A methylene blue based electrochemical assay to quantify haemoglobin in blood for point-of-care anaemia diagnosis

Anaemia is a blood disorder that affects individuals of all ages, genders, and ethnicities. It results from a decrease in the number or functionality of erythrocytes or haemoglobin (Hb), leading to a deficit in oxygen transport. Anaemia can be classified into different phenotypic groups, such as haemolytic, microcytic, macrocytic, hypochromic, aplastic, and Iron Deficiency Anaemia (IDA). Its diagnosis and classification rely on the measurement of several biomarkers, where Hb is ubiquitous in all of them. Current methods for anaemia diagnosis rely on a complete haemogram, which usually requires larger volumes of sample and hinders its integration into a Point-of-Care (PoC) device. Herein, we developed an analytical method based on the interaction between methylene blue (MB) and Hb on screen printed carbon electrodes (SPCE). The system has been optimized in PBS and subsequently evaluated in whole blood real samples. Different chemical lysing buffers for erythrocytes have been studied, and the erythrocyte's lysing buffer has been chosen as the most suitable for this analytical test. An evaluation of the calibration curves is presented, giving a LOD, LOQ, and sensitivity of 664 nM, 2.8 μM, and 0.22 μM^-1, respectively, allowing the clinically relevant quantification of Hb in only 1 μL of a neat blood sample. This detection method is feasible for the development of a cost-effective multiplexed PoC device with the integration of biosensors for other anaemia biomarkers, providing a fuller view to patients and clinicians of anaemia status and classification.