TECHNICAL SECTION

THE USE OF CONTROL CHARTS IN THE CLINICAL LABORATORY*

STANLEY LEVEY, P H . D . , AND E. R. JENNINGS, M.D.
From the Departments oj Physiological Chemistry and Pathology, Wayne University College
of Medicine, and Receiving Hospital, Detroit, Michigan

Constant supervision is necessary in order to obtain uniformly reliable values

in a busy clinical laboratory. Normally the director of the laboratory does not
have sufficient time to carry out a detailed supervision of the methods but, by
the use of control charts, it is possible to determine at a glance whether the
errors of analysis are beyond the statistical variation of the procedures employed.
Quality control by statistical methods is widely used in industry 7 to determine
whether the variation observed among items produced by a single machine or
an entire process is consistent with the hypothesis that a stable system of chance
causes is operating. This hypothesis may be tested by taking a number of groups
of observations at reasonable intervals of time, to see if the variations among
the averages of each group are consistent with the variations observed within
groups of observations made at the same time. The details of this technic are
furnished by the theory of statistical quality control. 1
In this laboratory the principle of the control chart provided a constant
check on the reliability of the numerous determinations run each day. I t made
it possible to distinguish between what might be termed statistical fluctuations
and actual error. In addition it offered a rational basis for action in correcting
a defective procedure. While the chart will show when a method is out of con-
trol, i.e., that the variation is greater than would be expected if chance alone
were operating, it remains for the analyst to study the cause and prevention of
the error. Since control-chart methods have not been widely used in the clinical
laboratory, we undertook a study of their application in our laboratory.

METHOD

In setting up the type of control chart used here, it is necessary to have

sufficient amounts of homogenous blood or plasma in which the concentration
of the material to be analyzed is stable over a long period. Also, the concentra-
tion of the substance estimated should be approximately in the range of normal
blood values, so that no special analytical steps would be required to check
its level.
The pooled blood or plasma derived from this blood was obtained from lots
discarded by the hospital blood bank. Whole blood was used to control the
urea method, and plasma was used to study the total protein, albumin, globulin
and chloride determinations. The blood or plasma was pooled and then dis-
tributed as 5-ml. samples in a large number of small test tubes. The latter were
well corked, stored in a "deep-freeze" cabinet maintained at —10 C. and kept
frozen solid until ready for analysis. Sufficient samples were prepared at this
* Received for publication, May 15, 1950.
1059

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 1060 LEVEY AND JENNINGS

time to last approximately one year. On the morning of the analyses two samples
each of plasma and whole blood were removed from the freezer and thawed at
about 30 to 35 C.
For the control of the analyses of the carbon dioxide combining-power of
plasma, the material obtained directly from the blood bank was not suitable
since its carbon dioxide combining-power was approximately 12 volumes per
100 ml. plasma. This low value was partially due to the citric acid content of
the preserving fluid. Sodium carbonate was added to the plasma in sufficient
quantity to bring the carbon dioxide combining-power up to about 60 to 80
volumes per 100 ml. The plasma thus fortified was distributed in many small
test tubes and stored in the frozen state.
Since the true value for the concentration of any of the control substances
was not known, an estimate of the true value had to be obtained from the data.
This estimate was obtained by averaging the individual values obtained from
the first 20 pairs of samples analyzed. These values were obtained over a period
of about a month so that any day-to-day factors influencing the analytical pro-
cedures were minimized. For the quality control study two samples for each
determination studied were sent twice a week (usually Tuesday and Thursday)
to the laboratory where they were numbered and treated as routine samples.
It was most important in this work to make sure that the test samples were not
given any preferential treatment. After the analyses were completed, the aver-
age value of the two samples and the value representing the difference in the
two results (the range) were plotted on the control chart.
For the preparation of the control charts, the mean value of any one pair of
determinations was plotted as the ordinate, with the date or order of analysis
as the abscissa. In charting the variation between duplicate analyses, the range
(the difference in values of duplicates) was plotted as the ordinate and the date
or order of the analyses as the abscissa. The statistical limits for the mean and
range were also placed on the control chart. Using two samples per determina-
tion, the limits for the mean and range are ± 1.88 X fi and 3.27 X R, respec-
tively. The value of R was obtained by averaging the range values of the first
20 pairs of samples studied. 1 The control limits for the mean and range are
approximately equivalent to 3 a limits. This means that, if a stable system of
chance causes is operating, only 3 out of 1000 observations should fall outside
the 3 o- limits. If an observed average or range is beyond the limits given above,
it is taken as evidence of lack of control since it has such a small probability
of occurring under controlled conditions. Examples of the use of the control
chart in the clinical laboratory are given below.
RESULTS
Urea Determination
The Karr Direct Nesslerization procedure was employed for the determination
of urea in blood.2 The control chart for this determination is given in Figure 1.
I t can be seen that on days 5, 6, 7 and 10 the values for the urea nitrogen were
far out of control (exceeded the statistical limits assigned to the method).

