Urinary carbonic anhydrase 1 excretion is a marker of hemolysis-triggering conditions suitable for point-of-care testing Open Access

1. Urinary carbonic anhydrase 1 excretion detects intravascular hemolysis across diverse conditions.

2. A lateral flow device testing urine for carbonic anhydrase 1 enables rapid detection of hemolysis in resource-limited settings.

Intravascular hemolysis accompanies diverse diseases, yet blood-based markers (plasma-free hemoglobin, haptoglobin, lactate dehydrogenase, bilirubin) are variably sensitive, non-specific, or impractical for point-of-care use. We evaluated urinary carbonic anhydrase 1 (CA1) as a mechanistically grounded, urine-based marker of hemolysis in a multi-center study spanning the UK, Bangladesh, and Peru. We enrolled 234 participants: healthy adults and adults with inherited anemias (Oxford), newborns in intensive care (London), children and adults with complicated or uncomplicated malaria (Bangladesh), and adults attending rural clinics for various medical reasons (Peru). Urine CA1 and hemoglobin (Hb) were quantified by ELISA and immunoblot, with CA1:Hb stoichiometry used to distinguish intravascular hemolysis from urogenital blood contamination. Correlations with PFH, LDH, bilirubin, Hb, CRP, and clinical variables were tested using regression, principal component analysis, and decision tree. Reference urines were predominantly CA1-negative/Hb-negative. In inherited anemias, urinary CA1 was highest in sickle cell disease and correlated with serum LDH and inversely with blood Hb. In neonates, longitudinal CA1 trajectories stratified infants into physiological, transient, and sustained hemolysis groups; higher CA1 associated with prematurity, elevated CRP, and lower WBC. Urinary CA1 was detected in Bangladeshi patients with elevated plasma-free hemoglobin (PFH) and in cases of intravascular hemolysis not identified by PFH. Among Peruvian participants, urinary CA1 correlated with raised C-reactive protein (CRP) consistent with inflammation being a pro-hemolytic trigger. To enable point-of-care testing, a lateral flow device was developed and verified for excellent sensitivity and specificity. Urinary CA1 provides a sensitive and practical readout of intravascular hemolysis suitable for point-of-care testing globally.