Mannose-binding lectin conjugated to quantum dots as fluorescent nanotools for carbohydrate tracing.

Quantum dots (QDs) have stood out as nanotools for glycobiology due to their photostability and ability to be combined with lectins. Mannose-binding lectin (MBL) is involved in the innate immune system and plays important roles in the activation of the complement cascade, opsonization, and elimination of apoptotic and microbial cells. Herein, adsorption and covalent coupling strategies were evaluated to conjugate QDs to a recombinant human MBL (rhMBL). The most efficient nanoprobe was selected by evaluating the conjugate ability to labelCandida albicansyeasts by flow cytometry. The QDs-rhMBL conjugate obtained by adsorption at pH 6.0 was the most efficient, labelingca.100% of cells with the highest median fluorescence intensity. The conjugation was also supported by Fourier transform infrared spectroscopy, zeta potential, and size analyses.C. albicanslabeling was calcium-dependent; 12% and <1% of cells were labeled in buffers without calcium and containing EDTA, respectively. The conjugate promoted specific labeling (based on cluster effect) since, after inhibition with mannan, there was a reduction of 80% in cell labeling, which did not occur with methyl-α-D-mannopyranoside monosaccharide. Conjugates maintained colloidal stability, bright fluorescence, and biological activity for at least 8 months. Therefore, QDs-rhMBL conjugates are promising nanotools to elucidate the roles of MBL in biological processes.

Evaluating viscoelastic properties and membrane electrical charges of red blood cells with optical tweezers and cationic quantum dots - applications to ß-thalassemia intermedia hemoglobinopathy.

Biomechanical and electrical properties are important to the performance and survival of red blood cells (RBCs) in the microcirculation. This study proposed and explored methodologies based on optical tweezers and cationic quantum dots (QDs) as biophotonic tools to characterize, in a complementary way, viscoelastic properties and membrane electrical charges of RBCs. The methodologies were applied to normal (HbA) and ß-thalassemia intermedia (Hbß) RBCs. The ß-thalassemia intermedia disease is a hereditary hemoglobinopathy characterized by a reduction (or absence) of ß-globin chains, which leads to α-globin chains precipitation. The apparent elasticity (µ) and membrane viscosity (ηm) of RBCs captured by optical tweezers were obtained in just a single experiment. Besides, the membrane electrical charges were evaluated by flow cytometry, exploring electrostatic interactions between cationic QDs, stabilized with cysteamine, with the negatively charged RBC surfaces. Results showed that Hbß RBCs are less elastic, have a higher ηm, and presented a reduction in membrane electrical charges, when compared to HbA RBCs. Moreover, the methodologies based on optical tweezers and QDs, here proposed, showed to be capable of providing a deeper and integrated comprehension on RBC rheological and electrical changes, resulting from diverse biological conditions, such as the ß-thalassemia intermedia hemoglobinopathy.

Interactions of mannose binding-lectin with red blood cells by employing cationic quantum dots.

Mannose-binding lectin (MBL) plays important roles by interacting with specific molecular patterns on cell surfaces, triggering first-line host defense. We investigated the MBL interaction with healthy red blood cell membranes as well as its effects on the membrane rheology. We explored electrostatic interactions between cationic quantum dots (QDs) and negatively charged red blood cell surfaces to quantitatively evaluate membrane electrical charges as well as to investigate the MBL binding to healthy erythrocytes. Results showed that cationic QDs labeled efficiently red blood cells. However, the MBL interaction with erythrocytes prevents the QD labeling. We also observed that red blood cells treated with MBL are more resistant to lysis, suggesting a membrane-stabilizing effect. Moreover, we used a fluorescent anti-MBL antibody and Candida albicans cells to further study the MBL interaction with erythrocytes. Our results of this comparative labeling suggested that either this probe was not effective to detect MBL bound to healthy red blood cells (by its carbohydrate-recognition domain) or the MBL binding to those cells might be occurring via another portion. Thus, our results demonstrated the ability of MBL interacting with healthy red blood cells and pointed out to a new role of this protein as a membrane-stabilizing molecule.