CdTe quantum dots conjugated to concanavalin A as potential fluorescent molecular probes for saccharides detection in Candida albicans.

Semiconductor colloidal quantum dots (QDs) have been applied in biological analysis due to their unique optical properties and their versatility to be conjugated to biomolecules, such as lectins and antibodies, acquiring specificity to label a variety of targets. Concanavalin A (Con A) lectin binds specifically to α-d-mannose and α-d-glucose regions of saccharides that are usually expressed on membranes of mammalian cells and on cell walls of microbials. Candida albicans is the most common fungal opportunistic pathogen present in humans. Therefore, in this work, this fungus was chosen as a model for understanding cells and biofilm-forming organisms. Here, we report an efficient bioconjugation process to bind CdTe (Cadmium Telluride) QDs to Con A, and applied the bioconjugates to label saccharide structures on the cellular surface of C. albicans suspensions and biofilms. By accomplishing hemagglutination experiments and circular dichroism, we observed that the Con A structure and biochemical properties were preserved after the bioconjugation. Fluorescence microscopy images of yeasts and hyphae cells, as well as biofilms, incubated with QDs-(Con A) showed a bright orange fluorescence profile, indicating that the cell walls were specifically labeled. Furthermore, flow cytometry measurements confirmed that over 93% of the yeast cells were successfully labeled by QD-(Con A) complex. In contrast, non-conjugated QDs or QDs-(inhibited Con A) do not label any kind of biological system tested, indicating that the bioconjugation was specific and efficient. The staining pattern of the cells and biofilms demonstrate that QDs were effectively bioconjugated to Con A with specific labeling of saccharide-rich structures on C. albicans. Consequently, this work opens new possibilities to monitor glucose and mannose molecules through fluorescence techniques, which can help to optimize phototherapy protocols for this kind of fungus.

Fluorescence plate reader for quantum dot-protein bioconjugation analysis.

We present here a new and alternative method that uses a Fluorescence Plate Reader in a different approach, not to study protein-protein interactions, but to evaluate the efficiency of the protein bioconjugation to quantum dots (QDs). The method is based on the QDs' native fluorescence and was successfully tested by employing two different QDs-proteins conjugation methodologies, one by promoting covalent binding and other by inducing adsorption processes. For testing, we used bioconjugates between carboxyl coated CdTe QDs and bovine serum albumin, concanavalin A lectin and anti-A antibody. Flow cytometry and fluorescence spectroscopy studies corroborated the results found by the Fluorescence Plate Reader assay. This kind of analysis is important because poor bioconjugation efficiency leads to unsuccessful applications of the fluorescent bioconjugates. We believe that our method presents the possibility of performing semi-quantitative and simultaneous analysis of different samples with accuracy taking the advantage of the high sensitivity of optical based measurements.