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 CONTROL CHARTS IN THE LABORATORY 1061

Because of the large number of urea determinations (up to 80) run per day, two
technicians usually worked as a team after the ammonia solutions were nessler-
ized, one technician matching the colors and the other recording the values on
the reports sent to the wards. Since only one technician was working on the
determinations during these days, it took considerably longer to complete a
series of tests. Under these conditions the test samples which were placed near
the end of the series developed turbidity at a more rapid rate than the standard.
By reducing the size of a run or having two persons work on the determinations,
this error was eliminated. On days 13, 19, 26 and 27 it was found that the average

Mean Value
mg. /IOO ml. Urea nitrogen
Blood
22|-
20 h
18
I6h
14
12
10'
8

Range (2 samples per determination)

4i-

_L _L -L
I 10 15 20 25 30 35 40
Order of analysis
FIG. 1. Control chart for the urea nitrogen determination. The values in the shaded
area are in control. The heavy line represents the mean value.

values for the urea determinations were again out of control. During the last
two periods mentioned, an intensive study of the variables in this method was
made and it was concluded that cooling the samples before nesslei'ization, a
process used all summer, brought the values into control. Cooling of the samples
before nesslerization had previously been done only during hot weather.
The allowable variations in the control of urea are relatively small from the
clinical point of view, being only ± 1.6 mg. per 100 ml.* During the entire study
* The control limits used in this stud}' are 3<r limits which were derived by calculations
from the variations of the range values. In many of the charts, it is difficult to ascribe
so many zero ranges to the operation of pure chance. The cause of this discrepancy appears
to be the lack of sensitivity of the analytical methods for the mathematical procedures
employed. There is probably considerable variation among samples listed as having a

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 1062 LEVEY AND JENNINGS

the range of the duplicate values was never beyond the limits of control. While
the mean urea values might be far beyond the control limits (21 mg. per 100 ml.)
the duplicates were only 1 mg. per 100 ml. apart. The range, therefore, was well
in control. From this chart one notes that high values were obtained when the
Karr urea method was out of control.

T
Mean Value i I r
meq./liter Plasma chloride
Plasma
70i-

15 20 25 30 35
Order of analysis
FIG. 2. Control chart for the determination of the chloride content of plasma. The values
in the shaded area are in control. The heavy line represents the mean value.

Plasma Chlorides
The control chart concerned with the chloride concentration of the pooled
plasma is shown in Figure 2. The method used for all the assays was the mercuric
nitrate titration of Schales and Schales.6 I t should be noted that the chloride
content of the test samples was markedly lower than the normal for blood.
This is probably due to the dilution of the whole blood at the blood bank with
a dextrose-citrate preserving solution. The determinations appeared to be in
control up to day 9 (Fig. 2). On this day the technician who normally conducted
the test left for her vacation. The new analyst, using the titration factor that
the regular worker had determined earlier, reported values which showed the
method to be out of control. On the fifteenth day the new technician determined

zero range value, but the method does not show it. This is similar to the variation which
would occur in measurements of length of 0.01 mm., when the gauge employed was accurate
to only 0.1 mm. Thus it should be recognized that the computed limits for the mean values
are probably narrower than the true 3a- limits.

