ARE THE CYTOKINES INTERLEUKIN-6 AND ANGIOGENIN STABLE IN FROZEN AMNIOTIC-FLUID

OBJECTIVE: To determine the stability of cytokines in frozen stored am niotic fluid samples, we measured angiogenin, a potent inducer of neov ascularization and interleukin-6, an inflammatory cytokine, in the sam e sample of midtrimester amniotic fluid 1 year apart. STUDY DESIGN: In this study paired aliquots of amniotic fluid kept at -70 degrees C we re immunoassayed for angiogenin and interleukin-6 at two different tim e periods 1 year apart. Inclusion criteria were (1) samples with clear ly identifiable numbers, (2) no evidence of breaks in sealing of sampl es, and (3) singleton gestation. Amniotic fluid was immunoassayed for angiogenin and interleukin-6 in July 1995 and 1996. Angiogenin sensiti vities were 0.078 and 0.026 ng/ml, interassay coefficients of variatio n were 3.8% and 4.6%, and intraassay coefficients of variation were 2. 7% and 2.9%, respectively, in 1995 and 1996. Interleukin-6 sensitiviti es were 1.74 and 2.37 pg/ml, interassay coefficients of variation were 8.9% and 2.6%, and intraassay coefficients of variation were 3.5% and 1.9%, respectively, in 1995 and 1996. Statistical analysis included p aired t test, Wilcoxon signed-rank test, and regression with p < 0.05 significant. Angiogenin and interleukin-6 values were normalized with natural log transformation for statistical analysis. RESULTS: Paired a mniotic fluid samples from 30 patients were immunoassayed from 1993 to 1995. The values of angiogenin were significantly lower in the 1996 a ssay compared with the 1995 assay (median 12.4 [range 5.6 to 61.3] vs 26.7 [range 13.6 to 159.2] ng/ml, p < 0.001). A significant correlatio n was found between the change in angiogenin levels and the year of th e sample, with older samples having the greatest change in values (r = 0.5, p = 0.008). The values of interleukin-6 were significantly lower in the 1996 assay compared with the 1995 assay (median 230.8 [range 4 0.9 to 3711.3] vs 289.2 [range 53.7 to 19100.0] pg/ml, p < 0.001). CON CLUSIONS: Angiogenin and interleukin-6 values in amniotic fluid appear to decrease with time despite optimal freezing conditions. The year o f sampling and length of storage should be taken into consideration wh en evaluating amniotic fluid cytokine levels from stored samples.