Ametropia detection using a novel, compact wavefront autorefractor.

INTRODUCTION: Despite the well-known reproducibility issues of subjective refraction, most studies evaluating autorefractors compared differences between the device and subjective refraction. This work evaluated the performance of a novel handheld Hartmann-Shack-based autorefractor using an alternative protocol, which considered the inherent variability of subjective refraction. METHODS: Participants underwent an initial measurement with a desktop autorefractor, two subjective refractions (SR1 and SR2) and a final measurement with the QuickSee Free (QSFree) portable autorefractor. Autorefractor performance was evaluated by comparing the differences between the QSFree and each of the subjective refractions with the difference between the subjective refractions (SR1 vs. SR2) using Bland-Altman analysis and percentage of agreement. RESULTS: A total of 75 subjects (53 ± 14 years) were enrolled in the study. The average difference in the absolute spherical equivalent (M) between the QSFree and the SR1 and SR2 was ±0.24 and ±0.02 D, respectively, that is, very similar or smaller than the SR1 versus SR2 difference (±0.26 D). Average differences in astigmatic components were found to be negligible. The results demonstrate that differences between QSFree and both subjective refractions in J0 and J45 were within ±0.50 D for at least 96% of the measurements. The limits of agreement (LOAs) of the differences between QSFree and SR1, as well as QSFree and SR2, were higher than those observed between SR1 and SR2 for M, J0 and J45 . CONCLUSIONS: A protocol was designed and validated for the evaluation of a refractive device to account for the variability of subjective refraction. This protocol was used to evaluate a novel portable autorefractor and observed a smaller difference between the device and subjective refractions than the difference between the two subjective refraction measurements in terms of mean bias error, although the standard deviation was higher.

Correction: Assesment of the QuickSee wavefront autorefractor for characterizing refractive errors in school-age children.

[This corrects the article DOI: 10.1371/journal.pone.0240933.].

Prediction of manifest refraction using machine learning ensemble models on wavefront aberrometry data.

PURPOSE: To assess the performance of machine learning (ML) ensemble models for predicting patient subjective refraction (SR) using demographic factors, wavefront aberrometry data, and measurement quality related metrics taken with a low-cost portable autorefractor. METHODS: Four ensemble models were evaluated for predicting individual power vectors (M, J0, and J45) corresponding to the eyeglass prescription of each patient. Those models were random forest regressor (RF), gradient boosting regressor (GB), extreme gradient boosting regressor (XGB), and a custom assembly model (ASB) that averages the first three models. Algorithms were trained on a dataset of 1244 samples and the predictive power was evaluated with 518 unseen samples. Variables used for the prediction were age, gender, Zernike coefficients up to 5th order, and pupil related metrics provided by the autorefractor. Agreement with SR was measured using Bland-Altman analysis, overall prediction error, and percentage of agreement between the ML predictions and subjective refractions for different thresholds (0.25 D, 0.5 D). RESULTS: All models considerably outperformed the predictions from the autorefractor, while ASB obtained the best results. The accuracy of the predictions for each individual power vector component was substantially improved resulting in a ± 0.63 D, ±0.14D, and ±0.08 D reduction in the 95% limits of agreement of the error distribution for M, J0, and J45, respectively. The wavefront-aberrometry related variables had the biggest impact on the prediction, while demographic and measurement quality-related features showed a heterogeneous but consistent predictive value. CONCLUSIONS: These results suggest that ML is effective for improving precision in predicting patient's SR from objective measurements taken with a low-cost portable device.

Predicting subjective refraction with dynamic retinal image quality analysis.

The aim of this work is to evaluate the performance of a novel algorithm that combines dynamic wavefront aberrometry data and descriptors of the retinal image quality from objective autorefractor measurements to predict subjective refraction. We conducted a retrospective study of the prediction accuracy and precision of the novel algorithm compared to standard search-based retinal image quality optimization algorithms. Dynamic measurements from 34 adult patients were taken with a handheld wavefront autorefractor and static data was obtained with a high-end desktop wavefront aberrometer. The search-based algorithms did not significantly improve the results of the desktop system, while the dynamic approach was able to simultaneously reduce the standard deviation (up to a 15% for reduction of spherical equivalent power) and the mean bias error of the predictions (up to 80% reduction of spherical equivalent power) for the handheld aberrometer. These results suggest that dynamic retinal image analysis can substantially improve the accuracy and precision of the portable wavefront autorefractor relative to subjective refraction.

