Analysis of haemolysis-based injury in immortalised proximal tubule epithelial cells is impaired by haemoglobin interference with bioluminescence assays.

Free haemoglobin (fHb) interference substantially reduces bioluminescence readouts in human ciPTECs, and removing fHb or haemolysed plasma before assays considerably mitigates this signal loss.

• Why it matters: Bioluminescence-based injury measurements in haemolysis-linked AKI models can be distorted, yet the injury mechanisms and assay reliability under fHb exposure remain unclear.
• What they did: The study exposed human-derived conditionally immortalised proximal tubule epithelial cells (ciPTECs) to fHb solutions and haemolysed plasma, then assessed bioluminescence, NF-B activity, and mitochondrial function.
• The result: fHb reduced bioluminescence signal output, with different effects between haemolysed plasma and fHb solutions; haemolysed plasma caused only a marginal NF-B increase and minimal mitochondrial impact, implying assay caution and limited injury severity in ciPTECs.

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