Amniotic Fluid Acetylcholinesterase Research Articles

The Outcome of a Uniform Program for Alpha‐Fetoprotein Screening in Early Pregnancy in Seven Regions in Sweden

Abstract. In 1979—1980 a multicenter study on serum alpha‐fetoprotein (S‐AFP) screening for detection of fetal neural‐tube and abdominal wall defects (NTD/AWD) was carried out in seven different regions in Sweden. A total of 22991 pregnant women, attending 84 public antenatal clinics, were offered an AFP test between their 15th and 18th completed gestational weeks. The average participation rate was 80.8%. The AFP assays were performed by five hospital laboratories, using the same commercially available assay method. In 19704 women screened, all with known pregnancy outcome, 17/22 fetuses affected by open NTD and 6/8 fetuses having AWD, gave S‐AFP levels exceeding 2 MOM. One placental haemangioma was recognized. Controlsampling was undertaken in 10.5% of the women. Amniocentesis was performed in 1.1% of all women screened with no false negative test results. However, three apparently normal fetuses were aborted at the parents request due to AFP levels in amniotic fluid exceeding 3 MoM.Thus, AFP screening in women at low risk enables detection of 3/4 of fetuses affected by NTD/AWD. A program with higher efficiency would be obtained by more accurate gestational dating by ultrasonography, by complementary diagnostic methods such as neurospecific acetylcholinesterase in amniotic fluid, by a reduction of the work‐load by shifting the action limit to 3 MOM and by continous appropriate training of the medical staff.

A neural tube defect specific form of acetylcholinesterase in amniotic fluid

A neural tube defect specific form of acetylcholinesterase in amniotic fluid

Progressive partial lipodystrophy in association with intrauterine death and growth retardation

Atresia and fistula of the esophagus are among the most common congenital anomalies, but their prenatal diagnosis is only rarely achieved by ultrasonography, amniofetography or cu-fetoprotein (AFP) determination. Intra-amniotic injection of radiopaque dye into the amniotic cavity and radiologic imaging of the fetal gastrointestinal tract have been used most frequently in pregnancies with hydramnios and the sonographic finding of absent fluid in the gastrointestinal tract. It is known, however, that partial passage of the x-ray contrast medium into the fetal intestine in cases of esophageal atresia can occur, thus giving rise to a false negative diagnosis. A simple and reliable biochemical marker present in the amniotic fluid would be of great practical importance. Amniotic fluid acetylcholinesterase as a possible marker for the prenatal diagnosis of esophageal atresia was recently suggested by a case report of a pregnancy characterized by fetal esophageal atresia, polyhydramnios, ambiguous amniofetography, a normal amniotic fluid AFP concentration, and an elevated amniotic fluid acetylcholinesterase level.’ To test the reliability of this proposed marker we retrospectively examined samples of clear amniotic fluid obtained at second-trimester amniocenteses of three pregnancies, each of which proved to be complicated by upper gastrointestinal atresia. Two of these cases had a positive acetylcholinesterase band on gel electrophoresis and normal AFP concentrations. In one case of an esophageal atresia with tracheoesophageal fistula a normal AFP value and no specific acetylcholinesterase band was found on electrophoresis.

Amniotic fluid acetylcholinesterase as a prenatal diagnostic marker for upper gastrointestinal atresias

Amniotic fluid acetylcholinesterase as a prenatal diagnostic marker for upper gastrointestinal atresias

Alphafetoprotein (AFP), concanavalin A non-reactive AFP and specific acetylcholinesterase in amniotic fluid from pathological pregnancies. Predictive values for open spina bifida

Alphafetoprotein (AFP) and concanavalin A non-reactive alphafetoprotein determination and the acetylcholinesterase (AchE) qualitative test have been performed on amniotic fluid samples from 33 normal pregnancies, 44 pregnancies with fetal malformations and 8 normal pregnancies with elevated amniotic fluid alphafetoprotein (3 false positive AFP results, 5 contaminations with fetal blood). The validities of these three tests in detecting abnormal pregnancies are compared. The usefulness of the existing complementary tests in the detection of neural tube defects in a low neural tube defect incidence area is discussed. Risk figures for open spina bifida according to the prior risk situation and the results of maternal serum AFP, amniotic fluid AFP, AchE qualitative test and ultrasound examination have been calculated.

Quantitative enzyme antigen immunoassay of acetylcholinesterase in amniotic fluid.

