QUANTITATIVE EXPERIMENTS WITH THE CUTANEOUS

TUBERCULIN REACTION

CLEMENS F. v. PIRQUET

(From the Departme,U of Pediatrics of the Johns Hopkins University)

(Received for publication, April 14, 1909)

The old tuberculin of Koch, an extract of the tubercle bacilli, causes

a local inflammation when it is used on the skin of a tuberculous mdi-
vidual.1 The usual signs of inflammation become manifest, namely,
rubor, tumor, dolor, calor, and microscopically the specific lesions of a
tuberculous inflammatory process are encountered.
From my first experiments I reached the conclusion that the inten-
sity of the reaction stands in a certain relation to the quantity of tuber-
culin used. Using progressive dilutions of tuberculin it was always
seen that the undiluted tuberculin elicited the most pronounced inflam-
mation whereas dilutions of 1 10 or 1 100 called forth smaller pap-
ules which were not as much raised. Dilutions of 1 1000 showed
only a positive reaction while the undiluted tuberculin gave rise
to a very intense inflammation. When this reaction was weak even
dilutions of 1 : 10 were indefinite or negative.
This quantitative relation of the reaction to the amount of the for-
eign body introduced in the skin established a certain connection of the
tuberculous cutaneous reaction with the “early reaction” after re-
peated Jenner vaccinations.
For the Jenner vaccination I proved2 that the effect of a first vaccina-
tion is not dependent on any dilution of the lymph, but that the
“early reaction,” which is only met with in repeated vaccinations,
shows itself quantitatively dependent on the amount of lymph used.

1 Berl. kiln. Worchenschr., 1907, 20; Wien. med. Wochenschr., 1907, 28.
3Wien. kiln. Worchenschr., 1906. Nr. 47.
152 CLEMENS F. v. PIRQUET

Fig. 1 is based on the measurements of the diameter of the local

reactions after the first Jenner vaccination. To obtain comparable
measurements it was found convenient to make small round abrasions
of the skin about 1 mm. in diameter. To accomplish this I used a

1234’S 74’, ft 12 12 74’

Tag mr4
Durchmesser der Hyperamle.
Papille.

FIG. 1. CURVES OF A FIRST VACCINATION WITH COWPOX.3

special instrument shaped like a chisel. The edge of the chisel is

pressed against the skin and with a rotary movement abrasions of suf-
ficient uniformity are produced. In fig. 1 we find registered the
body temperature and two curves. The lower curve expresses the
size of the central papule and is linear in character gaining about 1 mm.
‘Vaccination und vaccinale Allergie, Wien, Deutike, 1907.
 THE CUTANEOUS TUBERCULIN REACTION 153

daily. The same type of curve expresses the growth of bacterial cul-
tures on solid media.3 The upper curve represents the, development
of the areola. The areola appears between the eighth and tenth day
together with the general symptoms, gains rapidly in size, reaching
its maximum after two or three days. At the same time the growth

123 5675$ ff

Tag eauL/nei*v daersia /m#{216}/e//e

Durchmesser der ROtung:

c-o Erstfmpfung (Iinpfstelle 1).
Nacbimpfung am 3. Tage (Impfstelie 3). I
,, 5. ,. ., 5).
9. ,, ,, 12).
14. ,. ,, 21.)

FIG. 2. CONSECUTIVE VACCINATIONS. DEVELOPMENT OF “ALLERGY”3

of the central vesicle is completed and its contents lose their specific
activity. Dilution of the material used for the vaccination does not
change the character of the curves at all, but the whole train of events
is postponed.

Vaccination und vaccinale A5llergie, Wien, Deutike, 1907.

