60 Phenylalanine 
n 
oO 
Counting Efficiency (percent) 
rm 
° 
Oo 40 80 120 160 200 
240 
Suspension Concentration (mg/85ml) 
FIG.3. Self-calibration and internal-stand- 
ard counting curves for phenylalanine 
N 
fe} 
a 
fe} 
e "Cold" suspension plus 
internal C'* standard 
20 * c!4—- phenylalanine 
suspension 
Counting Efficiency (percent) 
| . 
900 1000 
1100 
Photomultiplier Voltage (volts) 
FIG. 4. Effect of light scattering and 
absorption on scintillation counting of 
suspensions 
TABLE 2—Counting Lyophilized and 
Wet Tissues 
Specific counting rate 
(dpm/mg) 
Tissue Lyophilized Wet 
Testes 18.8 + 0.6 19.0 + 0.4 
Liver We Ove ae 82.0 + 2.8 
Muscle 23.7 + 0.4 26.3 +0.5 
Spleen AD 2 Eee 44.54+1.6 
Sternum 19.8 +1.5 23.5 +0.8 
TABLE 3—Comparison of Suspension 
Counting with Conventional 
Specific counting rate 
(dpm/mg wet weight) 
Tissue Suspension Van Slyke-Folch 
Whole femur 18 + 2 19 +2 
Liver aiff ap 7] 40 + 3 
Testes ap il I 1 
Spleen 3) ae 444+5 
Thigh muscle 25 + 1 26 +1 
14 
sec for phenylalanine, 530 sec for bar- 
ium sulfate, 570 sec for calcium oxalate, 
and 310 sec for bacteria. 
Self-Absorption 
The suspended particle can be com- 
pared with the thin sample layer on 
a planchet. In the former case, there 
is 100% geometry between beta parti- 
cles escaping the solid and the detecting 
medium (the liquid scintillator). But 
the latter case finds about 50% geome- 
try the best realizable. A 1-mg/cm? 
BaCO; layer on a planchet produces 
a self-absorption factor of about 0.7 
(5). This corresponds to an average 
thickness of about 2.3 » and indicates 
very small effective particle size for 
the suspension with its smaller self- 
absorption. It was not established in 
this study whether the particles were 
actually as small as indicated or 
whether in wetting and partly per- 
meating the particles the liquid scintil- 
lator may have reduced their absorbing 
dimensions. 
The process of losing efficiency by 
self-absorption should act to degrade 
the beta spectrum such that the self- 
calibrated and internal-standard effi- 
ciencies will be the same for high ener- 
gies and different for low energies. 
Figure 3 is a plot of counting efficiency 
vs. photomultiplier voltage for C!*- 
phenylalanine and nonradioactive 
phenylalanine with added C!4-benzoic 
acid internal standard. At low volt- 
ages only the high-energy part of the 
C14 spectrum was being counted and 
the curves are almost identical. With 
increasing voltage the curves diverge in 
the direction of decreasing f as low C14 
energies rose above the lower dis- 
criminator and contributed to the 
counting efficiency. 
Light Absorption 
In earlier studies of homogeneous 
liquid scintillation counting of water 
samples, it was occasionally noticed on 
completing a count that the sample 
was very milky due to insufficient 
alcohol in the system. However, this 
opacity did not seem to interfere with 
counting. 
Similarly, in this study considerable 
opacity due to large quantities of 
a white suspension did little to inter- 
fere with the passage of scintillations 
from solution to photomultipliers. 
Figure 4 demonstrates this point for 
phenylalanine. 
The case of a colored suspension 
must be complicated by light absorp- 
tion. Whereas, with a white suspen- 
sion a photon will be reflected when 
striking a particle, the colored suspen- 
sion must absorb it. The lower curve 
in Fig. 4 shows this for highly colored 
liver tissue. 
C'-Analyses of Tissue 
The biologist needs a counting 
method for labeled animal tissues that 
is rapid, reproducible, and easy. The 
present method was compared in these 
respects with Van Slyke-Folch oxida- 
tion followed by BaCO; plate counting. 
Five pooled tissue homogenates pre- 
pared as described on p. 50 were made 
of bone, muscle, liver, spleen, and 
testes taken from rats injected with 
C!nitrogen mustard. Triplicate ali- 
quots of each homogenate were counted 
by each of the two methods and the 
averaged specific counting rates are 
compared in Table 3. The two meth- 
ods are essentially in agreement for all 
cases. 
The difference in operator time is 
significant. A rough estimate of oper- 
ator time on one sample of tissue homog- 
enate would be 100 min for the Van 
Slyke-Folch oxidation and BaCO; plate 
counting, 40 min for the scintillation 
counting of suspensions of lyophilized 
tissues, and 20 min for wet tissue 
homogenates. This estimate does not 
include time for the maintenance of 
equipment, etc., which probably would 
be about the same as operator time. 
BIBLIOGRAPHY 
D. L. Williams, F. N. Hayes, R. J. Kandel, 
W. H. Rogers, Nucteonics 14, No. 1, 62 (1956) 
. F. N. Hayes, R. G. Gould, Science 117, 480 
(1953) 
R. D. Hiebert, R. J. Watts, Nucueonics 11, 
No. 12, 38 (1953) 
J. R. Arnold, Science 119, 115 (1954) 
P. E. Yankwich, J. W. Weigl, Science 107, 
651 (1948) 
D. D. Van Slyke, J. Folch, J. Biol. Chem. 
136, 509 (1940) 
7, W.C. Pierce, E. L. Haenisch, ‘‘ Quantitative 
Analysis,"’ 3rd ed. p. 391 (John Wiley and 
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8. J. B. Niederl, H. Baum, J. S. McCoy, J. A. 
Kuck, Ind. Eng. Chem. Anal. Ed. 12, 428 
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9. N. H. Furman, ‘Scott's Standard Methods 
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Minneapolis, 1948) 
~ 
® 
2 AR w& 
Coming Next Month 
Scintillation Counting To- 
day, a NUCLEONICS Special 
Report on the Fifth Scintilla- 
tion Counter Symposium 
