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7 September 1963 S.A. TYDSKRIF VIR GENEESKUNDE 931 COMPARISON OF THE XENOPUS LAEVIS PREGNANCY TEST WITH AN HAEMAGGLUTINATION INHIBITION (pREGNOSTICON) TEST * W. M. POLITZER, MD. and F. E. SIMPSON, South African Institute for Medical Research, Johannesburg The present work deals with a study of the Xenopu5 TABLE J. COMPARISON OF RESULTS OF Xenopus laevis TEST laevis pregnancy test' carried out in parallel with a test WITH HAEMAGGLUTINATION INHIBITION TEST based on haemagglutination inhibition (HI). In this test Xenopus laevis HI test the antigen is human chorionic gonadotrophin (HCG), Number of test results results and erythrocytes sensitized to HCG serve as antigen car- riers. Addition of antiserum causes a variation in the sedi- specimens Positive Negative Positive Negative mentation pattern of the erythrocytes. In the presence of 43 43 43 pregnancy urine containing HCG (1,000 IV/I. or more) 57 57 57 sedimentation is prevented and a brown ring is formed 12 12 12 (positive result). In the absence of HCG, or when less than 1,000 IV / 1. are present, a homogeneous yellow-brown 3 3 3 sediment remains (negative result). In 15 cases discrepancies in the results occurred. Follow- MATERIAL AND METHODS up studies revealed the following data: in 12 cases giving negative Xenopus laevis and positive HI test results, 7 One hundred and fifteen specimens of urine received by were found to have normal pregnancies and one was a this Institute for routine pregnancy tests were subjected patient who died 2 weeks later of uraemia and hyperten- to the Xenopus laevis test and to the HI test. The precipi- sion; in this case examination per vaginam excluded preg- tation method used for the Xenopus laevis test was that nancy. In the remaining 4 cases (3 abortions and 1 ectopic described by Scott,' each of 4 frogs being injected with 2·5 pregnancy) the negative Xenopus laevis and positive HI m!. of extract corresponding to 50 rnI. of urine. tests were both regarded as correct. The HI (pregnosticon) system is composed of: Of the 3 patients with positive Xenopus laevis and nega- (a) Antiserum from rabbits immunized with HCG. (In tive HI test results, 2 were subsequently found to be preg- the freeze-dried state this is stable for 12 months when nant. The urine of the third patient gave a weak-positive stored in the refrigerator.) Xenopus laevis test result, i.e. lout of 4 frogs ovulated, (b) Erythrocytes sensitized to HCG. and this patient was found to be non-pregnant. (c) Suspension fluid for (b) which, when prepared, is The accuracy of the results was as follows: stable at 0 - 4°C. for 1 week. (d) Positive and negative control urine specimens sup- Xenopus laevis test HI test plied freeze-dried in vials. Correct positive 87/,0 96% All glassware used must be scrupulously clean, without Correct negative 98% 98/,0 the use of soaps and detergents. The method employed was as follows: DISCUSSION 4 m!. of suspension fluid was placed into the vial con- To our knowledge, there have been no studies comparing taining the erythrocytes sensitized to HCG, and shaken the Xenopus laevis test with the HI (pregnosticon) test. until a homogeneous suspension was obtained. After cen- The only information available is that of Wide' who trifugation 0'1 rnI. of urine was placed into the ampoule carried out the HI in various dilutions on 2,701 urine containing the freeze-dried antiserum, 1 rnI. of distilled specimens from 1,012 women and obtained the following water was placed into the vials containing the freeze-dried accuracy: correct positive 99,8%, correct negative 99'8/'0' positive and negative urine controls, and aliquots of 0·1 An interesting finding was that the urine of 15 women mt. were treated in the same way as the test urine. with early pregnancy, i.e. 31 - 37 days after the last men- Finally, 0·4 m!. of homogeneous erythrocyte suspension strual period, 8 gave positive results, I gave a negative was added to the antiserum/urine solution in the original and 6 gave doubtful results. In our study, 7 gave a posi- ampoule. tive HI test result at a time when the Xenopus laevis test The ampoule was placed in a specially designed rack was still negative, which indicates that the HI test is the which was shaken for approximately I minute. The rack more sensitive test. A further advantage of this test is its was then left standing undisturbed at room temperature simplicity, i.e. 40 tests can be set up and read in just over in a position away from heaters, sunlight, and vibration. 