154 CLEMENS F. v. PIRQUET

In fig. 2 we find the curves of the development of the areola

only. The same individual was vaccinated for the first time and from
that day on the vaccinations were repeated daily with the same amount
of lymph. The areola develops on the same day in vaccinations
which precede the point on which the character of the curve changes.
Vaccinations made after this time develop their areola inside of twenty-
four hours. This areola is much smaller, a vesicle is not formed and
the whole phenomenon disappears rapidly, wherefore reactions of this
type which I called “ Frilhreaction “ (early reaction) were overlooked.
This type of altered reactivity of the organism (“ Allergy “) may last
throughout life, gradually within a course of years reverting again
more and more to the type observed in the unimmunized individuals.
The height of the curve (the diameter of the inflamed area) of this
Friihreaction is dependent on the quantity of lymph used. With
progressive dilutions the curve elevates itself less and less from the
abscissa. The method of procedure does not permit the fixation of
an exact zero point on account of the slight traumatic reaction caused
by the use of the instrument alone. The minimal reactions after
vaccinations grade off into this traumatic reaction.
This quantitative relationship agrees very well with the theoretical
conceptions which I had previously formed about the Frtihreaction.4
After the first vaccination the pathogenic microorganisms multiply in
the skin without interference until the organism produces antibodies
which have a digestive action on these particular microorganisms.
During this digestion toxic substances are formed and these exercise
an irritating influence on the cells falling within their reach and lead
to the inflammatory symptoms.
The digesting antibodies are not produced immediately after the
first vaccination but only after an interval of eight to ten days. Accord-
ingly the areola with its rapidly rising curve appears at this time.
Microorganisms introduced after the formation of the antibodies
are immediately attacked. The effect of the toxic substances due to
their digestion becomes immediately noticeable as Friihreaction.
In the case of tuberculin vaccinations the conditions are altered

‘Verhandlungen der Deutschen Gesellschaft f#{252}r

Kinderheilkunde, Kassel, 1903.
Wien. kim. Wochenschr.. 1903, Nr. 26, 1906, Nr. 28, 47.
 THE CUTANEOUS TUBERCULIN ItEACTION 155

inasmuch as instead of living microorganisms extracts are used. Never-

theless there exists this similarity with the processes just discussed
that only such individuals give rise to a Friihreaction as have been
previously infected with tuberculosis. And in these we may assume
the presence of antibodies. The mode of action of the antibody is

dsr

FIG. 3. QUANTITATIVE EXPERIMENTS WITH TUBERCULIN.5

not cleared up as yet, neither is it of interest to us at present whether

we have to deal with a single substance or whether several components
enter into the reaction such as thermostable amboceptor, thermo-
labile complement, etc. The object of this investigation was to deter-
5Ergebnisse der inneren Med. u. Kinderheilkunde, 1908. I.
156 CLEMENS F. V. PIRQUET

mine in how far the intensity of the clinical phenomenon of inflamma-

tion is dependent on the concentration of the tuberculin used and the
role which the different content of different parts of the skin in anti-
bodies plays in this regard.
It was my intention to establish the laws governing the intensity
of the Fruhreaction. At first a large number of experiments were
carried on on different individuals. Each of these was vaccinated
with a series of progressively diluted tuberculin. The limits of error
widened to such an extent because of external circumstances beyond
our control, that I preferred to make the experiments on myself. With
each of the dilutions several vaccinations were made in order to gain
a good average and the measurements were made very frequently.
In this way the inexactitudes of the method were reduced to the small-
est possible extent.
Such an experiment was made in November, l907, and is illustrated
in fig. 3. At that time I made five vaccinations of each dilution.
The vaccinations, thirty in all, were made on the extensor surface
of my upper thighs. The technique and particularly the demonstra-
tion of the results have been somewhat improved since then and the
following mode of procedure was adopted.

EXPERIMENTAL PART.

Tuberculin. Alt Tuberculin Hoechst (from Koechl in New York,

dated March 8, 1908) was used for vaccinations. The dilutions were
prepared in nine watch glasses. At first 0.5 cc. of water was placed in
each watch glass then 0.5 cc. tuberculin in the first one. This was
thoroughly mixed. From this dilution (1: 2) 0.5 cc. was added to the
next watch glass, giving a dilution of 1 :4. In the same manner
further dilutions of 1 : 16, 32, 64, 128, 256, 512 were prepared.
Field of vaccination. The extensor and median surfaces of the upper
left thigh had been shaved the day before. It is preferable to shave
the day before because superficial abrasions can not be avoided and
these would allow an absorption of the tuberculin in other places than
the vaccination spots.
Close to the knee a control was prepared by placing a drop of water
with a glass rod on the skin, then followed nine rows each of four small
6 Ergebnisse der inneren Med. u. Kinderheilkunde, 1908. I.
 THE CUTANEOUS TUBERCULIN REACTION 157