Investigation of the Accuracy of a Low-Cost, Portable Autorefractor to Provide Well-Tolerated Eyeglass Prescriptions: A Randomized Crossover Trial.

PURPOSE: To compare patient preferences for eyeglasses prescribed using a low-cost, portable wavefront autorefractor versus standard subjective refraction (SR). DESIGN: Randomized, cross-over clinical trial. PARTICIPANTS: Patients aged 18 to 40 years presenting with refractive errors (REs) to a tertiary eye hospital in Southern India. METHODS: Participants underwent SR followed by autorefraction (AR) using the monocular version of the QuickSee device (PlenOptika Inc). An independent optician, masked to the refraction approach, prepared eyeglasses based on each refraction approach. Participants (masked to refraction source) were randomly assigned to use SR- or AR-based eyeglasses first, followed by the other pair, for 1 week each. At the end of each week, participants had their vision checked and were interviewed about their experience with the eyeglasses. MAIN OUTCOME MEASURES: Patients preferring eyeglasses were chosen using AR and SR. RESULTS: The 400 participants enrolled between March 26, 2018, and August 2, 2019, had a mean (standard deviation) age of 28.4 (6.6) years, and 68.8% were women. There was a strong correlation between spherical equivalents using SR and AR (r = 0.97, P < 0.001) with a mean difference of -0.07 diopters (D) (95% limits of agreement [LoA], -0.68 to 0.83). Of the 301 patients (75.2%) who completed both follow-up visits, 50.5% (n = 152) and 49.5% (n = 149) preferred glasses prescribed using SR and AR, respectively (95% CI, 45.7-56.3; P = 0.86). There were no differences in demographic or vision characteristics between participants with different preferences (P > 0.05 for all). CONCLUSIONS: We observed a strong agreement between the prescriptions from SR and AR, and eyeglasses prescribed using SR and AR were equally preferred by patients. Wider use of prescribing based on AR alone in resource-limited settings is supported by these findings.

Assesment of the QuickSee wavefront autorefractor for characterizing refractive errors in school-age children.

PURPOSE: To assess the performance of an open-view binocular handheld aberrometer (QuickSee) for diagnosing refractive errors in children. METHODS: 123 school-age children (9.9 ± 3.3 years) with moderate refractive error underwent autorefraction (AR) with a standard desktop device and subjective refraction (SR), with or without cycloplegia to determine their eyeglass prescription. Measurements with QuickSee (QS) were taken in 62 of these patients without cycloplegia (NC), and in 61 under cycloplegia (C). Differences in refraction values (AR vs SR vs QS) as well as the visual acuity (VA) achieved by the patients with each method (QS vs SR) were used to evaluate the performance of the device in measuring refractive error. RESULTS: The spherical equivalent refraction obtained by QS agreed within 0.5 D of the SR in 71% (NC) and 70% (C) of the cases. Agreement between the desktop autorefractor and SR for the same threshold was of 61% (NC) and 77% (C). VA resulting from QS refractions was equal to or better than that achieved by SR procedure in 77% (NC) and 74% (C) of the patients. Average improvement in VA with the QS refractions was of 8.6 and 13.4 optotypes for the NC and C groups respectively, while the SR procedure provided average improvements of 8.9 (NC) and 14.8 (C) optotypes. CONCLUSIONS: The high level of agreement between QuickSee and subjective refraction together with the VA improvement achieved in both study groups using QuickSee refractions suggest that the device is a useful autorefraction tool for school-age children.

Validation of an Affordable Handheld Wavefront Autorefractor.

