Improved quantitative Apt test for detecting fetal hemoglobin in bloody stools of newborns

We devised an improved spectrophotometric method, based on the differential susceptibility of various hemoglobins to alkaline denaturation, to measure the percentage of fetal hemoglobin (HbF) in bloody meconium so as to determine the source of the blood. The oxyhemoglobin of a sample was spectrophotometrically scanned from 450 to 650 nm before and after the addition of sodium hydroxide. With Allen baseline correction, the ratio of absorbance at 576 nm was linearly proportional to the percentage of HbF in the specimen as described by the equation: ratio = 0.01 (%HbF) + 0.0045 (r = 0.9971, Sy/x = 0.047). Preliminary studies on samples containing 10% or 80% HbF revealed that the method was fairly precise. The accuracy of this test was verified by comparing the results (y) with those obtained by radial immunodiffusion (x): y = 0.997x + 2.93 (r = 0.91). This preliminary study demonstrates that the modified test is simple, fast, and free from bilirubin interference. It should be useful for the diagnosis of gastrointestinal bleeding of the newborn on a stat basis.