drops of diluted tuberculin and finally one row with undiluted tuber-
culin. The distance at which the drops were placed was about 40
mm. with undiluted tuberculin, gradually decreasing to 25 mm. in the
lower dilutions.
After the droplets had been placed the instrument described above
was turned through each drop with as equal a pressure as possible,
starting from the control and the lowest tuberculin dilutions. With
sufficient practice the method gives very uniform round abrasions of
the skin of about 1 mm. diameter.
Observations. The experiment was started on the forenoon of the
twenty-fourth of March. The vaccinations were accomplished in
the time between 10:04 a. m. and 10 : 06 a.m.
10 : 09 a.m. Everywhere on the points of vaccination colorless
papules of 3 to 4 mm. diameter are seen. I mean slight elevations of
the skin, which do not differ in their color from the neighboring skin.
The total field, the surface of the upper thigh, is a little hyperemic,
very probably in consequence of the irritation due to the somewhat
painful manipulation.
10 : 12 a.m. All vaccination points were dried with cotton; only
where the higher concentrations had been used were the places still
moist on account of the glycerine content. Where the low concen-
trations were used the exposure to the air had dried the places.
10 : 18 a.m. The general hyperemia of the thigh has disappeared
and only in the neighborhood of some of the vaccinations a slight and
diffuse hyperemia is noted.
10 : 22 a m. All the vaccinations were touched first with wet then
with dry cotton in order to remove all of the adhering tuberculin.
10 : 30 a.m. On all vaccinations very slight papules are noted of an
“urticaria-like” character; on stretching the skin or on pressure they
fade, that is, they become paler than the neighboring skin. The
diameter of the papules varies from 3 to 5 mm., some of them are still
colorless, others slightly hyperemic with areolae not sharply defined
sending out processes to the diameter of about 8 mm.
10 :45 a.m. (40 minutes post vacciruitionem = 40 m. p. v.) All
places were drawn. They are nearly uniform papules of about 5 mm.
diameter “urticaria-like,” slightly hyperemic, mostly of round from,
some sending out delicate processes.
158 CLEMENS F. v. PIRQUET

11 : 15 a.m. (1 h. 10 m. p. v.) All lesions measured. Diameter

3 to 5 mm.
11 : 35 a.m. (1 h. 30 m. p.v. (Measuring. Diameter between 3 and
5 mm., but on an average somewhat smaller than before. The pap-
ules fade under pressure only very little urticaria-like, they are very
little elevated and very slightly hyperemic.
12 : 05 p.m. (2 h. p.v.) Very slightly hyperemic, small spots,
hardly elevated, not palpable, do not fade any more urticaria-like.
1 : 25 p.m. (3 h. 20 m. p.v.) All lesions sketched. The circumference
of the lesion produced by the instrument hardly elevated, not distinctly
hyperemic, having a diameter of about 2 to 3 mm. On one vaccina-
tion the specific reaction be#{231},rins
: it is the second place of the first row,
with undiluted tuberculin (“ 1 b “) . It is little elevated, somewhat
more hyperemic than the others and has a diameter of about 8 mm.
“8 a” also is a little larger than the rest, with a diameter of about 4mm.
1 : 55 p.m. (3 h. 50 m. p.v.) Drawing. “1 b” measures 9 mm.
it is indistinctly palpable. Specific reaction just beginning in 1 a,
2 c,4a.
2 :05 p.m. (4 h. p.v.) Drawing. 1 b measures 11 mm., indis-
tinctly palpable. A number of other places (1 c, 2 a, b, d, 16 a, 32 a,
show beginning reaction. The hyperemic areas are not yet raised and
measure 4 to 8 mm. The rest of the vaccination shows indefinite red-
ness of 2 to 3 mm. like the control.
2 :35 p.m. (ft h. p.v.) Drawing. 1 b is now definitely palpable.
Diameter 12 mm.
3 : 05 p.m. (5 h. p.v.) Drawing: New reaction in 8 c.
3 : 35 p.m. (5 h. 30 m. p.v.) Drawing. New reactions in 4 b,
16 b, 32 d.
4 :05 p.m. (6 h. p.v.) and 4 :35 p.m. (6 h. 30 m. p.v.) Drawing.
A number of vaccinations are now definitely palpable (1 b, 2 a, 4 a,
8 a), new reactions: 4 c, 8 c, 16 d, 128 b. In spite of the fact that some
of the very diluted vaccinations react already, 1 d, 4 d, and 8 d do not
show any reaction as yet.
In glancing over the field one might gain the impression as if the
higher concentrations were used on the lateral part of the thigh and the
lower concentrations on the medial part. De facto the higher con-
centrations stand proximally, the lower ones distally. This peculiar
 THE CUTANEOUS TUBERCULIN REACFION 159

