Triple-Junction Optoelectronic Sensor with Nanophotonic Layer

1 day ago - Interest in developing a rapid and robust DNA sequencing platform has surged over the past decade. Various next-/third-generation ...
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Triple-Junction Optoelectronic Sensor with Nanophotonic Layer Integration for Single Molecule Level Decoding Hsin-Yi Hsieh, Yu-Hsuan Peng, Sheng-Fu Lin, Li-Ching Chen, Teng-Chien Yu, Chung-Fan Chiou, and Johnsee Lee ACS Nano, Just Accepted Manuscript • DOI: 10.1021/acsnano.9b00019 • Publication Date (Web): 11 Mar 2019 Downloaded from http://pubs.acs.org on March 11, 2019

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Triple-Junction Optoelectronic Sensor with Nanophotonic Layer Integration for Single Molecule Level Decoding Hsin-Yi Hsieh*, Yu-Hsuan Peng, Sheng-Fu Lin, Li-Ching Chen, Teng-Chien Yu, Chung-Fan Chiou, Johnsee Lee Personal Genomics, Inc., Hsinchu Biomedical Science Park, Zhubei, Hsinchu 30261, Taiwan. *email: [email protected] Abstract Interest in developing a rapid and robust DNA sequencing platform has surged over the past decade. Various next-/third-generation sequencing mechanisms have been employed to replace the traditional Sanger sequencing method. In sequencing by synthesis (SBS), a signal is monitored by a scanning charge-coupled device (CCD) to identify thousands to millions of incorporated dNTPs with distinctive fluorophores on a chip. Because one reaction site usually occupies dozens of pixels on a CCD detector, a bottleneck related to the bandwidth of CCD imaging limits the throughputs of the sequencing performance and causes tradeoffs among speed, accuracy, read length, and the numbers of reaction sites in parallel. Thus, current research aims to align one reaction site to a few pixels by directly stacking nanophotonic layers (NPLs) onto a CMOS detector to minimize the size of the sequencing platforms and accelerate the processing procedures. This article reports a custom integrated optoelectronic device based on a triple-junction photodiode (TPD) CMOS sensor in conjunction with NPL integration for real-time illumination and detection of fluorescent molecules. Keywords: CMOS, filter, grating, nanophotonic layer, planar waveguide, triple-junction photodiode (TPD) Sequencing technology is improving rapidly to advance processing quantity and efficiency based on the massive parallelization of individual DNA sequencing reaction centers on a chip.1-3 Sequencing strategies include base-by-base 5’-to-3’ polymerase extension (sequencing by synthesis (SBS) or sequencing by polymerase), such as those commercialized by 454 Roche, Helicos, Illumina, Ion Torrent, Pacific Bioscience (PacBio), and Direct Genomics, and query-based 3’-to-5’ ligase enzyme extension (sequencing by ligation), such as those commercialized by AB SOLiD and Complete ACS Paragon Plus Environment

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Genomics.3-5 Currently, the overwhelming majority of strategies are based on the SBS process, and representative companies commercializing these strategies include Illumina (based on ensemble SBS for stepwise and short-read sequencing) and PacBio (based on single-molecule SBS for real-time and long-read sequencing).3,6 Illumina amplifies many thousands of identical copies of a DNA fragment together on defined surfaces as DNA clusters to ensure that the fluorescence signals of a cluster can be distinguished from each other. Massive clusters with their own clonal DNA template are added on complementary dNTPs in a cycle of reversible termination, and detection is performed by a charge-coupled device (CCD) to identify the fluorophore attached to the dNTPs. Once the collection of fluorescence signals is completed, the 3’ end blockade is removed to allow a repeating cycle of DNA synthesis. The iterative sequencing process in a cluster tends to lose synchrony after an average of ~150 base pairs.3,4,7 PacBio immobilizes DNA polymerase, clamps different long DNA fragments in the bottom of a zero-mode waveguide (ZMW) nanowell array, and monitors fluorescence-labeled dNTP uptake by the polymerase for DNA sequencing. The diameter of the nanowell bottom and the ZMW confines many thousands of individual picoliter regions in which only the incorporated fluorescence-labeled dNTPs can generate momentary pauses that can be visualized by continuous laser illumination and camera imaging. Owing to the ZMW nanowell with a stationary enzyme, sequencing can be performed in real time at 2-4 bp/sec. The average read length can reach 10-15 kb, which is ideal for de novo genome sequencing applications.7-10 The advantages and disadvantages of the abovementioned technology are complementary between Illumina and PacBio. Due to the combined clonal DNA templates, the raw read accuracy can reach >99.5% for Illumina. However, the properties of a short read generate a bias associated with ambiguous matching to a specific genomic location because of gene redundancy, with the occurrence of multiple subsequences in the genome performing the same function.11,12 Although PacBio can promise long-read sequencing, the very high error rate of 15% for single-read sequencing raises concerns about instrument usage.2,7