1 hour. Exactly 3 hours later the results were read and the pat- All pregnancy tests depend on the detection of HCG terns were interpreted as follows: positive result, a brown in the urine, and besides diagnosing normal pregnancy, ring at the bottom of the tube; negative result, the homo- are of great value in.detecting conditions such as hydatidi- geneous yellow-brown suspension remains. form mole, choriocarcinoma and malignant tumours of the testis. Pituitary gonadotrophin may also give a posi- RESULTS tive Xenopus laevis test result. In fact, in the Xenopus The results are shown in Table I. laevis test on a non-pregnant patient who was 28 years of age, lout of 4 frogs ovulated. Both pituitary and * This article was received too late for inclusion in the chorionic gonadotrophin are glycoproteins, and although supplement: South African Journal of Obstetrics and Gynae- cology-hence its publication here-Editor. pituitary gonadotrophin differs chemically from the 932 S . A.. MED I C A L J 0 URN A L 7 September 1963 chorionic hormone, the two substances cannot be differ- SUMMARY entiated by means of immunological and biological pro- The Xenopus laevis and the HI (pregnosticon) tests were cedures. During menopause there is a higher excretion of carried out on 115 specimens of urine submitted for preg- pituitary gonadotrophin. In the HI test concentrated urine nancy testing. The accuracy obtained was as follows: is used, and a false positive result may be obtained. In order to differentiate between amenorrhoea owing to Xenopus laevis test HI test pregnancy and menopause, the manufacturers suggest Correct positive 87% 96% diluting the urine with an equal amount of water which Correct negative 98/"0 98% will lower the concentration of urinary pituitary gonado- The HI test described is simple and rapid and the accu- trophins sufficiently to render a negative result. The speci- racy obtained compared favourably with the Xenopus mens submitted to this Institute are not always accom- laevis test. panied by detailed information, and difficulty may be We wish to thank the Director of the South African Insti- encountered in establishing the age of the patient. tute for Medical Research for permission to publish and the many gynaecologists and general practitioners for their assis- In 2 pregnant patients positive Xenopus laevis test re- tance in follow-up studies. We also wish to thank Organon sults were obtained with negative HI test results. As the Laboratories Ltd., London, and their local agents, Keatings Pharmaceuticals Ltd., for the material supplied. HI test is carried out in concentration only, sedimentation REFERENCES may have taken place owing to non-specific agglutinins, I. Scat!, L. D. (1940): Brit. J. Exp. Path.. 21. 320. but if dilutions had been carried out, sedimentation 2. Wide. L. (1962): Acta Endocr. (Kbh.), 41. suppl. 70. 