behavior finds its explanation in the fact that the lateral part of the
thigh had been used for the similar experiment already referred to,
about 1 years ago, whereas the medial part of the thigh is new ter-
ritory. I will return to the discussion of this phenomenon later on.
5 : 10 p.m. (7 h. 5 m. p.v.) Drawing. New reactions at 64 a, b, c.
Messuring.
5 :45 p.m. (7 h. 35 m. p.v.) Drawing. Now only the develop-
ment begins in 1 d, 4 d, 8 d and 16 b. All places of rows + to
are reacting.
6 : 10 p.m. (8 h. 05 m. p.v.) Measuring.

1 2 48 16 32 8 256 512 C

- -
a 13 14 13 12 15 9 6 5 3 4 3
k ,-. ,- -‘ ,-.- ,
17 12 7 7 5 6 7 6 4 3
C ,.-, .-# ,-
16 6 5 5 6 5 5 4 3
d 10
3 4 5 5 5 6 5 4 4 3
Note: The sign ‘‘ means distinctly palpable, -‘ indistinctly palpable. Those not marked are flat.

The appearance of hyperemia in 32 c, 64 d, 128 c and d, closes the

rows up to 128; 256 shows a reddening in 3 places, 512 in one place.
In the outstanding place of 256 the reddening begins at 8:35 (10k
h. p.v.), in 512, b is reddened at 8:45 p.v., c and d only at 14 h. 40 m.
and 17 h. 40 m. p.v.
6 : 50 p.m. (8 h. 45 m. p.v.) Drawing. Measuring. On the
lateral part of the field all the vaccinations up to 128 are distinctly
palpable. There exists no considerable difference in the size of the
undiluted tuberculin place up to 1 : 16, the diameter being about 15
mm.; 32, 64 and 128 are considerably smaller.
8 : 35 p.m. (10k h. p.v.) Measuring. 1 d is still quite insignificant
and not larger than 256 a.
10 : 15 p.m. (12 h. 10 m. p.v.) Drawing and measuring. The
form of most of the vaccinations is nearly perfectly round, only 1 b and
1 c have long processes making a lymphangietic connection between
the two.
11 :35 p.m. (13k h. p.v.) Measuring.
March 25. 12 : 45 a.m. (14 : 40 h. p.v.) Measuring.
3 45 a.m. (17 : 40 h. p.v.) Measuring.
160 CLEMENS F. v. PIRQUET

1 2 4 8 16 32 64 128 256 512 C

. % - -
a.... 15 13 15 13 18 14 11 7 5 4 3
b.... Th Th Th 9 #{237}#{244}
I I 4
,- .-, .-% %

C.... 17 18 13 10 8 8 8 6 5 4
. . - - -
d.... 9 13 13 7 7 8 8 7 3 5

5 : 45 a.m. (19 h. 45 m. p.v.) Measuring.

