The amount of kinetic energy in a beta particle differs from one decay to the next. However, each radioisotope has a typical energy spectrum, that is, a predictable range of energies. Typical energy spectra for different radioisotopes can be radically different in shape and magnitude, as with the commonly used 14-C and 3-H.
In liquid scintillation counting, the material containing radioisotopes is dissolved in an organic solvent containing an aromatic solute (the scintillant). When radioactive decay takes place, the energy of a beta particle is transferred by collision to an electron in the shell of the scintillant, exciting that electron. The electron then returns to its ground state, releasing a photon. The energy transfer can be from beta particle to solvent to scintillant, or directly to the scintillant, and usually there are multiple collisions per b particle. The number of photons emitted following each atomic disintegration is proportional to the energy of the released beta particle.
Now here is how the detection system works. The vial is lowered into a dark chamber with photoelectric detectors on each side. Each "flash" received by the detectors corresponds to one atomic disintegration. The detectors are connected, via photomultiplier tubes, to a microprocessor unit that records not only each event, but also the number of photons detected during each event (brightness of the flash). At brief intervals, such as 1/100 second, the instrument calculates the number of flashes per unit time, displaying them as counts per minute, or CPM.
No detection system perfect, of course. In liquid scintillation counting, cosmic rays, beta particles from decaying potassium in the glass vial, spontaneous discharges from the very sensitive photodetectors, and chemicals dissolved in the scintillation fluid all can contribute to spontaneous flashes of light that are recorded as counts. The CPM attributable to such sources are called background. Background counts are often so low relative to the activity being measured that they are ignored. However if the number of "real" counts is low, background counts can contribute to experimental error. It is a good practice to include a vial containing everything except added radioactivity as a control to determine the background level. Background CPM are then subtracted directly from the CPM for the experimental samples.
Unfortunately, there are two more little problems. No instrument is capable of recording all of the atomic disintegrations within a scintillation vial. Because of the geometry of the vial and photoelectric detectors some events go undetected. The maximum efficiency with which a low energy emitter such as tritium can be detected is about 70%. Worse yet, the energy of some photons is absorbed by chemicals in the solvent before the photon can reach the detector. The latter phenomenon is known as quenching. With the chemical quenching that is typical of most experiments, the usual counting efficiency for tritium is 30 to 40%, and sometimes much less. The amount of quenching can vary from sample to sample, therefore it is often necessary to estimate the efficiency of counting for each individual sample.
Remember that the amount of light detected is proportional to the energy of the beta particle that was released by the disintegrating nucleus. When you prepare to count samples, you select appropriate "windows", that is, ranges of light intensity that the instrument will record as counts. The instrument records the amount of light detected following an event, and if that amount is within the energy range for a particular window, the event is recorded as one count. It is ignored if it falls outside the selected range. Each window is given a channel number, and the count for each window is given as CPM for the corresponding channel.
As quenching takes place, the energy recorded for each event is less than it would be for an unquenched sample, since for each event the energy of some photons is absorbed before detection is possible. The energy spectrum is shifted to the left, and the greater the quenching the greater the shift. As the shift takes place, the ratio of counts in channel 2 to counts in channel 1 becomes smaller. That ratio is known as the sample channels ratio, or SCR.
Counting efficiency is positively related to the SCR. To get a conversion factor between SCR and efficiency, equal known amounts of the isotope are added to a series of vials. Progressively greater amounts of a quenching agent, such as carbon tetrachloride, are added to each vial. The vials are counted and CPM is divided by known DPM to get the fractional efficiency of counting. For example, if a vial with 20,000 DPM of tritium yields 5,000 CPM, the fractional efficiency of counting was 5,000/20,000 = 0.25, that is, 25% of the atomic disintegrations were detected. Fractional efficiency is plotted versus SCR to yield a quench curve. The instrument prints out CPM and SCR for each sample, therefore to get actual DPM in a sample the investigator must (1) subtract background CPM from the CPM for the sample, (2) determine fractional efficiency from the SCR for the sample, and (3) divide net CPM by the fractional efficiency.
Example. You are counting samples containing 14-C. Sample #1 gives you channel 1 CPM of 1323. The background counts were 23 CPM and the SCR was 0.5. From the quench curve, an SCR of 0.5 corresponds to an efficiency of 0.93. Then the estimated amount of radioactivity in the sample is (1323 - 23)/0.93 = 1398 DPM.
There are other problems associated with the measurement of radioactivity that are not so easily solved. However, for single label experiments the quench correction is all you need. Review the special safety cautions for radioisotope work, and you are ready to go.
Modern scintillation counters have a conveyor system that automatically feeds samples in order into the counting chamber. As each sample is counted the relevant information is printed. A typical printout includes preliminary information followed by specific information on a sample by sample basis. Every manufacturer uses a different system of terms and abbreviations, so either you will need access to the instrument manual or the printout from your instrument will have to be translated by an experienced individual.
Preliminary information may include the date, time, user i.d., and program selected. Common counting parameters are usually listed. The parameters may include: number of times each sample is counted; time period of counting each sample; number of times the entire set of samples is counted; type of quench correction; windows selected; special features such as criteria for cutting short a count if counts are very high, or automatic background subtract. The preliminary information must indicate that the counting parameters were appropriate for your type of samples.
The information supplied for each sample typically includes: sample i.d., which may be position number in a rack, position in order of counting, or both; channel numbers (windows) and corresponding counts per minute; time of counting; elapsed time since the run was started (this is important for radioisotopes with very short half lives); sample channels ratio or other measure of quenching.
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