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 CONTROL CHARTS IN THE LABORATORY 1063

the titration factor herself and the values she reported immediately fell in line
with the previously controlled data. The original technician returned on the

~~I I I
Mean Value
gm./lOO ml.
Plasma Total plasma protein
7r .

U
5
Range ( 2 samples per determination )
1,0
T
a5
ll!llllllllllllllllllllllllllllllUII!llllll!«llllll
0 |1U11U11III1U11UIIU1

_L J_
10 20 30 40
Order of analysis

1 1 1 1 1 1 1 1
Mean Value
g m . / K DO ml. Plasma albumin
Plas Tia
5_ • • -

4
_ •
3<
Range ( 2 samples per determination )
I.Op
0.5 h

I I JL I _L _L
I 10 20 30 40
Order of analysis
F I G . 3. Control charts for the determination of total plasma protein and albumin. T h e
values in t h e shaded area are in control. T h e heavy line represents the mean value.

nineteenth day and, with the titration factor she had determined, all analyses
were within the predicted limits until the twenty-eighth day, at which time it
was suggested to the operator that the titration factor was low. When this was
found to be true upon checking, a new factor was determined and used in all
succeeding analyses.f The factor of the reagent is now rechecked weekly. Also,

t On examination of Figure 2 there appear t o be two population groups present, one

covering days 16 to 25 and the other consisting of all values beyond day 25. Using a mean
of 64.2 mEq. per liter for the first group and <r of 1.11, one finds t h a t the 3<r mean range

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 1064 LEVEY AND JENNINGS

because of individual- variations in evaluating the end point of this titration,

each analyst determines a titration factor herself. While there was some varia-
tion of the average values beyond the control limits, the differences between
duplicate samples analyzed at the same time were always within definable
limits.

Plasma Proteins and. Albumin

The determination of the protein content of the pooled samples of plasma
was accomplished using the biuret reaction and the salt fractionation of Kingsley.3
I t should be noted that the allowed variation of the control limits is only ±

Mean Value Carbon dioxide

Vol./100 ml combining power
Plasma
90

15 20 25
Order of analysis
FIG. 4. Control chart for the determination of the carbon dioxide combining-power of
plasma. The values in the shaded area are in control. The heavy line represents the
mean value.

0.25 Gm. per 100 ml. for the total protein and ± 0.17 Gm. per 100 ml. for
albumin (Fig. 3). On days 32 and 33, two consecutive mean values were
beyond the control limits. In addition, one set of determinations showed a range
difference too great to be a controlled variation. No explanation for this lack
of control could be found in spite of a rather thorough investigation of all the
reagents and the use of new standards. On day 34 the analyses spontaneously
came back into control and remained in control. The reason for this could not
be ascertained. Since the determination of the protein content of serum or
plasma is an analysis in which many errors can occur which are unknown to the
analyst, quality control is a valuable tool for the over-all policing of this method.
derived from the data to be 60.9 to 67.5 mEq. per liter. The second group had a mean of
61.8 and a a- of 0.84; thus, the 3<r limits would be 59.3 to 64.3 mEq. per liter. The discrepancy
between these mean limits and those derived from the range values given in Figure 2 is
probably due to the relative insensitivity of the analytical procedure (too many zero
range values).

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 CONTROL CHARTS IN THE LABORATORY 1065

The control chart for albumin showed scattered values which were out of control
but there was no general trend that would indicate a constant error creeping
into the determination. During the time the range values for the albumins were
out of control, the total protein determination was fluctuating beyond the
defined limits.