10 : 20 a.m. (24 h. 15 m. p.v.) Measuring.
12 m. (25 h. 55 m. p.v.) Measuring and drawing.
Now all the delayed vaccinations are nearly fully developed, the rows
presenting a fairly uniform aspect and the dependence of the reaction
on the dilution of the tuberculin is readily recognizable. Only the first
3 rows do not show this dependence : the 12 points with the undiluted,
half and four times diluted tuberculin do not differ much.
All vaccinations including 1 : 64 are distinctly palpable. In 128
three places are distinctly palpable, the places in 1 : 256 are indistinctly
palpable, those in 512 not palpable, although they differ distinctly from
the control by the diameter of the hyperemia. At. the same time an
involution becomes noticeable, since these places had been somewhat
larger before and a little more elevated.
2 : 10 p.m. (28 h. 05 m. p.v.) Measuring.
5 : 35 p.m. (31 h. 30 m. p.v.) Measuring. Nearly all vaccinations
present the phenomenon of the Halo-formation. The vaccination
papule proper is much elevated and has a slight yellowish pigmenta-
tion aside from its more intense redness. Now it is surrounded by a
slightly red halo. The papule itself is smaller than in the previous
measuremejlts, while the halo has a diameter up to 24 mm. The vac-
cinations with diluted tuberculin have no real papules any more, but
the surroundings of the lesions produced by the instrument are sim-
ilarly reddened like the halos of the more concentrated vaccinations.
It is surprising that the control too participates in this hyperemia.
At 6 :30 p.m. (32 h. 25 m. p.v.) by daylight and at 6 :35 (32 h. 30
m. p.v.) by electric light measurements were taken to see whether the
difference in the illuminations gives a pronounced error. This was
not the case. The halo-formation is now very distinct, only 1 d does
not yet show it.
 THE CUTANEOUS TUBERCULIN REACTION 161

March 26. 12 :05 a.m. (38 p.v.) Measuring. The halos are
sti]llarger than the last time, and 1 d has a halo too.

7 : 30 a.m. (45 h. 35 m. p.v.) Measuring. The halo can only be

‘“I

FIG. 4. FIELD OF VACCINATION UNDER TANGENTIAL ILLUMINATION.

seen indistinctly in few places. The papules are sharply defined,

yellowish red. The exudation has decreased considerably. Most of
the vaccinations from 1: 16 down are indistinctly palpable.
162 CLEMENS F. v. PIRQUET

3 :05 p.m. (53 h. p.v.) Measuring. Some places stillshow halo,

presenting mostly a very large (up to 35 mm.) indefinite reddening.
March 27. 9 : 30 a.m. (3 days p.v.) Drawing, measuring, photo-
graph taken.

The halos have disappeared. The papules +, , , are all dis-

tinetly palpable, much pigmented. to B1 are indistinctly pal-
pable with few exceptions. Tie rest can hardly be recognized as re-
actions ; th are not any more to be distinguished from the control.
This photograph (fig. 4) is taken under tangential illumination so
that the elevations can be seen. Those at the proximal rows are
very pronounced, and it is evident that even the eighth and ninth
rows are somewhat elevated.
Under
vertical illumination (fig. 5) only the difference in the color-
tion can be seen. The photograph illustrates the general arrange-
ment of the vaccinations, while the details can be seen better in the
drawing on fig. 6.
March 28. (4 days p.v.) Measuring. The pigmentation is about
equally pronounced, the papules are still less palpable.
March 29. (5 days p.v.). Measuring. Only few places are still dis-
tinctly palpable (2 d, 4 c, d). In some places the small crusts of the
center are fallen off.
March 30. (6 days p.v.). Only 4 c is indistinctly palpable, others not
at all. On the lateral part the vaccinations can only be recognized
by slight pigmentations, on the medial part at the thigh they are a
little bit more distinct, yellowish red.
March 31.(7 days p.v.). Measuring. All crusts fallen off, all vaccin-
ations flat. The places ,on the, lateral part and from. 1 : 64 down
hardly recognizable any more.
April 7. (14 days p.v.). The center of the vaccination can be recog
 THE CUTANEOUS TUBERCULIN REACTION 163

nized by a circle of peeling skin of a diameter of 2-3 mm. There is no

difference with regard to the dilutions. In sonic places on the medial

FIG. 5. FIELD OF VACCINATION UNDEII VERTICAL ILLUMINATION.

part a very slight grayish-red discoloration is all that remains of the

pigmentation. Besides, a very fine transversal corrugation of the
epidermis is noted.
164 CLEMENS F. v. PIRQUET

4:

!i!

_____ d .

FIG. 6. FIELD OF VACCINATION (DaAwIN).

