Lipid-Specific Labeling of Enveloped Viruses with Quantum Dots for Single-Virus Tracking

Abstract

Quantum dots (QDs) possess optical properties of superbright fluorescence, excellent photostability, narrow emission spectra, and optional colors. Labeled with QDs, single molecules/viruses can be rapidly and continuously imaged for a long time, providing more detailed information than when labeled with other fluorophores. While they are widely used to label proteins in single-molecule-tracking studies, QDs have rarely been used to study virus infection, mainly due to a lack of accepted labeling strategies. Here, we report a general method to mildly and readily label enveloped viruses with QDs. Lipid-biotin conjugates were used to recognize and mark viral lipid membranes, and streptavidin-QD conjugates were used to light them up. Such a method allowed enveloped viruses to be labeled in 2 h with specificity and efficiency up to 99% and 98%, respectively. The intact morphology and the native infectivity of viruses were preserved. With the aid of this QD labeling method, we lit wild-type and mutant Japanese encephalitis viruses up, tracked their infection in living Vero cells, and found that H144A and Q258A substitutions in the envelope protein did not affect the virus intracellular trafficking. The lipid-specific QD labeling method described in this study provides a handy and practical tool to readily “see” the viruses and follow their infection, facilitating the widespread use of single-virus tracking and the uncovering of complex infection mechanisms. IMPORTANCE Virus infection in host cells is a complex process comprising a large number of dynamic molecular events. Single-virus tracking is a versatile technique to study these events. To perform this technique, viruses must be fluorescently labeled to be visible to fluorescence microscopes. The quantum dot is a kind of fluorescent tag that has many unique optical properties. It has been widely used to label proteins in single-molecule-tracking studies but rarely used to study virus infection, mainly due to the lack of an accepted labeling method. In this study, we developed a lipid-specific method to readily, mildly, specifically, and efficiently label enveloped viruses with quantum dots by recognizing viral envelope lipids with lipid-biotin conjugates and recognizing these lipid-biotin conjugates with streptavidin-quantum dot conjugates. It is not only applicable to normal viruses, but also competent to label the key protein-mutated viruses and the inactivated highly virulent viruses, providing a powerful tool for single-virus tracking.

State-of-the-Art of Nanodiagnostics and Nanotherapeutics against SARS-CoV-2

“…Previously, QD-labeling of the enveloped viruses has been proved as a practical approach to investigate the cell internalization mechanisms. 127 Furthermore, QDs could be used in developing a variety of fluorescence (Förster) resonance energy transfer (FRET)-based sensors. 128 In such systems, the energy of excited electrons in donor QDs could be transferred to an acceptor fluorescent molecule or particle.…”

Emerging Advances of Nanotechnology in Drug and Vaccine Delivery against Viral Associated Respiratory Infectious Diseases (VARID)

“…Researchers also developed anionic and cationic polyamidoamine with PAMAM on MERS-COV infectious, with their intervention showing improved antiviral responses [130]. KK-46 dendrimer is a peptide-based compound used for intracellular delivery of anti-SARS-CoV-2 siRNA for inhibition of virus replication [131]. Finally, astrodimer sodium is a four-lysine dendrimer with a poly anionic charge that has been shown to inhibit viral infections in VeroE6 cells and reduce the replication of virus [132].…”

Aerobic and Anaerobic Bacterial and Fungal Degradation of Pyrene: Mechanism Pathway Including Biochemical Reaction and Catabolic Genes

“…M. vanbaalenii PYR-1 metabolizes pyrene in two paths: the first path consists of deoxygenating pyrene via C1 and C2 leading to the formation of o-methylated derived to the pyrene 1-2-diol, while the second one is related to the deoxygenation at C4 and C5 of the K-region, resulting in the complete degradation. The deoxygenation of pyrene in C4 and C5 is catalyzed by the ring hydroxylating oxygenase α-subunit (A) and β-subunit (B) encoded by nidAB [ 44 ]. However, the deoxygenation of pyrene in C1 and C2 to form 1-2-dihydroxypyrene is encoded by nidA3B3 [ 44 ].…”