Continuous reference curves for hematology and biochemistry analytes in pregnancy - a prospective study

Pregnancy involves dynamic metabolic changes that cannot be accurately interpreted using population reference intervals (RI). The aim was to develop continuous reference curves for common hematology and biochemistry laboratory analytes in pregnancy and to assess their agreement with the RIs for non-pregnant women, which are routinely reported for pregnant women.

A prospective study was conducted at the Clinical Hospital Centre Rijeka, Croatia, from September 2022 to December 2025. Inclusion criteria were age ≥18 years, singleton pregnancy, normal ultrasound and prenatal screening findings, and favorable pregnancy outcome. Exclusion criteria were illness in the last month, pregnancy-related complications, medically assisted fertilization, and chronic medication use. All participants provided informed consent and completed a questionnaire on lifestyle habits and health status. Continuous reference curves for 43 laboratory analytes were generated using the mathematical-statistical GAMLSS model. Agreement with the discrete RI was expressed as the percentage deviation from the continuous reference curves.

The analysis included 739 samples from 542 pregnant women between the 11th and 38th gestational weeks. Of the 43 laboratory analytes, 35 showed discrepancies when compared to non-pregnant RI. Falsely elevated interpretations ranged from 5.6 to 64.9 %, and falsely lowered from 3.4 to 62.0 %. Graphical representations of the continuous reference curves show pronounced dynamics compared to the existing discrete RI.

The use of RIs for non-pregnant women results in substantial differences in the interpretation of laboratory results in pregnant women. Continuous gestational age-specific reference curves account for nonlinear physiological changes and provide a physiologically appropriate framework for interpreting laboratory results during pregnancy.