SIGNIFICANCE: There is a critical need for tools that increase the accessibility of eye care to address the most common cause of vision impairment: uncorrected refractive errors. This work assesses the performance of an affordable autorefractor, which could help reduce the burden of this health care problem in low-resource communities. PURPOSE: The purpose of this study was to validate the commercial version of a portable wavefront autorefractor for measuring refractive errors. METHODS: Refraction was performed without cycloplegia using (1) a standard clinical procedure consisting of an objective measurement with a desktop autorefractor followed by subjective refraction (SR) and (2) with the handheld autorefractor. Agreement between both methods was evaluated using Bland-Altman analysis and by comparing the visual acuity (VA) with trial frames set to the resulting measurements. RESULTS: The study was conducted on 54 patients (33.9 ± 14.1 years of age) with a spherical equivalent (M) refraction determined by SR ranging from -7.25 to 4.25 D (mean ± SD, -0.93 ± 1.95 D). Mean differences between the portable autorefractor and SR were 0.09 ± 0.39, -0.06 ± 0.13, and 0.02 ± 0.12 D for M, J0, and J45, respectively. The device agreed within 0.5 D of SR in 87% of the eyes for spherical equivalent power. The average VAs achieved from trial lenses set to the wavefront autorefractor and SR results were 0.02 ± 0.015 and 0.015 ± 0.042 logMAR units, respectively. Visual acuity resulting from correction based on the device was the same as or better than that achieved by SR in 87% of the eyes. CONCLUSIONS: This study found excellent agreement between the measurements obtained with the portable autorefractor and the prescriptions based on SR and only small differences between the VA achieved by either method.

H-EM: An algorithm for simultaneous cell diameter and intensity quantification in low-resolution imaging cytometry.

Fluorescent cytometry refers to the quantification of cell physical properties and surface biomarkers using fluorescently-tagged antibodies. The generally preferred techniques to perform such measurements are flow cytometry, which performs rapid single cell analysis by flowing cells one-by-one through a channel, and microscopy, which eliminates the complexity of the flow channel, offering multi-cell analysis at a lesser throughput. Low-magnification image-based cytometers, also called "cell astronomy" systems, hold promise of simultaneously achieving both instrumental simplicity and high throughput. In this magnification regime, a single cell is mapped to a handful of pixels in the image. While very attractive, this idea has, so far, not been proven to yield quantitative results of cell-labeling, mainly due to the poor signal-to-noise ratio present in those images and to partial volume effects. In this work we present a cell astronomy system that, when coupled with custom-developed algorithms, is able to quantify cell intensities and diameters reliably. We showcase the system using calibrated MESF beads and fluorescently stained leukocytes, achieving good population identification in both cases. The main contribution of the proposed system is in the development of a novel algorithm, H-EM, that enables inter-cluster separation at a very low magnification regime (2x). Such algorithm provides more accurate brightness estimates than DAOSTORM when compared to manual analysis, while fitting cell location, brightness, diameter, and background level concurrently. The algorithm first performs Fisher discriminant analysis to detect bright spots. From each spot an expectation-maximization algorithm is initialized over a heterogeneous mixture model (H-EM), this algorithm recovers both the cell fluorescence and diameter with sub-pixel accuracy while discriminating the background noise. Finally, a recursive splitting procedure is applied to discern individual cells in cell clusters.

Quality of eyeglass prescriptions from a low-cost wavefront autorefractor evaluated in rural India: results of a 708-participant field study.

OBJECTIVE: To assess the quality of eyeglass prescriptions provided by an affordable wavefront autorefractor operated by a minimally trained technician in a low-resource setting. METHODS AND ANALYSIS: 708 participants were recruited from consecutive patients registered for routine eye examinations at Aravind Eye Hospital in Madurai, India, or an affiliated rural satellite vision centre. Visual acuity (VA) and patient preference were compared between trial lenses set to two eyeglass prescriptions from (1) a novel wavefront autorefractor and (2) subjective refraction by an experienced refractionist. RESULTS: The mean±SD VA was 0.30±0.37, -0.02±0.14 and -0.04±0.11 logarithm of the minimum angle of resolution units before correction, with autorefractor correction and with subjective refraction correction, respectively (all differences p<0.01). Overall, 25% of participants had no preference, 33% preferred eyeglass prescriptions from autorefraction, and 42% preferred eyeglass prescriptions from subjective refraction (p<0.01). Of the 438 patients 40 years old and younger, 96 had no preference and the remainder had no statistically significant difference in preference for subjective refraction prescriptions (51%) versus autorefractor prescriptions (49%) (p=0.52). CONCLUSION: Average VAs from autorefractor-prescribed eyeglasses were one letter worse than those from subjective refraction. More than half of all participants either had no preference or preferred eyeglasses prescribed by the autorefractor. This marginal difference in quality may warrant autorefractor-based prescriptions, given the portable form factor, short measurement time, low cost and minimal training required to use the autorefractor evaluated here.