3. Shapiro, H. A. and Zwarenstein, H. (1933): Proc. Ray. Soc. S.A., might have been prevented and a positive result obtained. October 1933 (see Trans. Ray. Soc. S.A., 1934, 22, LXXV). PASSING EVENTS: IN DIE VERBYGAAN Dr. Kalman Ezekowitz, specialist anaesthetist, has commenced Dr. Gerald M. Lurie, paediatrician, of Cape Town, has practice at 1004 S.A. Mutual Buildings, Gardiner Street, Dur- changed his residential address to 'Albatross', Blair Road, ban. Telephones: Rooms 31-2660, residence 8-5272. Camps Bay. Telephone 39-7348. Dr. Kalman Ezekowitz, narkotiseur, het begin praktiseer te * * '" S.A. Mutual-gebou 1004, Gardinerstraat, Durban. Telefoon: College of General Practitioners, Cape of Good Hope Faculty. Spreekkamers 31-2660, woning 8-5272. A symposium on Asthma will be held at the Vineyard Hotel, Newlands, Cape, on Tuesday 1 October 1963, at 8.15 p.m. The '" * * Dr. B. Gentin, intemis, het begin praktiseer in vennootskap speakers will be Dr. Hendrik Muller, physician, Dr. S. C. Shore, paediatrician, Dr. A. W. Spratt, general practitioner, met dr. T. G. Armstrong te Norwich Union House 403, and Dr. N. V. Storr, allergist. The talks will be followed by Smithstraat, Durban. Telefoon: Spreekkamer 2-8328, woning an open discussion and refreshments will be served. The 2-6912. meeting is being sponsored by Warner Pharmaceuticals and Dr. B. Gentin, specialist physician, has commenced practice all doctors are cordially invited to attend. in partnership with Dr. T. G. Armstrong at 403 Norwich Union House, Smith Street, Durban. Telephones: Rooms * * * Society for the Promotion of Undergraduate Research. A 2-8328, residence 2-6912. meeting of the ucr Medical School Society for the Promo- * * * tion of Undergraduate Research (SPUR) will be held on Mon- South African Paediatric Association (M.A.S.A.), Natal Sub- day 9 September at 8 p.m. in the Pharmacology Lecture group. The next meeting of this Subgroup will be held on Theatre, Medical School, Observatory, Cape. The following Tuesday 10 September 1963 at 8 p.m. in the Medical School, student papers will be read: 'Myasthenia gravis and the Durban. Dr. N. M. Mann will speak: on his recent travels in thymus', by E. J. Coetzee; and 'Art in psychiatry', by I. N. the USA and some problems of the newborn. Bernadt. All interested persons will be welcome at the meeting. INTERNATIONAL CONFERENCE ON THE PORPHYRIAS An International Conference on the Porphyrias will take Dr. Allan G. Redeker, School of Medicine, University of place in Cape Town at the Department of Medicine, Medical Southern California, Los Angeles. School, University of Cape Town, from 18 - 20 September Dr. A. Magnus, Institute of Dermatology, St. John's Hos- 1963. A visit will also be paid to the Karl Bremer Hospital pital for Diseases of the Skin, London. on 21 September. The Conference is sponsored by the Coun- Dr. Birgitta Haeger-Aronson, Department of Clinical cil for Scientific and Industrial Research, and the Conference Chemistry, Malmo General Hospital, University of Lund, proceedings will be opened by Prof. D. Craib, Associate Ad- Malmo, Sweden. viser in Medical Research to the CSIR. In addition to a number of South African workers on the Attendance at this Conference is by invitation. porphyrias, the following overseas visitors, all of whom are After the Conference in Cape Town the overseas visitors experts in this field, will be attending: will leave on 21 September for visits to the medical schools at Prof. C. Rimington, Professor of Chemical Pathology, Uni- Durban, and later Johannesburg and Pretoria, and also to versity of London, University College Hospital Medical the Onderstepoort Veterinary Institute. School, London. The purpose of this Conference is: Dr. A. Goldberg, Senior Lecturer in Medicine, University 1. To study the porphyrias as seen in South Africa, which Department of Medicine, Gardiner Institute, Western Infir- has the unenviable distinction of having the highest incidence mary, Glasgow. of porphyrias in the world; Dr. H. D. Barnes, Department of Chemical Pathology, St. Mary's Hospital, London. 2. To discuss the research work done in this field at the Dr. A. M. Gajdos and Madame Gajdos-Torok, Department various medical schools; and of Medicine, Hotel-Dieu, Paris. 3. To come to some agreement if possible on the classifi- Prof. R. Schmid, Department of Medicine, University of cation and nomenclature of the porphyrias. Other controver- Chicago. sial matters will also be discussed.
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