Performance of the amniotic fluid foam stability—50 percent test

Pe1formance of the amniotic fluid foam stability50 percent test A bedside precedure for the prenatal detection of hyaline metnbrane disease GEOFFREY SHER, F.A.C.O.G., M.R.C.O.G.(LoND.), F.C.O.G.(S.AFR.) BER.\JARD E. STATLAND, M.D .. PH.D. DENNIS E. FREER, M.S. JOH \J C. HISLEY, F.A.C.O.G. Chapei Hill and CharlottP, North Carolina, and Sacramento, California A study was conducted on 394 patients in order to evaluate a modified foam stability (FS) test for the pre·natal evaluation of fetal pulmonary maturity. We termed the assay the "FS-50 test" because of its dependence upon the maintenance of a 50% ethanol concentration in the final assay mixture. The FS-50 test was regarded as "positive" when an uninterrupted ring of stable bubbles formed around the meniscus of the tube at the air-fluid interface, following vigorous shaking. Anything less was re!~arded as a "negative" result. Neonatal hyaline membrane disease (HMO) did not occur in association with a "positive" FS-50 test result. However, when the FS-50 test was "negative," 35.8% of the neonates developed HMO. In 164 of the 394 patients studied, sufficient amniotic fluid was available so as to enable us to perform the FS-50 and the lecithin/sphingomyelin (L/S) ratio assay c:oncurrently. LIS ratio values of greater than 2.0 correctly predicted a sufficient degree of fetal pulmonary maturity so as to protect against the subsequent development of HMO in 97.8% of the cases. LIS ratio values of less than 2.0 were predictive of HMO in 28.4% of the cases studied, while a "negative" FS-50 test correctly predicted that HMO would subsequently occur in 46.9% of the cases. The FS-50 test promises to provide a reiiabie screening method ior the prenatai detection of HMO. The practicability and sensitivity of this simple assay makes it a potentially valuabl•3 addition to our diagnostic armamentarium. (AM. J. Oasrer. GYNECOL. 134:705, 1979.)

THE IDEAL prenatal test of fetal pulmonary maturity should be simple, sensitive. accurate, and inexpensive. At present, the two most widely used methods of predicting fetal pulmonary maturation are the lecithin/ sphingomyelin (LIS) ratio' and foam stability tests. 2 - 4 The LIS ratio has been found to be reliable in normal obstetric cases but has shown less predictive value in From the De/Jartment of Obstetrics and Gvnecoloi!V. North Carolina M;morial H;spital, the Depart~ent a/Hospital Laboratories, University of California, Davis, and the Departme rzt of Obstetrics and G)•necolog;·, Charlotte

Memorial Hosf;ital. Received/or publicationjulv 25, 1978. Revised S'ptember 12, 1978. Accepted September 18, 19 78. Reprint requests: Dr. Geoffrey Sher, Reno Women's Clinic, 129 W. Sixth St., Reno, Nevadn 89503 0002-9378/79/ 140705+03$00.30/0 © 1979 The C. V. Mosby Co.

complicated situations. 5- 7 The assay is time consuming, requires sophisticated equipment and technical expertise, and in most laboratories cannot be performed on a stat basis. We recently reported on a preliminary evaluation of a modified amniotic fluid, foam stability test for the prenatal detection of neonatal hyaline membrane dis· ease (HMD). 4 The assay is a modification of a procedure first described by Clements and co!!eagues 2 in 1972. We termed the assay the FS-50 test because it makes use of 100% ethanol in a I : I ratio with uncentrifuged amniotic fluid to produce an ethanol volume fraction of 50% in the final assay mixture. This distinguishes it from the procedure originally described by Clements' group in which the final ethanol volume fraction in the assay mixture was 47.5%. We were able to demonstrate that a positive result using the FS-50 705

706 Sher et al. ·\rn

Table I. Clinical presentations of the 394 patients No. of

Clinical presentations

cases

Normal pregnancy (including elective cesarean sections) Hypertension (toxemic and nontoxemic) Preterm delivery Diabetes mellitus (insulin-dependent) Intrauterine growth retardation Rhesus disease Third trimester bleeding Chronic renal disease Total

160 94 83 47 20 17 12 5 438

Table II. FS-50 test versus clinical outcome 1Veortatal pubnonai) status FS-50 test

Normal

HMD

+

285 70

39

0

Table III. LIS ratio and FS-50 results versus incidence of HMD LIS ratio FS-50test

+ Total

<1.0 j1.1-1.5j1.6-2.0 j2.1-3.0

0/1 * 8/10 8/11

0/7 12/17 12/24

0/27 1/12 1/39

0/38 2/9 2/47

I

>3.0

Total

0/42 0/1 0/43

0/llS 23/49 23/164

*Ke =Cases of HMD Y Total cases

Table IV. Predictive values of LIS ratios and of FS-50 test results Coruiitional probability

Based on 164 cases(%)

Based on 394 cases(%)

P(Normal I LIS > 2.0) P(HMD LIS ,;;;; 2.0) P(Normal FS-50 plus) P(HMD FS-50 minus)

97.8 28.4 100 46.9

100 35.8

I I I

test provided lOOo/c .~ssurance of protection against the subsequent development of HMD in the neonatal period. 1 While the results of our preliminary study were extremely encouraging, the sample size was relatively small (i.e., only eight cases of HMD in a total of 104 patients studied) and the assay "vas perf{)rmed under optimal conditions by a laboratory-trained technologist.3 This present study presents an evaluation of the FS-50 test under routine clinical conditions with the assay being performed by a number of clinicians as a