Direct Characterization of Polymer Encapsulated CdSe/CdS/ZnS Quantum Dots.

Surface engineering advances of semiconductor quantum dots (QDs) have enabled their application to molecular labeling, disease diagnostics and tumor imaging. For biological applications, hydrophobic core/shell QDs are transferred into aqueous solutions through the incorporation of water-solubility imparting moieties, typically achieved via direct exchange of the native surface passivating ligands or indirectly through the adsorption of polymers. Although polymeric encapsulation has gained wide acceptance, there are few reports addressing the characterization of the adsorbed polymers and existing theoretical analyses are typically based on simple geometric models. In this work, we experimentally characterize and quantify water-soluble QDs prepared by adsorption of amphiphilic poly(maleic anhydride-alt-1-tetradecene) (PMAT, MW~9000) onto commercially available CdSe/CdS/ZnS (CdSe/CdS/ZnS-PMAT). Using x-ray photoelectron spectroscopy (XPS) we determined that ~15 PMAT molecules are adsorbed onto each QD and sum frequency generation (SFG) vibrational spectra was utilized to investigate the mechanism of interaction between PMAT molecules and the QD surface. Importantly, when employed together, these techniques constitute a platform with which to investigate any polymer-nanoparticle complex in general.

Design and Clinical Evaluation of a Handheld Wavefront Autorefractor.

PURPOSE: To introduce a novel autorefractor design that is intended to be manufacturable at low cost and evaluate its performance in measuring refractive errors. METHODS: We developed a handheld, open-view autorefractor (the "QuickSee" [QS]) that uses a simplified approach to wavefront sensing that forgoes moving parts and expensive components. Adult subjects (n = 41) were recruited to undergo noncycloplegic refraction with three methods: (1) a QS prototype, (2) a Grand Seiko WR-5100K (GS) autorefractor, and (3) subjective refraction (SR). Agreements between the QS and GS were evaluated using a Bland-Altman analysis. The accuracy of both autorefractors was evaluated using SR as the clinical gold standard. RESULTS: The spherical equivalent powers measured from both autorefractors correlate well with SR, with identical correlation coefficients of r = 0.97. Both autorefractors also agree well with each other, with a spherical equivalent power 95% confidence interval of ±0.84 diopters (D). The difference between the accuracy of each objective device is not statistically significant for any component of the power vector (p = 0.55, 0.41, and 0.18, for M, J0, and J45, respectively). The spherical and cylindrical powers measured by the GS agree within 0.25 D of the SR in 49 and 82% of the eyes, respectively, whereas the spherical and cylindrical powers measured by the QS agree within 0.25 D of the SR in 74 and 87% of the eyes, respectively. CONCLUSIONS: The prototype autorefractor exhibits equivalent performance to the GS autorefractor in matching power vectors measured by SR.

Recovery and normalization of triple coincidences in PET.

PURPOSE: Triple coincidences in positron emission tomography (PET) are events in which three γ-rays are detected simultaneously. These events, though potentially useful for enhancing the sensitivity of PET scanners, are discarded or processed without special consideration in current systems, because there is not a clear criterion for assigning them to a unique line-of-response (LOR). Methods proposed for recovering such events usually rely on the use of highly specialized detection systems, hampering general adoption, and/or are based on Compton-scatter kinematics and, consequently, are limited in accuracy by the energy resolution of standard PET detectors. In this work, the authors propose a simple and general solution for recovering triple coincidences, which does not require specialized detectors or additional energy resolution requirements. METHODS: To recover triple coincidences, the authors' method distributes such events among their possible LORs using the relative proportions of double coincidences in these LORs. The authors show analytically that this assignment scheme represents the maximum-likelihood solution for the triple-coincidence distribution problem. The PET component of a preclinical PET/CT scanner was adapted to enable the acquisition and processing of triple coincidences. Since the efficiencies for detecting double and triple events were found to be different throughout the scanner field-of-view, a normalization procedure specific for triple coincidences was also developed. The effect of including triple coincidences using their method was compared against the cases of equally weighting the triples among their possible LORs and discarding all the triple events. The authors used as figures of merit for this comparison sensitivity, noise-equivalent count (NEC) rates and image quality calculated as described in the NEMA NU-4 protocol for the assessment of preclinical PET scanners. RESULTS: The addition of triple-coincidence events with the authors' method increased peak NEC rates of the scanner by 26.6% and 32% for mouse- and rat-sized objects, respectively. This increase in NEC-rate performance was also reflected in the image-quality metrics. Images reconstructed using double and triple coincidences recovered using their method had better signal-to-noise ratio than those obtained using only double coincidences, while preserving spatial resolution and contrast. Distribution of triple coincidences using an equal-weighting scheme increased apparent system sensitivity but degraded image quality. The performance boost provided by the inclusion of triple coincidences using their method allowed to reduce the acquisition time of standard imaging procedures by up to ∼25%. CONCLUSIONS: Recovering triple coincidences with the proposed method can effectively increase the sensitivity of current clinical and preclinical PET systems without compromising other parameters like spatial resolution or contrast.

