Liquid scintillation counting is the preferred method for the
measurement of low-energy beta-emitting radioisotopes commonly used in
cell-fractionation studies (see Figure 18). It is convenient, sensitive,
and rapid for routine measurement of radiation in hydrocarbons, other
organic compounds, and aqueous solutions containing such isotopes as ³H,
¹⁴C, and ³²P.
Liquid scintillation solutions share with other scintillating materials
the property of converting into visible light the energy deposited in
them by ionizing radiation. In theory, if a sample of a beta emitter is
dissolved in a liquid scintillator solution, every beta particle emitted
will be absorbed completely because the range of penetration of beta
particles in liquids is quite short (ranging from 0.008 millimicron for
³H to 7.9 millimicrons for ³²P in a medium of unit density). The kinetic
energy of the beta particles is largely used up in the ionization and
excitation of the most abundant molecular species present, the solvent
in which the scintillating material was dissolved. A fraction of the
energy thus expended by each beta particle is transferred from excited
solvent molecules to scintillator molecules; thus the electrons in the
atoms of the scintillator molecules are raised to an excited (higher
energy) state. When these electrons return to the ground, or unexcited,
state, a fraction of them emit a photon of light. Thus each beta
particle produces a burst of photons.
[Illustration: Figure 18 _Technician placing a tray of samples in a
liquid scintillation counter. The radioactivity of each sample is
recorded as the trays revolve._]
If a vessel containing the liquid scintillator and the radioactive
sample is placed near a suitably sensitive instrument known as a
photomultiplier tube, each burst of scintillator photons activates this
device and causes it to release a burst of photoelectrons. Each burst of
photoelectrons is multiplied successively in a series of electronic
steps; as a result, there is a suitably large electrical-output pulse to
be recorded.
One of the principal advantages of the liquid scintillation method is
the ease of sample preparation. We need only transfer a known volume of
a liquid sample or weigh a given mass of a solid sample into a sample
bottle, add a known amount of the liquid scintillator solution, and stir
until there is a homogeneous solution. Samples thus prepared are placed
in a refrigerated counting apparatus. After a short waiting period to
allow time for the samples to cool and for a natural, short-lived
phosphorescence (due to exposure to room light) to subside, the samples
are ready to be measured.
[Illustration: Figure 19 _Placing radioactive samples in a refrigerated
unit for liquid scintillation counting._]
One disadvantage of liquid scintillation counting is that different
compounds show different degrees of quenching (loss of emitted photons),
and the effect must be checked for each class of compounds in each
concentration range. This checking is usually done with an internal
standard technique, the sample being counted before and after a
standard, or known, emitter is added.
Another difficulty is that the best scintillating solvents are not the
best chemical solvents for most biological materials. The solubility
problem is also aggravated by the low temperatures at which liquid
scintillation counters are usually operated for more effective
instrument performance.
With the method we have described, we can obtain a fairly accurate idea
of the rate of RNA synthesis in a given tissue. There are other things
we would like to know about RNA. The first of these is the kind of RNA
being synthesized. During alkaline digestion all kinds of RNA are broken
down into their component nucleotides; we must therefore use other
methods if we wish to know the kind of RNA in which the radioactivity of
the precursor has been incorporated.