Hemozoin and hemoglobin characterization by optical absorption towards a miniaturized spectrophotometric malaria diagnostic system

Malaria is a parasitic disease with high prevalence in regions with inefficient healthcare conditions. Currently, the most used diagnostic methods in endemic regions are microscopy and rapid diagnostic tests by immunochromatographic assays. This work proposes an alternative miniaturized optical diagnostic method by spectrophotometry, for hemozoin (Hz) and hemoglobin (Hb) quantification and classification. Hz is a subproduct of the malaria parasite, produced by degradation of Hb, inside the red blood cells and, as the malaria parasite is proliferating, the Hz concentration increases and the Hb decreases. Hz and Hb have typical absorption spectra, with characteristic absorption peaks, that change their shape according to their concentrations. In this work, it was studied the optical absorbance of Hz and whole blood (WB) samples, with different concentrations of both Hz and Hb. Additionally, with the aim of miniaturization, different solution volumes and different optical paths were considered. The miniaturization capacity was tested using 1 mm optical path quartz cuvettes and 3 mm optical path polydimethylsiloxane wells, where WB samples with different Hz concentration were introduced. The results showed high linearity of the absorbance curve, as a function of the concentration, with R-2 between 0.99 and 0.97, for volumes of 350 mu L with 1 mm optical path and 21.2 mu L with 3 mm optical path, respectively, and proved the accuracy of the method. Overall, this work presented the spectrophotometric basis and proof of concept towards the development of a lab-on-a-chip device for malaria detection in WB.Work supported by FCT with reference project UID/EEA/04436/2013, by FEDER funds through the COMPETE 2020 – Programa Operacional Competitividade e Internacionalização (POCI) with the reference project POCI-01-0145-FEDER-006941. SO Catarino thanks FCT for the SFRH/BPD/108889/2015 grant, supported by national funds from Ministérios da Ciência, Tecnologia e Ensino Superior and by FSE through the POCH - Programa Operacional Capital Humano. The authors thank Diana Pinho from the IP Bragança for providing the blood samples.