From unseen to seen: tackling the global burden of uncorrected refractive errors.

Worldwide, more than one billion people suffer from poor vision because they do not have the eyeglasses they need. Their uncorrected refractive errors are a major cause of global disability and drastically reduce productivity, educational opportunities, and overall quality of life. The problem persists most prevalently in low-resource settings, even though prescription eyeglasses serve as a simple, effective, and largely affordable solution. In this review, we discuss barriers to obtaining, and approaches for providing, refractive eye care. We also highlight emerging technologies that are being developed to increase the accessibility of eye care. Finally, we describe opportunities that exist for engineers to develop new solutions to positively impact the diagnosis and treatment of correctable refractive errors in low-resource settings.

Addressing Key Technical Aspects of Quantum Dot Probe Preparation for Bioassays.

Fluorescent semiconductor nanoparticles, or quantum dots, have become a promising platform for the engineering of biofunctional probes for a variety of biomedical applications, ranging from multicolor imaging to single-molecule tracking to traceable drug delivery. Advances in organometallic synthesis have enabled preparation of hydrophobic quantum dots with high quantum yields and narrow size distribution, offering bright optical materials with narrow size-tunable emission profiles. At the same time, polymer encapsulation procedures provided a simple and versatile methodology for transferring hydrophobic nanoparticles into physiologically-relevant aqueous buffers. Taken together, hydrophobic nanoparticle platforms and polymer encapsulation should offer great flexibility for implementation of novel probe designs. However, the success of the encapsulation and purification depends on many factors often overlooked in the scientific literature, such as close match between nanoparticle and polymer physicochemical properties and dimensions, slow dynamics of polymer arrangement on the nanoparticle surface, and the size and charge similarity of resultant polymer-coated quantum dots and empty byproduct polymer micelles. To make this general hydrophobic nanoparticle modification strategy accessible by a broad range of biomedical research groups, we focus on the important technical aspects of nanoparticle polymer encapsulation, purification, bioconjugation, and characterization.

Luminescent quantum dots for molecular toxicology.

Recent developments in nanotechnology have made available a host of new approaches for the improved quantitative detection of biomarkers due to the enhanced sensitivity of nanoparticle-based assays. The majority of molecular toxicology studies revolve around sensitive measurement of cell-death (apoptosis) and cell-health biomarkers present in living cells or formalin-fixed and paraffin embedded (FFPE) tissue samples. In this regard, semi-conductor quantum dots (QDs) which exhibit high brightness, photo-stability and degree of multiplexing, are predicted to have a significant impact on research in molecular toxicology. Due to these superior photophysical properties of QDs as compared to traditional fluorophores and the unsurpassed versatility of QDs as enabling components for new assays, these nanoparticles promise to facilitate new discoveries in molecular toxicology. Indeed, multiplexed QD-based assays have been incorporated into cell imaging, flow cytometry and other homogenized sample-based assays for detecting multiple biomarkers including those associated with cell injury and apoptosis.

Method for determining the elemental composition and distribution in semiconductor core-shell quantum dots.