An enzyme immunoassay for acetylcholinesterase in amniotic fluid is described. Rabbit antiserum (IgG fraction) against human erythrocyte membrane acetylcholinesterase is first attached to microtitre plates. Samples containing acetylcholinesterase are added and the enzyme activity is then measured, with acetylthiocholine as substrate and Ellman's reagent as coupling salt. Results are comparable with those by qualitative determination of the enzyme after polyacrylamide gel electrophoresis, except for blood-contaminated amniotic fluids. One person can perform 200 enzyme analyses per day. Intra- and interassay coefficients of variation are less than 5% at concentrations from 15 to 450 arb. units/L. The mean catalytic concentration in 400 amniotic fluid samples was 21 arb. units/L (range 5 to 70 arb. units/L) with 1000 arb. units/L for a human serum pool as standard. All 39 cases of neural tube defects and two-thirds of cases with omphalocele and gastroschisis had abnormally high values, exceeding 75 arb. units/L.

A potential systematic error in using lysivane as inhibitor in the measurement of amniotic fluid acetylcholinesterase by the Ellman method.

The measurement of low levels of cholinesterase or acetylcholinesterase by the Ellman method requires correction for a non-enzymatic increase in absorption at 412 millimicron that is due both to non-enzymatic hydrolysis of the acetylthiocholine substrate and to modification of the colour reagent. The rate of increase in absorption is dependent on temperature and pH. Addition of an acidic solution of lysivane to the assay solution for selective measurement of amniotic fluid acetylcholinesterase gives rise to a shift in pH; the use of methanol is suggested as an easier method of dissolving the inhibitor and does not affect the pH of the assay, obviating any need to redetermine the background absorption. There is, however, no improvement in ability of the method to predict pregnancies associated with neural-tube defects.

Amniotic fluid acetylcholinesterase as a marker in prenatal diagnosis of esophageal atresia

Amniotic fluid acetylcholinesterase as a marker in prenatal diagnosis of esophageal atresia

Acetylcholinesterase in neural tube defects: A model using chick embryo amniotic fluid

Acetylcholinesterase was measured in amniotic fluid from normal chick embryos and embryos with neural tube defects. Neural tube defects were induced in the chick embryos by three procedures, removal of albumen, mechanical disruption of the closed neural tube or injection of tetanus toxin. The concentration of acetylcholinesterase in amniotic fluid from untreated normal embryos changed throughout the period examined (5–14 days incubation) but was stable at 0.5 Ul −1 over the time period 6–11 days. Amniotic fluid taken from treated embryos with neural tube defects at 8 days always contained a higher concentration of acetylcholinesterase than fluid from sham operated but otherwise normal embryos, mean 40.9 Ul −1,S.E.M. = 10.1 Ul −1, versus 1.0 Ul −1,S.E.M. = 0.2 Ul −1. The range of values (6.1–393 Ul −1) was clearly separated from the normal values, range 0.0–5.5 Ul −1. In 13 cases with developmental abnormalities other than neural tube defects, the concentration of acetylcholinesterase was elevated in only one. Two different forms of acetylcholinesterase, as shown by gel electrophoresis, were present in fluid form both normal and defective embryos. These forms were also present in blood plasma, cerebrospinal fluid and in the high speed supernatant from brain extracts, the latter tissue contained an additional form of greater electrophoretic mobility. After irreversible inhibition, enzyme activity in amniotic fluid recovered slowly; only half the control value was reached by 140 h compared with complete recovery in the tissues of the embryo within 19 h. Histochemical staining for acetycholinesterase showed that the spinal cord in the region of the lesion contained high concentrations of the enzyme. The possible sources of acetylcholinesterase in amniotic fluid are discussed. This chicken model of neural tube defects provides support for the use of acetylcholinesterase tests in the detection of neural tube defects clinically, and provides a model for experimentation with this system.

Prenatal diagnosis of open neural tube defects: Milunsky A, Sapirstein V. Prenatal diagnosis of open neural tube defects using the amniotic fluid acetylcholinesterase assay. Obstet Gynecol 59:1 (1982)

Prenatal diagnosis of open neural tube defects: Milunsky A, Sapirstein V. Prenatal diagnosis of open neural tube defects using the amniotic fluid acetylcholinesterase assay. Obstet Gynecol 59:1 (1982)

Amniotic Fluid Acetylcholinesterase Electrophoresis as a Secondary Test in the Diagnosis of Anencephaly and Open Spina Bifida in Early Pregnancy

C. R. F. Cancer Epidemiology and Clinical Trials Unit. University of Oxford. Radcliffe Infirmary. Oxford. England

Amniotic fluid acetylcholinesterase: a retrospective and prospective study of the qualitative method.