Carbon Dioxide Combining-Power of Plasma

This property of the test plasma was determined by the method of Van Slyke
and Cullin,6 employing the volumetric Van Slyke gasometric apparatus. The
mean value for the test samples used was 81 volumes per 100 ml. plasma while
the control limits ranged from 77.4 to 84.6 volumes per 100 ml. (Fig. 4). All
the mean values were in control, although one of the range values appeared to
be out of control. To be in control, the difference in duplicates could be as great
as 11.7 volumes per 100 ml. plasma.

DISCUSSION

The control chart is an important aid in keeping the variation of any single
chemical analysis of a clinical laboratory within set statistical limits. Some of
the advantages of the control chart are as follows: The control chart offers a
simple method of checking the resultant effect of all factors influencing the
accuracy of a given test, e.g., the reagents, standards, time factors and instru-
ments used in the analysis. I t offers a basis for action in initiating correction
of a method that is not functioning properly. Also, it improves the general
accuracy of the laboratory, since the technicians become control-conscious and
readily detect and report a test that is out of control.
By using the control chart it is possible to tell at once whether or not a method
is working well. If the method is out of control, the chart usually cannot give
the reason and it is up to the analyst to determine the cause of the difficulty.
Sometimes it is possible to note the deterioration of reagents or standards by
observing a trend in the control curve. However, the possibility of deterioration
of the substance in the frozen control standards should also be considered. In
setting up control charts for the gamma globulin method of Kunkel, 4 it was
found that the gamma globulin content of the normal plasma used as a standard
had completely disappeared on storage in the frozen state after three months.
The data presented in this work were obtained by seven technicians and three
student technicians; and yet with the exception of some of the chloride deter-
minations, there were no constant analytical differences in the mean values which
could be attributed to any one worker. But even with respect to chloride analysis,
when the new worker determined a constant for the reagent, the analyses were
again in control. I t is possible by the use of the control chart to discover an
inept analyst.
The use of control charts need not be limited to the determinations described
here. Any of the chemical analyses routinely done in the laboratory, such as
glucose, nonprotein nitrogen or hemoglobin, could be used for control studies.

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 1066 LEVEY AND JENNINGS

SUMMARY

The application of the principle of the control chart is made to the chemical
division of the clinical laboratory. Examples of the use of these charts for the
evaluation of the functioning of the determination of the urea content of the
blood, and the chloride, protein and carbon dioxide combining-power of plasma
are given. By using the control chart, it is possible to ascertain at a glance the
over-all functioning of any determination thus studied. It also provides a basis
of action in correcting a defective method.
Acknowledgments. T h e authors are indebted to D r . Benjamin Epstein of the D e p a r t m e n t
of Mathematics of Wayne University for aid in this study, and to Mi$s Jean Reed and Mrs.
Violet Greenberg for technical assistance. j

REFERENCES
1. American Society for Testing Materials: Manual on the Presentation of D a t a . Philadel-
p h i a : The Society, 1947, 47 p p .
2. K A R R , W. G.: A method for determination of blood urea nitrogen. J. L a b . and Clin.
Med, 9: 329-333, 1924.
3. K I N G S L E Y , G. R.: T h e direct biuret method for the determination of serum proteins
as applied to photoelectric and visual colorimetry. J. L a b . and Clin. Med., 27: 840-
845, 1942.
4. K U N K E L , H . G.: Estimation of alterations of serum gamma globulin by a turbidimetric
technique. Proc. Soc. Exper. Biol, and Med., 66: 217-224, 1947.
5. P E T E R S , J. P., AND VAN SLYKE, D . D . : Quantitative Clinical Chemistry. Vol. 2. Balti-
more: Williams & Wilkins Co., 1932, 957 p p .
6. SCHALES, 0 . , AND SCHALES, S. S.: A simple and accurate method for the determination
of chlorides in biological fluids. J. Biol. Chem., 140: 879-884, 1941.
7. WERNIMONT, G.: Use of control charts in the analytical laboratory. Ind. E n g . Chem.
Anal. Ed., 18: 587-592, 1946.

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