( of natural size.)
 THE CUTANEOUS TUBERCULIN REACTION 165

CONSTRUCTION OF CURVES DEMONSTRATING THE COURSE OF REACTION

OF EACH VACCINATION POINT

To gain an exact picture of the course of the reaction a system of

ordinates was constructed. The abscissa denotes the time, the ordi-
nates the extension that is the diameter of the inflammatory reaction in
mm. In correspondence with the single measurements 35 ordinates
were erected on the abscissa.
Diameter
18
mm

10
mm

48 hours
FIG. 7. COURSE OF REACTION OF POINT +A (UNDILUTED TUBERCULIN, LATERAL SIDE).

The system of ordinates was constructed once and served for all the
reactions. A piece of tracing-paper was placed on top of this. So the
results obtained from each of the places of vaccination were plotted
on a separate sheet of tracing paper. At the same time different signs
were used to denote the degree of elevation of the reddened area.
#{149}
= distinctly palpable, o = indistinctly o = not palpable.
After entering all the points it was found that a simple curve does
not include all the points, but frequently differences of 1 to 2 mm. and
even more were noted in consecutive measurements. These differences
are doubtless dependent on errors in measuring. These errors are mostly
 166 CLEMENS F. V. PIRQUET

due to the fact that the redness does not end abruptly and that the cir-
cumference is not a perfect circle. Therefore the numerous measure-
ments are placed between two arbitrary lines, one of which passes
through the highest values, and the other through the lowest.
Then a curve was drawn falling between these two auxiliary lines
and touching as many points of measurement as possible. It would
have been preferable to give all the forty separate curves, which would
represent the record of all the measurements taken, but it must suffice
to discuss a few of these curves as specimens.

18
mm
Diameter

10
mm

0 6 12 24 hours 48

FIG. 8. COURSE OF REACTION OF POINT +D (UNDILUTED TUBERCULIN, MEDIAL SIDE)

We see in fig. 1 the beginning of the traumatic reaction at 5 mm.,

then the curve drops for several hours. About four hours after the
vaccination the specific reactioi begins and with it a rapid rise of the
curve.
12 h. p.v. the maximum is nearly reached and the curve declines
gradually after 24 h. p.v. ‘

The place d, located on the medial side of the thigh, shows a con-
siderably different curve (fig. 8). Here a period of coinplete latency
lasts for about 7 hours, then the specific rise occurs, but it does not
 7

THE CUTANEOUS TUBERCULIN REACTION 167

begin with an angle of 85 as in + a, but with one of about 50#{176}

and
the curve reaches its highest point only after 24 hours. Later the
curve does not decline, but the papule remains visible to the same ex-
tent during the time of observation. Here the auxiliary lines could
be dispensed with, since the points do not oscillate much.
The curves of the tuberculin dilutions recall these two main types.
The earlier the reaction sets in the more rapid is the rise of the curve.
As a rule,the reactions set in later, the curves rise less rapidly and do
not reach the same height as with the undiluted tuberculin.
The following are two typical curves of tuberculin dilutions:
Diameter
13
mm

10
mm

0 6 12 24. hours . 48

FIG. 9. COURSE OF REATION OF TUBERCULIN 32 TIMES DILUTED.

1: 32 b. (fig. ‘9.) Period of latency, 5 hours; the curve rises at an

angle of 70#{176};
maximum of 11 mm. between 12 and 24 hours; gradual
decline.
1 : 256 a (fig. 10) shows one of the smallest elevations, still permit-
ting a sharply defined observation of the specific course of the reaction.
Period of latency, 7 hours. Rise of the curve at an angle of about
60#{176},
maximum only 5 mm, then gradual decline. The papule becomes
distinctly palpable for a very short time.
The halo formation was not considered in constructing the curves;
this phenomenon will be dealt with in a later publication.
 168 CLEMENS F. V. PIRQUET

CONSTRUCTION OF AVERAGE CURVES FOR EACH CONCENTRATION OF

TUBERCULIN.

