane and Lipman 1953 Mandeles and Kammen 1966). The method is quicker and more convenient than DEAE-cellulose chromatography. Mandeles and Kammen (1966) used Norit A suspended in 1 mM phosphate 1 mM pyrophosphate buffer at pH 6.0 at a concentration of 100 mg/ml. The nucleotide solution is adjusted to pH 3 and 5 mg Norit for every optical density unit ( 40 pg) of nucleotides is added. The charcoal is collected on a filter paper and washed with water. (A good quantitative measure of or radio-activity may be obtained by counting the dry filter paper in a Geiger-Muller thin window counter.) The nucleotide material is eluted with a small volume of a mixture of ethanol, water and ammonia (600 400 6.5 v/v/v) which is then removed by drying under reduced pressure at 40°C. 

Table 1. Comparison of the growth of recombinant Saccharomyces and the parent yeast in glucose and xylose media. The units used are Klett Units, i.e. the optical density units measured by a Klett-Sm-nmerson photoelectric colorimeter ...

Spectrophotometry DNA concentration can be determined by measuring of the absorbance at 260 nm (A260) hi spectrophotometer using a quartz cuvette. For greatest accuracy, readings should be between 0.1 and 1.0. An absorbance of one optical density unit (1 OD) at 260 nm corresponds to 50 pg/ml of genomic DNA (see Table 3.6). This relation is valid only for measurements made at neutral pH therefore, samples should be diluted in a low-salt buffer with neutral pH (e.g. Tris-HCl, pH 7.0). To measure multiple samples, the cuvettes must be matched. 

KU = Klett Units, the optical density units measured by Klett-Summerson photoelectric colorimeter. 

The FTIR spectra were measured on a Shimadzu FTIR-8300 spectrometer over a range of 700-6000 cm-i with a resolution of 4 cm i and 100 scans for signal accumulation. Before spectra recording, powder samples were pressed into thin self-suppwrting wafers (8-30 mg/cm2) activated in a special IR cell under chosen conditions and further in vacuum (p < 10-3 mbar). FTIR sp>ectra are presented in the optical density units referred to a catalyst sample weight (g) in 1 cm cross-section of the light flux. 

The concept of absorbance units at 260 nm, A bo (aIso known as optical density units. ODU) often causes confusion for students. By definition, one unit is the amount of sample that gives a UV absorbance at X = 250 nm of 1.0 if the sample is dissolved in 1 ml volume and measured in a l-cm pathlength cuvette. Since aborbance is proportional to concentration (Beer s Law) and the volume is 1 ml, the number of Ajsq units of a sample is proportional to the number of moles of strands. Knowledge of the number of A260 units is important for planning UV melting experiments to optimize sensitivity. 

Fig. 3. Phosphate standard curve for the malachite green assay. The assay is linear over the range 1-200 iiM phosphate. Maximum sensitivity is limited by the optical density units on the spectrophotometer, which declines above 2.5 units. The inset shows the standard curve for a typical GTPase assay and is linear in the assay sensitivity range. All values are n = 3 and standard deviations are shown, but are normally smaller than the symbols.

To determine kinetic and thermodynamic parameters involved in these equilibria by direct relaxation measurements, very sensitive and rapid T-jump apparatuses appeared to be necessary. This objective can be attained by a fast Raman laser T-jump (time resolution about 10 s) and by a repetitive microwave T-jump (sensitivity better than 10 optical density units). 

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