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Experience with the six hour rat test for pregnancy
SANFORD KAMINESTER, M.D., F.A.C.S., BROOKLYN, (From the Long Islana College of Medicine)
N. Y.
NNUMERABLE pregnancy tests have been described in medical literature. Most of them have one or more disadvantages which have kept them from being generally accepted. It was not until Aschheim and Zondek described their technique that there became available a pregnancy test which met most of the requirements for such a procedure. A pregnancy test, in order to be of much value, must meet certain specifications : ‘its degree of accuracy must approach 100 per cent; it must be reasonably rapid and relatively simple. To meet these requirements, much experimental work has been done. It would seem that the ideal test should be a chemical one, thus removing the source of error inherent in all work which depends upon biological assay. Last year, Salmon and Geistl and their associates at the Mount Sinai Hospital used, as a pregnancy test, an observation which had been made many times before. They reported that within six hours after the injection of the urine of pregnant women into suitable rats a definite erythema of the ovaries developed. This was constant and their results compared favorably with those obtained with the Aschheim-Zondek or Friedman tests. In our laboratory, in 107 cases, we have had the same experience. Technique
I
Female rats are used as the test animals. They must be about four weeks old and weigh about 30 grams. Since these animals have a spontaneous estrus cycle and ovulate spontaneously at maturity, they cannot be used when they become much larger. Two animals are employed and two cubic centimeters of urine are injected subcutaneously into eaah. After six hours, the animals are asphyxiated with illuminating gas. In positive cases, the ovarian hyperemia is manifested by a reddening of the ovary. This redness is grossly visible in good light. Usually the uterus, too, is edematous and red looking although the interpretation of the test depends on the ovarian and not the uterine conditions. In the negative cases, the ovaries are small and white and the uterus is a whitme, threadlike structure. In 107 cases, the following results were obtained : *Presented
at a meeting
of the Brooklyn
Gynecological
265
Society,
October
1, 1043.
266
AMERICAN
JOURNAL
RESULTS TOTAL 39
OF
OBSTETRICS
AND
IN PREGNANT
WOMEN
RAT TEST POSITIVE
1st trimester 37 2nd trimester 2 2 3rd trimester 13 13 Duration unknown 9 9 Total 62 61 (eliminating dead fetus ease)
NEQATIVE
2 (a,b)
1
GYNECOLOGY
FRIEDMAN TEST TOTAL POSITIVE NEQATIVE 18 20 2 (be)
19
18
1
a-Test in rat negative but Friedman test positive. Clinically the patient was shown to be pregnant. b-Both rat and Friedman test were negative. In this case the fetus had been dead for about 3 months. c--Friedman test negative but rat test positive. This patient was shown to be pregnant clinically. RESULTS
IN NONPREGNANT TOTAL 15
RAT TEST POSITIVE
WOMEN NEQATIVE
AND IN MALES TOTAL
FRIEDMAN TEST POSITIVE NEQATIVE
Malea Yl 15 0 0 Nonpregnant Women 29 (a) 28 9 0 Totals 44 1 43 9 0 d-As this was one of the early cases, the error may have been due to inexperience in interpreting the findings.
0 9 9
At the beginning of the study, samples of urine were tested from male students, known pregnancies and known nonpregnancy cases. The identity of the subject was not known to the individual reading the result. Later, rat tests were done on the urines submitted for routine Friedman tests and the readings were made before the Friedman tests were completed. In those cases where there was no Friedman test and the samples were received from an outside source, follow-up was requested on the basis of the clinical course. In this way it is felt that all possible sources of error have been covered. After this careful investigation it was found that 61 women out of 62 who were definitely pregnant gave a positive rat test while in one the test was negative. In 19 of these pregnant women the Friedman test was also done with 18 positive and one negat.ive result. It is interesting to note that in the instance in which the rat test was negative, the Friedman test was positive, and in the case in which the Friedman test was negative, the rat test was positive. One patient in whom the fetus had been dead for three months gave a negative rat test and Friedman test. In 44 instances in which there was no pregnancy, 43 rat tests were negative and one was positive. In 9 of these in which the Friedman test was also done, no positives were obtained. It may be possible that the positive rat test which was obtained in a nonpregnant woman was due to inexperience in interpreting the findings since this was one of the early cases.
BAMDXSTEk
:
&fX
HOT3k
RAT
‘J!EST
POR PREGNANCY
267
Certain technical and economic details may be mentioned. Since the rats are of value for a very short span of their life (about two weeks) it is necessary to have a large colony on hand so that young are always available. If possible, the results should be read in daylight since color determinations are notoriously difficult under artificial light. And it is well to be sure that the individual making the readings is not color blind. For this reason, women often do better than men. In conclusion we feel that, although the number of cases reported is not large enough to warrant an absolute statement as to the value of this test, enough work was done and the results were sufficiently accurate to justify the assumption that this may prove to be a valuable addition to our laboratory tests for pregnancy. It certainly warrants further trial by ourselves and others. Although it does not obviate all the difficulties inherent in biologic testing, this method short,ens t.he time from some 36 or 48 hours down to six hours.