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In response to the demand for improved performance among various gene-sequencing machines, postprocessing error-correction procedures associated with optical signal alignment and analysis, quality scoring, reference genome alignment and assembly, and variant calling are essential.2,11 The tremendous data processing load derived from massively synchronous SBS reactions forms the bottleneck of sequencing technology with respect to the bandwidth limitations of the read rate of the CCD detector.1 Going forward, improvements in signal intensity by increasing the photon collection efficiency will eliminate the data processing complexity of the error correction, and the mitigation of redundant access of data bandwidth from unnecessary pixels for imaging (~100 pixels per sequencing site in previous technologies) will increase the sequencing rate and parallel SBS reaction throughputs. PacBio and Illumina’s cutting-edge PacBio Sequel™ System and Illumina iSeq™ 100 Sequencing System were launched in late 2016 and early 2018, respectively. One notable innovation of these systems is the placement of the sequencing platform of the original SBS techniques on complementary metal-oxide-semiconductor (CMOS) sensors with millions of sequencing reads on a chip using semiconducting integration technology.13 In machine vision, the performance merits of CMOS over CCD, such as its higher speed, lower noise, lower power consumption, and higher defect pixel tolerance, promise more applications for fabrication integration.14-16 Owing to the direct stacking of the SBS reaction nanowell array onto the detection pixels in a one-to-one set alignment, the signal change in the effective reaction space can be absolutely converted for algorithmic decoding, reducing the imaging process load originating from the drift of z-axis focusing and x/y-axis displacement. Furthermore, the photon collection efficiency can be significantly improved by the close contact of the nanowells with the CMOS detectors.17-20 The great success of this integration offers the possibility of compiling all data on a miniature device with real-time analytical functionality, massively parallel capabilities, and single-molecule sensitivity.3,20,21 Despite the launch of the PacBio Sequel™ and Illumina iSeq™ systems, limited technical data on their NPL design or stacked nanostructures have been reported.22-24 Essentially, the integrated device should be composed of the following basic elements: the reaction sites (such as a nanowell or

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nano-/micro- patterns with a heterogeneous surface),20,25 an illumination mechanism for exciting fluorescent molecules (such as grating coupling into a waveguide under/surrounding the reaction sites or external light source),25-30 an excitation light-blocking mechanism (such as interference or absorption filters),20,30,31 components for light distinction (such as a Bayer filter mosaic, beam splitting/dispersion structure, or plasmonic color filter),9,31-34 and a light-sensitive detector (such as a CMOS

sensor).

Other

optional

components,

including

microfluidics,16,20

temperature-

controlling/sensing units,20,30 electric field applying/electronic recording units,18,19,35 and light condensers,36-38 can also be embedded in NPLs. Various combinations and properties of NPL designs could be utilized for CMOS integration. However, the NPL design and arrangement will largely dominate the system performance and the complexity of the decoding algorithm. Hence, we demonstrate a completely custom NPL-integrated TPD CMOS system39 for distinguishing among fluorescent molecules. A specially designed TPD CMOS sensor possessing an array of 256x256 pixels with three spectrum sensitive photodiodes vertically stacked in a pixel was employed for emission light detection, and its optical performance was analyzed before and after NPL integration. The functions of the photonic nanostructures of the devices, including the grating coupling planar waveguide (for supplying excitation light), metal pinhole (for excitation light reduction), interference/absorption filter(s) (for selective wavelength rejection/transmission) between the nanowells (as reaction sites) and the sensor pixels, are herein discussed.