The forty curves obtained in the manner outlined above were now
utilized for the construction of average curves for each concentration.
The main curves of each of the four vaccination places forming a row
of a given concentration were drawn on one paper and the arithmetic
mean of two of these curves was determined. In this way two aver-
age curves were obtained. Halving again the distance between a series
of points on these two curves the average curve of all four original
curves was obtained. This average curve begins with the time of the
rise of the latest original curve.

Diameter
8
mm

5
mm
,(
z
o

-1 0

0 6 12 24 hours 48
FIG. 10. CORSE OF REACTION OF TUBERCULIN 256 TIMES DILUTED.

Of the ten average curves I reproduce that of the dilution 1 :4 as an

example (fig. 11.) Here papule a develops first, on the lateral part of
the row, its curve shows a rapid and high rise,one hour later papule b
develops. The curves of both papules show a distinct decline after
reaching the maximum, as in the example given in fig. 7 representing + a.
The reaction of c and d sets in later and their curves resemble that of
+ d, they rise slowly but do not decline later on, denoting a more chronic
process.

COMBINATION OF THE AVERAGE CURVES TO A GENERAL PICTURE.

All ten average curves were drawn in one table. The picture ob-
tamed in this way became indistinct since the numerous lines crossed
each other several times. Therefore I prefer to arrange the material
in two tables each with five curves.
 r

THE CUTANEOUS TUBERCULIN REACTION 169

The firsttable (fig. 12) contains the curves of the undiluted tuberculin
and the dilutions in the geometric progression of four (, ,.1, and
the second table (fig. 13) reproduces the intermediate curves , ,
J 1’l
5T2
The curves show certain differences. Some show a rapid rise, others
a slower one. But this behavior cannot be attributed to the dilution
as such. As we have seen above in the curves of 1 a and 1 d other
factors enter here into consideration. The rapid rise depended here
essentially on the location of the vaccination places on the thigh.
Diameter
20
mm

10
mm

0 6 12 24 hours 48
FIG. 11. AVERAGE CURVE OF DILUTION 1:4 + AVERAGE POINTS OF TWO SINGLE CURVES

#{149}
AVERAGE POINTS OF ALL FOUR CURVES.

On the other hand it isevident that the absolute height of the curves,
that is, the size of the diameter of the papule, is dependent on the
dilution. -f, and apparently do not present any essential difference
in this regard, but all the others arrange themselves according to the
dilution. The close approach of and th may be explained by the
difficulties connected with the exact measurement of such weak reac-
tions.
 170 CLEMENS F. V. PIRQUET

mm Diameter
16

14

12

10

12 24 36 hours 48
FIG. 12. AVERAGE CURVES 1, 4, 16, 64, 256.

mm Diameter
16

16

2
12
/

I0 8
-

1/
6

4 L ----. 128
,

2
/

0 6 12 24 36 hours 48

FIG. 13. AVERAGE CURVES 2, 8, 32, 128, 512.

 THE CUTANEOUS TUBERCULIN REACTION 171

This brings us to the main object of our investigation. Is it pos-

sible to establish definite quantitative relations between the dilution
of tuberculin and the degree of its action?
As the result of former investigations I reached the conclusion that
the diameter of the papules was inversely proportional to the cubic
root of the dilution.7 This formula, however, seems not to cover
the process.
Which part of the curves lends itself best to gain some information
with regard to our main question? The rising part of our curves can
hardly be utifized, since other factors exercise an influence. To
exclude these influences it will be necessary to conduct new experiments
on a region of skin which has not served for similar experiments.
It is possible to use as a standard of comparison the highest eleva-
tion of the curves without reference to time, or the diameter of the
reactions at a given time.
In the following table the maximum as well as the figures for the
diameters after 24 and 48 hours are given; that is, the figures were de-
duced from the average curves.