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Chapel Hill and the Charlotte \1emorial Ho~pital in Charlotte; both are institutions affiliated with the University of :\'orth Carolina School of \lcdiLinc ..c\mniotic tluid was obtai ned bv amniocentesis f>erlormed for clinical reasons . l'ulmonan status of the neonate was deli ned as either ··hvaline metnhrane disease"' or '"normal"' on the basis ol criteria previoush tldined in our earlier publication.' 'vVe perfitrmed the FS-:"JO rest on each uf the amniotic Huid specimens obtained from the :~94 patients tileluded in the study. Both institutions had at their disposal prcviouslv pro\·ided Kin1ax glass tube~ tneasuring 5 em x 1.:-1 t m. each of which contained exanlv 1.0 ml of 100~{ (absolute) ethanol \\·hich had been dispensed into the tubes in a moisture-free environment. All tubes were scaled with an air-tight, screw-on cap. This procedure insured maintenance of the anhydrous nature of the absolute ethanol reagent. The procedure of the FS-50 tt:st ,,·as performed as follows: One milliliter of freshh obtained amniotic fluid was dispensed into a glass tube containing l.tl nil of ethanol. The tube was recapped and then vigoroush shaken for a period of 30 seconds. It was then allowed to settle for I :i seconds after which the air-fluid interface ''as examined for the absence or presence of stable foam. The presence of an uninterrupted ring ol bubbles around the entire meniscus of the tube was interpreted as .. positive" and anything le~' was called ··negatin···.' In [()~ of the :l~J.t patients studied, the lecithin/ sphingomyelin (LIS) ratio w,ts also performed on the same spenmen ol amniotic Huid. !he LIS ratio was performed according t.o the method described by Gluck and associates 1 and included the acetone precipitation step.

Results ~rable II ctHnpares the results obtained \Vith the FS-50 test versus neonatal clinical outcome. The incidence of hyaline membrane disease was 119(,. In no instance did hyaline membrane disease occur in association with a "positive"" FS-:JO test. HMD ocrurred in

Volume 134 Number 6

39 of 109 cases where the FS-50 test was "negative'' (i.e., 35.8%). Table III depicts the results obtained on each of the 164 cases where both the LIS ratio and FS-50 test were performed on the same specimen. In this subset, the incidence of hyaline membrane disease was approximately I4'7c
Amniotic fluid foam stability-50% test 707

States alone it is responsible for approximately 25,000 deaths and numerous serious sequelae each year. The major method for the prenatal detection of this disease is the LIS ratio which is employed to measure surfactant in the amniotic Huid. Unfortunately, however. this assay is expensive and requires the availability of a significant amount of technical expertise. In Addition, it is time consuming and therefore is not readily available on a 24 hour stat basis. Moreover. we as well as other workers have noted hyaline membrane disease in association with LIS ratio Yalues of greater than 2.0, especially in pregnancies complicated by diabetes mellitus and Rhesus disease. 4 • ,_ 7 In addition, this studv revealed that when LIS ratio values were less than 2.0, 71.6'7c of the neonates had mature lungs, as compared with 53.1% when the FS-50 test was "negative." The FS-50 test as described here, by providing prefilled glass tubes containing the required amounts of absolute ethanol, significantly simplifies the shake test procedure, thereby reducing the likelihood of variation in the final ethanol volume fraction. We are convinced that it is this factor more than any other which has avoided the occurrence of "false positive" results in our study. Ail that is required of the analyst (e.g., the clinician doing the assay) is the ability to pipette the prescribed volume of amniotic fluid and then to shake the assay mixture for 30 seconds. \>\'bile the results would suggest that the FS-50 test provides a reliable screening method for hyaline membrane disease, final clinical acceptance of this assav must a\vait the outcome of a more extensive studv.

Comment Hyaline membrane disease is one of the major causes of perinatal death throughout the world. In the United

REFERENC:ES

I. Gluck, L., Kulovich, M. V ., Borer, R. C., Jr., et a!.: Diagnosis of the respiratory distress syndrome by amniocen· tesis, AM. j. 0BSTET. GYNECOL. 109:440, 1971. 2. Clements, J. A., Platzer, A. C. G., Tierney, D. F., et al.: Assessment of the risk of the respiratory distress syndrome by a rapid test for surfactant in amniotic fluid, N. Engl. J. Med. 286:1077, 1972. 3. Edwards, J., and Baillie, P.: A simple method of detecting pulmonary surfactant activity in amniotic fluid, S. Afr. Med.]. 47:~~070, 1973. 4. Statland, B. E.. Sher, G., Freer, D. E., et al.: Evaluation of a

modified foam stability (FS-50) test. Am. ]. Clin. Pathol. 69:514, 1978. 5. Doran, T. A., Malone, R. M., Benzie, R. T., et al.: Amniotic fluid tests for fetal maturity in normal and abnormal pregnancies, AM.]. OBSTET. GYNECOL. 125:586, 1972. 6. Gluck, L., and Kulovich, M. V.: Lecithin/sphingomyelin in ratios in amniotic fluid in normal and abnormal pregnancy. AM. J. OasTET. GYNECOL. 115:539, 1973. 7. Morrison,]. C., Whybrew, W. D., Bucovaz, E. T., et al.: Amniotic fluid tests for fetal maturity in normal and abnormal pregnancies, Obstet. Gynecol.. 49:20, 1977.