Results/Discussion NPL structures and functions. To effectively optimize and compare the NPL nanostructures, one nanowell, one grating waveguide, four pinhole diameters, and one interference filter are arranged on an integrated TPD CMOS with or without color filters. The schematic design of the NPL-integrated TPD CMOS and cross-sectional SEM images of the grating area and the four pinhole sizes with embedded color filters are shown in Fig. 1. The detailed fabrication process of the NPLs is described in Supplementary Note. 1.

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The grating structure can couple up to 50% of the incident light and redistribute the energy uniformly into the Ta2O5 (n=2.16) planar waveguide by the total reflective confinement between the upper and lower claddings of SiO2 (n=1.46).28,29 The function of the T-shape nanowell bottom, which is exposed to a 20-nm Al2O3 layer on the waveguide, was to load or immobilize individual biomolecules for fluorescent reaction observation in the evanescence field, minimizing the background signal from the liquid solution and excitation light scattering. However, the intensity of the excitation light scattering generated from the incident light collision at the grating coupler, the nanowell structure, and the interfaces between the waveguide and the claddings conceal the tiny signals associated with the biomolecular reactions. Thus, three components in the NPLs were arranged between the reaction sites and the TPD sensors to suppress the wavelength of the excitation light: a metal pinhole, an interference filter, and a color filter. The opening diameters of the pinholes allowed different angles of fluorophore emission (as a point-source signal) and excitation light scattering (as background noise, which is largely distributed over a few tens of degrees around the incidence direction)40-42 to pass through. The optimized opening size of the pinhole was first assigned a diameter of 2 μm (offering an optical density (OD) of 2.8 for blocking excitation scattering) for maximum emission collection efficiency, and ø1.0-μm, ø1.5-μm, and ø2.5-μm pinholes were formed for comparison (refer to Supplementary Fig. S1(l)). Considering that the filter OD of a fluorescence microscope is ~9,43 excitation blocking with an OD of 6 should be then achieved with filters (Supplementary Fig. S2). The interference filter exhibits an excellent blocking OD and high emission transmission with a very steep spectral edge in a nonfluorescent thin film layer. However, the major drawback is that the equivalent spectrum shifts due to the angle of incidence (AOI).43 the color filter (0.5 μm, OD 1.04 at 488 nm), owing to the autofluorescence property of the organic absorption filter and non-AOI sensitivity, was arranged under the interference filter (2.6 μm, OD 4.87 at 488 nm) to absorb larger-angle scattering leakage from the interference filter. The NPL design and filter properties are detailed in Supplementary Note. 2.

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TPD optical properties. The specially designed TPD CMOS possesses three stacked P-N junction photodiodes. The radiation absorption wavelength of each junction was defined by vertically isolating an appropriate penetration depth of silicon to generate its own distinct light responsivity.15,44 The response curve peaks of junction one (JN1), junction two (JN2), and junction three (JN3) of our TPD occurred at 440 nm, 570 nm, and 690 nm, respectively. After the deposition of the interference filter and/or color filter, different excitation blocking performances could be achieved (Supplementary Fig. S3 and Supplementary Note. 3). A basic criterion of a high-performance detector is linearity, an essential characteristic for developing a decoding algorithm.14,45 Unless a pixel reaches full-well capacity, the collected photons of each junction should be proportional to the converted electronic signal. The dynamic range, representing the quality of an image sensor, is defined as the ratio of the highest detectable illumination level to the reference dark noise difference (spatially or temporally). A high dynamic range provides a wide dynamic intensity range for identifying different types of fluorescence or BioLED signals.14,15,45 In Fig. 2 and Supplementary Fig. S4, the nonlinearity of the normal TPD and NPL-integrated TPD exhibits a 45 dB in each junction and under all integration conditions; however, the dynamic range of the NPL-integrated TPD decreases significantly with long integration times, especially those longer than 500 ms (30-40 dB). The degradation in performance after NPL integration was caused by the increasing peak-to-peak noise level and leakage current accumulation during the integration time, which originates from the stress-induced dislocation of the CMOS sensor.14,15 This issue might be eliminated by reducing the warpage and/or temperature of thinfilm deposition during NPL integration.