1 2 4 8 16 3264 128 256512

After24hours 15.4 15.2 14.8 11.4 10.4 9.3 7.7 6.6 3.8 3.8
After48hours 13.4 12.8 12.5 9.8 7.7 7.2 5.5 3.8 2.3 2.2
Maximum 15.4 15.4 14.9 11.4 10.5 9.9 8.0 7.2 5.5 4.3

In fig. 14 I tried to construct a continuous curve from the figures given

for 24 hours p.v. The upper line designates the curve prepared on
the basis of the data given in this paper. The curve passes nearly
through all points with the exception of *and ; it is evident that one
too high, the other too low. The same mistake is to be seen in 256
and 512 which fall on the same point. 256 should have been at
about 5 mm., 512 below 4.
The lower curve represents the results of the experiment of Novem-
ber 1907. The elevation of the curve is lower throughout, but shows
about a similar course, with the difference that the end does not decline
‘Medizinische Klinik, 1907, Nr. 40.
 172 CLEMENS F. v. PIRQUET

as rapidly. This difference can be accounted for by a difference in the

calculations adopted at that time.
The experiment itself, arranged like our experiments above, using
the time as abscissa was given in fig. 3.

SIGNIFICANCE OF THE CURVE ON FIG. 14

I had supposed that the type of the curve in itself might furnish
an idea with regard to the physical chemical mechanism of the reaction.
After consultation with Prof. H. Jones, who kindly interested him-

mm Diameter

16

14

12

10

FIG. 14. DILUTION AND DIAMETER.

self in my problem, one of the curves of fig.14 was given to the mathe-
maticians of the University for their inspection. The report of Pro-
fessor Morley and of Mr. Sparrow, who were kind enough to examine it,
is as follows: “The curve is approximately a hyperbola with an
asymptote parallel to the axis of abscissa. Without formulating a
definite theory as to the manner of propagation of the inflamma-
tion it is impossible to say whether the curve satisfies the require-
ments of the mass action law or not.” It is my intention to make
fuller experiments, in order to exclude the possible influence of concen-
tration of salts and glycerine on the initial part of the curve.
 THE CUTANEOUS TUBERCULIN REACTION 173

CONCLUSIONS

The cutaneous tuberculin reaction depends on at least two factors.

One of these factors is the tuberculin, the other, furnished by the
organism, can be considered as an antibody, whose origin dates back
to a previous infection of the organism with tubercle bacilli.
In the experiments just described the first factor has been subjected
to definite variations in using tuberculin of different concentrations.
Diameter
20
mm

10
mm

6 12 24 hours 48

FIG. 15. INFLUENCE OF BOTH FACTORS ON THE COURSE OF REACTION.

1 a large amount of both tuberculin and antibody.

1 d large
amount of tuberculin and little antibody.
128 amount
a large of antibody and little tuberculin.
128 d small amounI of both.

The effect of the progressive dilutions showed itself in the diminution

of the intensity of the reaction. A similar occurrence had been noted
before inthe “FrUhreaction” in cowpox vaccination. A definite thathe-
matical expression for the effect could not be elicited thus far; further
investigations avoiding certain sources of interference will be under-
174 CLEMENS F. V. PIRQUET

taken, and I hope that it will then be possible to determine that these
phenomena can be classed under the mass action law. At present, it
would appear as Professor H. Jones remarked to me, that the active
mass of the second factor is very much smaller than the active mass
of the tuberculin used in the reaction.
The presence of the second factor can only be surmised from the
following considerations: An individual, free from any tubercular
infection, does not give any cutaneous reactions with tuberculin.
In persons who give the reaction, a marked difference exists between
different areas of the skin. In skin areas which have been subjected
previously to one or repeated applications of tuberculin, the reaction
proceeds in a different manner.
Here again a striking analogy with the results of repeated smallpox
vaccinations becomes manifest,8 and it may be concluded that besides
an accumulation of antibody in the general organism a local accumu-
lation occurs.
The inflammatory reaction is supposed to rest on a union of both the
factors given, with the production of a toxic principle. This toxic
principle acts on the tissues producing the inflammatory papules and
reddened areas.
Fig. 15 demonstrates the influence of the concentrations of tuberculin
and supposed antibody on the course of the reaction.
The most prominent influence of the concentration of the antibody seems
to lie in the time in which the reaction appears and reaches its mixi mum,
whereas the absolute height of the curve, signifying the intensity of the
reaction seems to be dependent on the concentration of the tuberculin.
It is of interest to note that in every instance a period of latency was
recorded, even when the high concentration of both factors induced
a relatively rapid appearance of the reaction.

‘Vaccination und Vaccinale Allergie, Wien, Deurike, 1907.