Reference 1. Salmon, V. J., Geist, S. H., Salmon, A. A., and Frank, erinol. 2: 167-170, 1942. 264 BERKELEY
PLACE
I. I>.:
J. Clin. Endo-
N. Y.
NNUMERABLE pregnancy tests have been described in medical literature. Most of them have one or more disadvantages which have kept them from being generally accepted. It was not until Aschheim and Zondek described their technique that there became available a pregnancy test which met most of the requirements for such a procedure. A pregnancy test, in order to be of much value, must meet certain specifications : ‘its degree of accuracy must approach 100 per cent; it must be reasonably rapid and relatively simple. To meet these requirements, much experimental work has been done. It would seem that the ideal test should be a chemical one, thus removing the source of error inherent in all work which depends upon biological assay. Last year, Salmon and Geistl and their associates at the Mount Sinai Hospital used, as a pregnancy test, an observation which had been made many times before. They reported that within six hours after the injection of the urine of pregnant women into suitable rats a definite erythema of the ovaries developed. This was constant and their results compared favorably with those obtained with the Aschheim-Zondek or Friedman tests. In our laboratory, in 107 cases, we have had the same experience. Technique
I
Female rats are used as the test animals. They must be about four weeks old and weigh about 30 grams. Since these animals have a spontaneous estrus cycle and ovulate spontaneously at maturity, they cannot be used when they become much larger. Two animals are employed and two cubic centimeters of urine are injected subcutaneously into eaah. After six hours, the animals are asphyxiated with illuminating gas. In positive cases, the ovarian hyperemia is manifested by a reddening of the ovary. This redness is grossly visible in good light. Usually the uterus, too, is edematous and red looking although the interpretation of the test depends on the ovarian and not the uterine conditions. In the negative cases, the ovaries are small and white and the uterus is a whitme, threadlike structure. In 107 cases, the following results were obtained : *Presented
at a meeting
of the Brooklyn
Gynecological
265
Society,
October
1, 1043.
266
AMERICAN
JOURNAL
RESULTS TOTAL 39
OF
OBSTETRICS
AND
IN PREGNANT
WOMEN
RAT TEST POSITIVE
1st trimester 37 2nd trimester 2 2 3rd trimester 13 13 Duration unknown 9 9 Total 62 61 (eliminating dead fetus ease)
NEQATIVE
2 (a,b)
1
GYNECOLOGY
FRIEDMAN TEST TOTAL POSITIVE NEQATIVE 18 20 2 (be)
19
18
1
a-Test in rat negative but Friedman test positive. Clinically the patient was shown to be pregnant. b-Both rat and Friedman test were negative. In this case the fetus had been dead for about 3 months. c--Friedman test negative but rat test positive. This patient was shown to be pregnant clinically. RESULTS
IN NONPREGNANT TOTAL 15
RAT TEST POSITIVE
WOMEN NEQATIVE
AND IN MALES TOTAL
FRIEDMAN TEST POSITIVE NEQATIVE
Malea Yl 15 0 0 Nonpregnant Women 29 (a) 28 9 0 Totals 44 1 43 9 0 d-As this was one of the early cases, the error may have been due to inexperience in interpreting the findings.
0 9 9
At the beginning of the study, samples of urine were tested from male students, known pregnancies and known nonpregnancy cases. The identity of the subject was not known to the individual reading the result. Later, rat tests were done on the urines submitted for routine Friedman tests and the readings were made before the Friedman tests were completed. In those cases where there was no Friedman test and the samples were received from an outside source, follow-up was requested on the basis of the clinical course. In this way it is felt that all possible sources of error have been covered. After this careful investigation it was found that 61 women out of 62 who were definitely pregnant gave a positive rat test while in one the test was negative. In 19 of these pregnant women the Friedman test was also done with 18 positive and one negat.ive result. It is interesting to note that in the instance in which the rat test was negative, the Friedman test was positive, and in the case in which the Friedman test was negative, the rat test was positive. One patient in whom the fetus had been dead for three months gave a negative rat test and Friedman test. In 44 instances in which there was no pregnancy, 43 rat tests were negative and one was positive. In 9 of these in which the Friedman test was also done, no positives were obtained. It may be possible that the positive rat test which was obtained in a nonpregnant woman was due to inexperience in interpreting the findings since this was one of the early cases.
BAMDXSTEk
:
&fX
HOT3k
RAT
‘J!EST
POR PREGNANCY
267
Certain technical and economic details may be mentioned. Since the rats are of value for a very short span of their life (about two weeks) it is necessary to have a large colony on hand so that young are always available. If possible, the results should be read in daylight since color determinations are notoriously difficult under artificial light. And it is well to be sure that the individual making the readings is not color blind. For this reason, women often do better than men. In conclusion we feel that, although the number of cases reported is not large enough to warrant an absolute statement as to the value of this test, enough work was done and the results were sufficiently accurate to justify the assumption that this may prove to be a valuable addition to our laboratory tests for pregnancy. It certainly warrants further trial by ourselves and others. Although it does not obviate all the difficulties inherent in biologic testing, this method short,ens t.he time from some 36 or 48 hours down to six hours.
Reference 1. Salmon, V. J., Geist, S. H., Salmon, A. A., and Frank, erinol. 2: 167-170, 1942. 264 BERKELEY
PLACE
I. I>.:
J. Clin. Endo-