Coupling and scattering. The NPL substrate was equipped with an imaging device with sufficient illumination energy for exciting fluorescent molecules in the picoliter reaction sites, and the emission signal could be effectively collected into the TPD with an adequate signal-to-noise ratio (SNR) to

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monitor specific changes in bioreactions. The grating nanostructures on the Ta2O5 waveguide couple the excitation energy into the waveguide. The period and depth affect the coupling efficiency of a grating coupler, and the coupling angle is extremely sensitive, showing up to a 10-fold intensity variation over an incident angle range of ±0.5°. The propagation loss of the waveguide, which is highly proportional to the surface roughness of the interface between the Ta2O5 and SiO2 layers, causes energy dissipation in the planar waveguide.23,46 The upper and lower claddings of SiO2 should be thicker than 500 nm, and the average roughness (Ra) should be controlled to less than 0.5 nm.46 In Supplementary Movie S1, the maximum planar waveguide coupling efficiency occurs at an incidence angle of only 3.7°, and the line laser is moved to the inner edge (over a few tens of micrometers) of the grating pattern. As shown in Fig. 3, four regions with different pinhole sizes were measured with the best coupling angle of 3.7±0.2°. The propagation loss is lowest in the region with the smallest pinholes: it is -6.1 dB/cm in the regions with the ø1.0-μm and ø1.5-μm pinholes, -7.6 dB/cm in the region with the ø2.0-μm pinhole, and -9.1 dB/cm in the region with the ø2.5-μm pinholes. Although the Ra of the interface in the four pinhole regions was controlled to be 0.5 nm (data not shown), the formation of a small crack extending from the edge of the pinhole resulted in a higher propagation loss. This issue could be resolved to reach a propagation loss of -3 dB/cm by fine-tuning the chemical-mechanical planarization (CMP) procedure. The OD performance of the 8 split designs for the suppression of the laser scattering incident onto the TPD was directly measured by the NPL-integrated TPD CMOS. The scattering mapping intensity of the 256x256 array features 8 regions, as shown in Fig. 4. The dark intensity levels of JN1, JN2, and JN3 were 10.0±3.0, 22.6±4.9, and 12.4±3.1 mV, respectively. Regions 1-4, each of which features a pinhole that has a different size and that is embedded with a color filter, showed distinctly lower scattering signals than those of regions 5-8, which were not embedded with a color filter. A large pinhole provides a high probability of transmitting scattered light through the interference filter over a large angle; thus, a color filter is essential for absorbing the leakage energy. Furthermore, the scattering

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intensities at columns 0-20 were higher than those at the other columns due to the strong scattering that occurs under the grating area. This issue could be addressed by elongating the distance between the grating and the sensor array or by replacing the Ti metal layers (on the upper cladding and under the bottom cladding) with a stronger light-absorbing opaque material.

Decoding algorithm and artificial sequence testing. The spectrum-distinguishing algorithm basically utilized three responsivities of the TPD to create a set of junction intensities for the classification of different wavelengths. For a fixed intensity of incidence, each wavelength should acquire a distributive cluster on a three-dimensional (3D) junction map with at least 100 data points and average a few tens of pixels as a decoding reference value. Once the clusters of incident light do not overlap, the received signals can be decoded by attributing the signal to the wavelength with the minimum distance on the 3D junction map. For a variable intensity of incidence, such as that observed for fluorescence, the absolute junction value should be normalized to exclude the intensity fluctuation. Here, the values of JN1 and JN3 were divided by that of JN2 to form the x-axis and y-axis of a twodimensional (2D) junction map, respectively. The degree of difficulty of decoding is intimately related to the SNR of the unknown pulses. As shown in Supplementary Tab. S1 and Fig. S5, our normal TPD CMOS demonstrated a decoding limitation associated with the illumination (~3125 ph/(pixel·data), 25-ms integration time, and 150-ms duration) for an artificial sequence of fixed or random pulses/intervals. As shown in Fig. 5(a) and Supplementary Fig. S6(a), the junction values of the 3D map were proportional to the incident intensity, but the divergence (∇) of the distribution of 100 continuous data increased in going from strong to weak incident intensities. Portions of the four artificial light clusters overlapped with each other (resulting in error decoding) at intensities of 5625 and 2813 ph/(pixel·data), with divergence values of 0.0452 and 0.0785, respectively. The obscure boundary between two (625 and 656 nm at the intensity of 5625 ph/(pixel·data)) or three clusters (590, 625, and 656 nm at the intensity of 2813 ph/(pixel·data)) led to the low decoding accuracy, which is also referred to as the decoding accuracy rate in