After retrospective evaluation with stored samples, the qualitative acetylcholinesterase (AChE) test has been used prospectively in conjunction with alpha-fetoprotein (AFP) assay on 986 amniotic fluid specimens received in this laboratory during 1980. The main value of AChE is in classifying fluids in which the AFP level is near the threshold between normal and abnormal. Among abnormal pregnancies with raised AFP levels, neural-tube defects can generally be distinguished from other abnormalities by careful appraisal of the AChE gel pattern, but confirmation of these other fetal abnormalities may require high resolution diagnostic ultrasonography and perhaps fetoscopy. Neural-tube defects with false negative AFP levels can be detected by AChE, but AChE gives occasional false positives so that it cannot be relied on in isolation for the diagnosis of neural-tube defects.

Chapter 4. Prenatal Diagnosis of Open Neural Tube Defects

This chapter describes the prenatal diagnosis of open neural tube defects (NTD). Spina bifida and anencephaly, which collectively account for nearly all NTD, are serious fetal malformations resulting from incomplete closure of the neural tube during embryogenesis. The two occur in approximately a 1:1 ratio, and roughly 20% of spina bifida lesions are skin-covered. Mean alpha-fetoprotein (AFP) concentrations in amniotic fluid fall steadily throughout the second trimester, emphasizing the need to document gestational age before interpreting test results. AFP is the major fetal serum protein during the early weeks of pregnancy, occurring in concentrations as high as 300 mg/100 ml at the twelfth week of gestation. It is found that about 40% of anencephalic and 20% of open spina bifida pregnancies are associated with bloodstaining. Brown samples also appear on occasion. These latter rarely have intact red cells but may have detectable hemoglobin F. Rather than being procedure-related, these brown samples are felt to indicate an earlier spontaneous hemorrhage. A report indicates that identification of a specific molecular form of acetylcholinesterase (AChE) in amniotic fluid can be useful both in diagnosing neural tube defects and in identifying false-positive AFP results.

Quantitative and qualitative assay of amniotic-fluid acetylcholinesterase in the prenatal diagnosis of neural tube defects

In 110 amniotic fluids the specific acetylcholinesterase was determined quantitatively and qualitatively. In the quantitative assay there were a considerable number of false positives and false negatives, although the mean value of the normal controls differed significantly from that of neural tube defect pregnancies. By the electrophoretic separation of acetylcholinesterase, however, all fluid samples with borderline alpha-fetoprotein levels or fetal blood contamination could be correctly classified. With the exception of one skin-covered spina bifida all neural tube defects in the second trimester could be identified by this method. The second fast-moving band characteristic of the specific acetylcholinesterase was also present in abdominal wall defects and intrauterine death.

Particulate acetylcholinesterase in amniotic fluid and its implications for neural tube defect screening.

Acetylcholinesterase activity in mid-trimester amniotic fluid has been determined by a quantitative spectrophotometric method and a qualitative electrophoretic technique in a limited (76 samples) retrospective study. Differential centrifugation studies on amniotic fluid from normal pregnancies demonstrated the presence of particle-associated acetylcholinesterase sedimenting only at relatively high centrifugal forces. It is postulated that this particle-associated activity was a contributory factor to an observed false-positive incidence of 6.6% when quantitative acetylcholinesterase measurement was used to discriminate between normal pregnancies and those associated with neural tube defects. Discrimination on the basis of electrophoretic analysis of acetylcholinesterase resulted in no misclassification of pregnancies. Particulate acetylcholinesterase would not be expected to influence the interpretation of the electrophoretic pattern due to its size-related exclusion from gel pores. The implication of these observations in the accurate prenatal diagnosis of neural tube defects is discussed.

