Channels ratio counting

Figure 3-16. Channels ratio counting of quenched samples. Quenching increases sequentially from sample 1 to sample 4. Arrows above the figure indicate the range of the A and B channel discriminators. (Courtesy of Beckman Instruments, Inc.)...

In column 110, it is also theoretically possible that glycine com-plexed with the added humic acid and that it was sequestered in the aqueous phase of the Teflon eluate and bound to the Teflon bed. To test this explanation, a 1/10-scale parfait column was constructed and 4 liCi of 14C-glycine, 40 fig total, was applied in 800 mL of synthetic hard water (column 123). In this experiment, the alcohol and solvent 1 conditioning washes were combined with the standard eluates of each bed before counting. These solutions were not concentrated before counting. Quench correction was by the channels ratio method. 

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... 

Using the data obtained from samples 1 to 4, the known number of 14C disintegrations per minute added to each of these samples, and considering the counts per minute data obtained from the widest window as a reflection of overall counting efficiency (the total possible number of counts per minute that can be detected in each sample), construct a channels ratio quench correction curve similar to that shown in Figure 3-9. 

The channel 1/channel 2 ratio is a value that will be provided by the instructor. It is a correction factor that takes into account the fact that some 14C counts per minute were counted in channel 1. The instructor determined this ratio by scintillation counting of a sample containing a known number of 14C disintegrations per minute in channel 1 and channel 2 under conditions identical to your experiment. Refer to Section I, Experiment 3 for further discussion of the channel ratio method of quantifying radioactivity. 

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. 

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

What was the efficiency of counting and the absolute radioactivity (dpm) in the sample These data give a channels ratio of... 

In practice a sample is counted normally as described for the channels ratio method and then counted a second time with a source of y-rays (usually Cs or Ba) positioned in the center of the counting chamber just below the sample vial (see Figure 3-19). Any quenching that occurs has the same effect on the efficiency and spectrum of the Compton electrons as it does on those of the sample j8 particles. Since this technique has incorporated the procedures of both internal standardization and channels ratio correction methods, it is no surprise that the data obtained must be treated as described above for both of these techniques. The first or normal count rate obtained in each channel arises only from the sample and may be represented as follows ... 

With a statistically significant channels ratio in hand, it is only necessary to relate this ratio to a specific efficiency. In the channels ratio technique this was done by determining the ratios and efficiency of counting for a set... 

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. 

Counting Quenched Samples Using the Channels Ratio Technique... 

E. T. Bush, Anal. Chem., 35 1024 (1963). General Applicability of the Channels Ratio Method of Measuring Liquid Counting Efficiencies. 

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 radioactivity of samples was measured by liquid scintillation with toluene+POPOP or with Triton X-100 (Isocap-300).Counting efficiencies,determined via the channel ratio method and quench correction curve,were sufficiently constant (about 70-80%).Gas exchange measurements of respiration of leaf were conducted by manometric tech-nique(9).It was found that the chosen concentrations of exogenous substrates had no effect on dark respiration of leaves studied plants. 

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,. 

It has been established that SCR and ESCR cannot be used to compensate for quenching due to variations in water content in emulsion counting (Noujam et al.,1976). However the results discussed below show that these channels ratios can be used... 

The variation of channels ratios with volume in the vial has recently been reported (Knoche et al., 1979). It is essential when accurate counts are required to keep both the volume of counting mixture cind vial type constcint. Otherwise it is necess try to calibrate the liquid scintillation counter being used. 

Figures 1 and 2 illustrate the relationship between counting efficiencies of the beta standards and the channels ratios for channels A and B respectively. Background should be subtracted from each channel before computation of channels ratios for increased accuracy. In computing channels ratios for low activity samples, it is very important to use net rather than gross counts.

An initial measurement is made with each channel adjusted to 10% gain. The resultant channels ratio C/B is used to obtain the counting efficiency for channel B from Figure 2. 

The sample is then recounted with each channel set for 100% gain. By reference to the channels ratio A/C and Figure 1 the counting efficiency for channel A is obtained. 

Calibration curves for this method were constructed by use of single beta transitions only. Further tests of the method were necessary to determine the usefulness of the procedure for measurement of samples that contain radionuclides that emit X-rays, gamma rays, auger and compton electrons, and alpha particles. Table 1 compares those efficiencies predicted from the channels ratio calibration curves with the actual counting efficiencies for several radionuclides which decay by various means. Efficiencies obtained by counting with the wide-beta counter are listed for comparison. The predicted results are in good agreement with the actual efficiencies except in the case of and to a lesser... 

The channels ratio method is satisfactory for all types of quenching. It is also less time consuming since only one coxmt is required in two-channel scintiUation counter. Moreover, it is the most accurate method for counting efficiency determination. 

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