Channels ratio external standard

For general purpose tracer work, however, and particularly in polymer chemistry, the liquid scintillation counter surpasses all other instruments in its sensitivity and adaptability. There is no question on the author s mind that at the present time such an instrument would be the first choice, particularly where tritium, carbon-14 or sulphur-35 were involved. Samples for assay are dissolved in a phosphor whose major solvent usually consists of toluene, toluene-alcohol, or dioxan. Many polymers and low molecular weight compounds are readily soluble in these solvents. Prospective users should not be deterred by alleged complications due to "variable quench effects" as these effects are readily corrected for via internal or external standards or the channels ratio method (7, 46, 91). Dilution quench corrections, though valid, are tedious and unnecessary. Where samples are insoluble in phosphor they may be suspended (e.g. as gels or as paper cut from chromatograms, etc.) or they can be burnt and the combustion products absorbed in a suitable phosphor solution. A modification of the Schoniger flask combustion technique is particularly suitable for this purpose (43—45). 

OF THE RADIOACTIVITY CONCENTRATION Radioactivity measurements are carried out by the liquid scintillation counting procedure in -spectrometers using an external standard device which permitted the counting efficiency to be determined by the channel ratio method (explained for instance by Dyer (1980)). 

The channels ratio method makes use of existing counts within the sample vial. This method is suitable when large numbers of counts are present, but it becomes very time consuming with samples containing few counts, because a long time is required to accumulate sufficient counts for statistical accuracy. Most modern scintillation counters therefore employ an automatic external standardization system of quench analysis to avoid the time required for the internal channels ratio method. This method utilizes a specially selected external y radiation source carried in a lead-shielded chamber that is buried in the instrument. Before the regular counting of the sample, the external standard is... 

This experiment demonstrates two methods for analysis of quenching within samples, namely, the channels ratio method and the automatic external standardization method. Either or both of the methods may be demonstrated with the quench series of bottles described in the following protocol, depending on the capabilities of your particular scintillation counter. 

If the channels ratio method is used, proceed with Steps 3 to 8. If the automatic external standardization method is used, proceed with Steps 9 to 13. 

Three methods have evolved to ascertain the degree of efficiency loss both within the instrument and as a result of quenching. These techniques are termed (1) internal standardization, (2) channels ratio quench correction, and (3) external standard channels ratio quench correction. Determination of counting efficiency by internal standardization may be performed in two steps. The sample is first accurately counted followed by the addition of a precisely known quantity of radioactivity to the vial (50,000-80,000 dpm C or 100,000-150,000 cpm H). It is important for the amount of added radioactivity to be considerably larger than that originally present in the vial. The sample is then counted a second time. The first count is the sample cpm and the second count is the sample cpm + (efficiency)(standard dpm). That is. 

Dividing equation 3-22 by 3-21 gives the channels ratio for the external standard ... 

Figure 3-20. External standard channels ratio quench correction curves. These data were obtained using the procedures outlined in steps 3-50 to 3-57 of the experimental section.

Example. A sample of C was counted using the external standard channels ratio technique and the following data were obtained. 

Once these standard curves have been prepared the multiply labeled sample may be counted in the presence and absence of the external standard. The counting rate in channel B (cpm C). and the channels ratio value may be used in conjunction with the quench correction cutve (Figure 3-20) to calculate the absolute dpm of C using equation 3-24. 

Radioactivity Determinations of Multiply Labeled Samples Using the External Standard Channels Ratio Technique... 

Plot the efficiency (in channel A) of each quenched standard as a function of the external standard channels ratio value it yielded. This type of plot is shown in Figure 3-20. 

Tl. Takahashi, I. T., and Blanchard, F. A., Counting quenched liquid scintillation samples by using an outside-the-instrument gamma source and an external-standard channels-ratio method. Anal. Biochem. 35, 411-423 (1970). 

The applicability of the sample channels ratio (SCR) and external standard channels ratio (ESCR) for determining the efficiency of counting of quenched samples was studied using phenolphthalein, chloroform, and alkaline extracts of sheep s liver as quenching agents when counting CO,. 

The preparation of samples labelled with tritium always requires more careful attention than samples labelled with carbon-14. Furthermore, when the samples are low in activity, then a procedure which results in a heavily quenched sample may necessitate a prohibitive amount of counting. Quenching is probably the greatest obstacle in sample preparation and is the term applied to any effect which reduces the light output by the system. The determination of quenching is synonymous with the determination of counting efficiency. The most common methods used to determine counting efficiency are (1) internal standard (2) channels ratio, and (3) the external standard method. As these methods have been described in detail recently, they are not further discussed [74, 78, 205]. 

Usually more than two channels are required to determine the ideal quantification procedure, and the use of external and internal standards is the most highly recommended way of producing the calibration curve. The particular example of benzoic acid was taken to demonstrate how LOD was determined (analytical signal-to-noise ratio <2 1 Fig. 2) in combination with the low variability observed between replicate analyses. Fig. 2 also clearly illustrates the establishment of minimum and maximum concentrations in order to define the response linear range for each compound using the UV detection system involving single A. Comparative measurements were taken to... 

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