AMNIOTIC FLUID ACETYLCHOLINESTERASE ELECTROPHORESIS AS A SECONDARY TEST IN THE DIAGNOSIS OF ANENCEPHALY AND OPEN SPINA BIFIDA IN EARLY PREGNANCY: Report of the Collaborative Acetylcholinesterase Study

Fifteen centres collaborated in a study to determine the value of amniotic fluid acetylcholinesterase (AChE) electrophoresis in the diagnosis of fetal open neural-tube defects (NTDs) between 13 and 24 weeks of gestation among women with positive amniotic fluid alpha-fetoprotein (AFP) results. Pregnancies with positive amniotic fluid AFP results were identified from about 34 000 tested; on average they had values greater than the 99·6th percentile for non-NTD pregnancies. Amniotic fluid samples were available from 1099 pregnancies with positive AFP results, and the percentages which yielded positive AChE results were as follows: anencephaly 99·6% (476/478); open spina bifida 99·4% (333/335); non-NTD pregnancies (i) without serious malformation and not ending in miscarriage 6% (8/125), (ii) without serious malformation but ending in miscarriage 47% (34/73), (iii) exomphalos 75% (47/63), (iv) congenital nephrosis 0% (0/11), (v) other serious malformations 50% (7/14). The gel-AChE test was thus found to be very effective when applied to amniotic fluid samples with positive AFP

Gel electrophoresis of amniotic fluid acetylcholinesterase as an aid to the prenatal diagnosis of fetal defects

The presence or absence of a specific acetylcholinesterase (AChE) band was determined by polyacrylamide gel electrophoresis on 272 second trimester amniotic fluid samples. The AChE band was absent from 176 normal samples, including seven which had been scored as false positives by alphafetoprotein (AFP) assay. It was present in all 30 samples from open neural tube defects, of which four had been scored as false negatives by AFP assay. In the remaining 66 pregnancies with abnormal outcome, an AChE band was in general present when AFP was raised and absent when it was normal. However, all six cases of congenital nephrosis had raised AFP and no AChE band, while two of 30 pregnancies ending in spontaneous abortion had an AChE band and normal AFP. These results suggest that AChE electrophoresis is a valuable confirmatory technique for the early prenatal diagnosis of fetal abnormalities.

Amniotic fluid acetylcholinesterase and prenatal diagnosis.

Raised specific acetylcholinesterase (AChE) activity in amniotic fluid was consistently found to be associated with fetal intrauterine death and neural tube defects, but in just under half of the samples from pregnancies with spina bifida, the increase was marginal. Elevated AChE levels were occasionally found in fluid samples contaminated by maternal blood, and in nearly half of the samples which were a brown or yellow-brown colour but where there was a normal fetus. Nine fluids, five of which were contaminated with fetal blood, had a false positive result on alpha-fetoprotein (AFP) assay (elevated AFP levels and a normal fetus), but showed normal AChE activities. It is suggested that AChE estimation might be useful in prenatal diagnosis as a second test for those fluids which have an elevated AFP level.

Amniotic fluid acetylcholinesterase isoenzyme patterns in the diagnosis of neural tube defects

Acetylcholinesterase isoenzyme patterns were determined in amniotic fluid from 140 normal pregnancies and from 68 pregnancies with various fetal abnormalities. A fast acetylcholinesterase isoenzyme was detected in all cases of neural tube defect, exomphalos and intrauterine death. The fast isoenzyme was not seen in any of the normal pregnancies. Blood-contaminated amniotic fluids with false positive elevation of alphafetoprotein gave a single isoenzyme band. The qualitative demonstration of a fast acetylcholinesterase isoenzyme in amniotic fluid provides a useful adjunct to the alphafetoprotein assay in the prenatal diagnosis of neural tube defects.

Amniotic-Fluid Acetylcholinesterase as a Possible Diagnostic Test for Neural-Tube Defects in Early Pregnancy

Raised levels (greater than or equal to 4.5 munits/ml) of acetylcholinesterase (AChE) activity in amniotic fluid at 14--23 weeks of pregnancy were significantly associated with open fetal neural-tube defects. Out of 72 pregnancies correctly classified by the amniotic-fluid alpha-fetoprotein (A.F.P.) test, 2 of 56 without neural-tube defects and all 16 with open neural-tube defects (8 with anencephaly and 8awith open spina bifida) had raised levels of AChE. Out of 5 pregnancies misclassified by the A.F.P. test (4 without neural-tube defects and 1 with open spina bifida), only 1 was misclassified by the AChE test--namely, one of those without a neural-tube defect. Thus, only 3 of the 77 pregnancies tested were misclassified by the quantitative AChE test. A qualitative test for an isoenzyme of AChE found in cerebrospinal fluid correctly classified these 3 pregnancies. These findings suggest that the analysis of AChE in amniotic fluid may be a useful test in the diagnosis of open neural-tube defects.