In the biological sciences, the use of core-shell quantum dots (QDs) has gained wide usage but analytical challenges still exist for characterizing the QD structure. The application of energy-dispersive X-ray spectroscopy and X-ray photoelectron spectroscopy (XPS) to bulk materials is relatively straightforward; however, for meaningful applications of surface science techniques to multilayer nanoparticles requires novel modifications and analysis methods. To experimentally characterize the elemental composition and distribution in CdSe/CdS/ZnS QDs, we first develop a XPS signal subtraction technique capable of separating the overlapped selenium 3s (core) and sulfur 2s (shell) peaks (both peaks have binding energies near 230 eV) with higher precision than is typically reported in the nanoparticle literature. This method is valid for any nanoparticle containing selenium and sulfur. Then we apply a correction formula to the XPS data and determine that the 2 nm stoichiometric CdSe core is surrounded by 2 CdS layers and a stoichimetric ZnS monolayer. These findings and the multiapproach methodology represent a significant advancement in the detailed surface science study of multilayer nanoparticles. In agreement with recent surprising findings, the time-of-flight secondary mass spectrometry measurements suggest that the surface sites of the QDs used in this study are primarily covered with a mixture of octadecylphosphonic acid and trioctylphophine oxide.

Monodisperse magnetic nanoparticles for biodetection, imaging, and drug delivery: a versatile and evolving technology.

Advances in nanotechnology have pushed forward the synthesis of a variety of functional nanoparticles (NPs) such as semiconductor quantum dots (QDs), magnetic and metallic NPs. The unique electronic, magnetic, and optical properties exhibited by these nanometer-sized materials have enabled a broad spectrum of biomedical applications. In particular, iron-oxide-based magnetic NPs have proved to be highly versatile deep-tissue imaging agents, having been incorporated into clinical applications due to their biocompatibility. This Interdisciplinary Review will focus on the recent advances in strategies for the synthesis and surface modification of highly monodisperse magnetic NPs and their use in imaging, drug delivery, and innovative ultrasensitive bioassays.

Quantum dot nanobarcodes: epitaxial assembly of nanoparticle-polymer complexes in homogeneous solution.

Multiplexed nanobarcodes have been prepared with quantum dots (QDs) and alternating amphiphilic copolymers consisting of hydrocarbons and maleic anhydride groups. In homogeneous solution, the QD-polymer complexes grow epitaxially into nanobeads of narrow size dispersity, which has been previously achieved only for micrometer-sized beads in the presence of solid supports. As a result of this new nanostructure formation mechanism, more than 250 QDs can be loaded into a nanobead of 100 nm in diameter. A model assay for sensitive detection of human prostate specific antigen has also been demonstrated using the QD-nanobeads as fluorescent reporters. This nanoparticle-polymer self-assembly technology is capable of producing a variety of nanostructures and is expected to open new opportunities in nanoparticle-based ultrasensitive detection and imaging.

Ultrasensitive detection and molecular imaging with magnetic nanoparticles.

Recent advances in nanotechnology have produced a variety of nanoparticles ranging from semiconductor quantum dots (QDs), magnetic nanoparticles (MNPs), metallic nanoparticles, to polymeric nanoparticles. Their unique electronic, magnetic, and optical properties have enabled a broad spectrum of biomedical applications such as ultrasensitive detection, medical imaging, and specific therapeutics. MNPs made from iron oxide, in particular, have attracted extensive interest and have already been used in clinical studies owing to their capability of deep-tissue imaging, non-immunogenesis, and low toxicity. In this Research Highlight article, we attempt to highlight the recent breakthroughs in MNP synthesis based on a non-hydrolytic approach, nanoparticle (NP) surface engineering, their unique structural and magnetic properties, and current applications in ultrasensitive detection and imaging with a special focus on innovative bioassays. We will also discuss our perspectives on future research directions.

Quantum dots for cancer molecular imaging.

Quantum dots (QDs), tiny light-emitting particles on the nanometer scale, are emerging as a new class of fluorescent probes for biomolecular and cellular imaging. In comparison with organic dyes and fluorescent proteins, quantum dots have unique optical and electronic properties such as size-tunable light emission, improved signal brightness, resistance against photobleaching, and simultaneous excitation of multiple fluorescence colors. These properties are most promising for improving the sensitivity of molecular imaging and quantitative cellular analysis by 1-2 orders of magnitude. Recent advances have led to multifunctional nanoparticle probes that are highly bright and stable under complex in-vivo conditions. A new structural design involves encapsulating luminescent QDs with amphiphilic block copolymers, and linking the polymer coating to tumor-targeting ligands and drug-delivery functionalities. Polymer-encapsulated QDs are essentially nontoxic to cells and small animals, but their long-term in-vivo toxicity and degradation need more careful studies. Nonetheless, bioconjugated QDs have raised new possibilities for ultrasensitive and multiplexed imaging of molecular targets in living cells and animal models.