Facile and Rapid One‐Step Mass Preparation of Quantum‐Dot Barcodes

Abstract

Since their creation in 1949 by Woodland and Silver for grocery and warehouse inventory, [1] the broad application and utility of barcodes has continuously expanded. Today, in response to the demand for high-throughput multiplexed detection for elucidation of biomolecular mechanisms and to advancing personalized molecular diagnostics and therapeutics, molecular barcodes are much needed for inventorying biological molecules. [2][3][4] Within the context of molecular barcoding, two platforms capable of fulfilling the needs for code recognition and multiplexed detection exist: graphical barcodes utilizing structural recognition and spectroscopic barcodes using the unique optical properties of an embedded material. [2] Whereas each platform maintains the potential to construct large barcode libraries, limitations in barcode stability, reproducibility, or readout impose serious limitations. Graphical barcodes, [5][6][7][8][9][10][11] such as etched polymeric or striped metallic structures, suffer a multitude of problems from the complex instrumentation required for both synthesis and readout, the slow data collection rate (approximately 3 Hz for recent polymeric structures [11] compared to several kilohertz detection of microbeads by flow cytometry), to the unstable dispersion of these structures in buffer or media, making them unfit for mainstream applications. While spectroscopic barcodes employ a colorimetric readout, fluorescence-based barcodes are rapidly detected using flow cytometry [12,13] or miniaturized home-built systems.[14] Raman barcodes still require planar array readout after lengthy detection protocols.[15] From a time-efficiency and sensitivity standpoint, fluorescence-based microbead barcodes are therefore best suited for applications in high-throughput multiplexed detection.Fluorescent quantum dots (QDs) boast narrow Gaussian emission line shapes, resistance to photobleaching, high quantum yields, and single-wavelength excitation, whereas molecular dyes suffer from both photobleaching and redtailed emission; [16,17] thus, QDs are ideal fluorophores for barcoding. However, the construction of QD barcodes has shown minimal advance since their conception in 2001 [18] because of difficulties encountered in the mass production of robust and reproducible barcoded materials. Current methods to prepare QD barcodes include the "swelling" technique, [18] QD entrapment inside layer-by-layer charged polymer coatings [19] or mesoporous silica microbeads, [20] and polymerizable QD encapsulation. [21,22] Microbead "swelling" and layer-by-layer techniques result only in surface-level loading of QDs into the polymer, [18,23] which are thus exposed to pH values and environmental factors that destabilize their fluorescence intensity. [24,25] Stability of the barcode (fluorescence profile) requires that the QDs be positioned well within the polymer matrix and do not leak from the bead, and thus, techniques to encapsulate the QDs during the polymerization step were developed. [21,22] However, this process is lengthy, r...

Droplet microfluidic technology for single-cell high-throughput screening

“…Multiplexing this coding scheme for 2 different colors will bring the range of accessible codes to 10,000 optical codes. A number of alternative coding strategies can be envisioned, including fluorescent techniques based on microbeads (30,31) or coded particles (32) that would permit both real-time deconvolution and postprocessing data analysis, positional encoding (18), or the use of DNA (33,34) or peptide molecules for coding, which would alleviate the challenges of spectral availability but would require a sorting step and postprocessing data analysis.…”

Diverse Applications of Nanomedicine

“…There are several methods to prepare QD barcodes: (a) the “swelling” technique, where polystyrene beads are mixed in a solvent to enable beads to increase in volume, which allows the diffusion of hydrophobic QDs into the outermost layer, 140 (b) layer-by-layer (LbL) assembly of different emitting QDs onto the surface of the beads, 141 (c) incorporation of QDs inside a silica NP with a hydrophobic core, 142,143 and (d) microfluidic flow focusing to incorporate QDs inside the microbeads. 144−146 Each method of barcode preparation has a different fluorescence signal stability. Some formulations tend to expose QDs to small ions in the buffer, which leads to high read-out variability.…”

Point of Care Technologies for HIV

“…However fluorescence is used for the broadest range of different types of POC assays [119] for reasons of sensitivity and, more recently, the ready availability of a range of different colors of efficient fluorophores, including quantum dots, quantum-dot barcodes, and fluorescent nanoparticles, providing improved limits of detection. Fluorescence in some cases can detect single particle LODs enabling multitarget multiplexing [120]. …”