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Supplementary Tab. S1. After NPL integration, the filter(s) altered the responsivity of the normal TPD (Supplementary Fig. S3 (d)-(g)) to block blue and green light at wavelengths shorter than 550 nm. The 3D junction map is remeasured in Fig. 5(b) and Supplementary Fig. S6(b) to examine the change in performance. As previously mentioned, the fabrication process increased the noise variation; compared to that of the normal TPD, the degree of divergence increased 1.02~1.43-fold under 1.3-fold illumination. When coupling a 473-nm laser into the planar waveguide via a grating, various types of scattering and instability of the laser power generated a noisy intensity signal affecting the SNR. As shown in Fig. 5(c) and Supplementary Fig. S6(c), the NPL with a ø2-μm pinhole and a color filter could effectively block scattering under laser powers of 50 and 250 mW. However, the divergence of the 3D reference map increased 10-fold for the NPL with a ø2-μm pinhole without a color filter under an incident power of only 50 mW. This result indicated that the color filter embedded under an interference filter can provide better scattering inhibition. As shown in Supplementary Fig. S6, the junction values of the 3D map from 16 neighboring pixels varied owing to the doping variation in the triple-junction photodiode. Thus, further improvement of the decoding algorithm to achieve a higher decoding accuracy can be attained by comparing the spatial distances using the individual junction map of each pixel (instead of the overall junction map averaged from many pixels).

Fluorescent molecule detection ability. Because a nanowell on a planar waveguide is utilized to confine a reactive region for fluorescence detection, the quantification of fluorescent molecules in the nanowell provides valuable information for assessing the performance of an NPL-integrated sensor. The penetration depth of the Ta2O5 planar waveguide was simulated as shown in Supplementary Fig. S7, and the effective region was determined to be ~29 nm (with 1/e intensity) from the surface of the Ta2O5 waveguide. In the effective region, the average intensities per pixel of PSB (RED) and PSB (YG2) were similar to that of QD625; however, the total number of photons detected for the PSBs (~26

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pixels) was approximately 2-3-fold higher than that detected for QD625 (~13 pixels). As shown in Fig. 6(a)-(c), the 2D junction map of five PSBs (one ø200-nm PSB in a ø210-nm nanowell) was measured, and it matched the distribution of the calculated map. (Due to the nonconstant intensity of real fluorescence, such as the variation in the distance of the evanescence field, variation in an excitation intensity, quenching/bleaching, and single molecular spectrum shift,47 the junction intensity should be normalized to eliminate intensity variations.) The discreteness among the five clusters enabled decoding for identifying different PSBs. Furthermore, Alexa 488, Atto 514, and Atto 532 were measured under the decodable conditions as shown in Fig. 6(d)-(f) and Supplementary Fig. S8. The signal intensities of the three dyes at a concentration of 10 μM were all detectable. However, the clusters of the three dyes were adequately separated in the 2D junction map for decoding only when the concentration exceeded 100 μM (14.5 dyes). Considering that the dark noise peak-to-peak variations of JN1/JN2/JN3 were 6.92/5.01/3.91 mV, the minimum dye numbers for an SNR of 3 of Alexa 488/Atto 514/Atto 532 were 0.96/1.73/1.78 under an integration time of 150 ms. These results prove that the NPL-integrated TPD is sensitive enough to monitor fluorescent dyes at the single-molecule level (Supplementary Tab. S2). The distinguishability is also closely related to the original spectrum difference among the selected dyes. Thus, for the single-molecule level applications, the dyes should have sufficient emission spectrum separation, such as long Stoke shift dyes,48 to provide better decoding resolution by the NPLintegrated TPD CMOS with single blue laser excitation.

Conclusions The use of nanostructure-integrated CMOSs as fluorescence signal detection systems in the field of DNA sequencing has recently evolved. Vertically stacking photodiodes in a pixel can equip one pixel with different photon absorption properties for blue to red wavelengths from shallow to deep junctions. In this study, we developed a triple-junction photodiode CMOS and integrated the CMOS

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with nanophotonic layers, including nanowells, a waveguide, metal pinholes, and filters. We succeeded in developing a suitable decoding algorithm and demonstrating the distinguishability of the differences in wavelength from artificial light and fluorescent molecules. This miniaturized device can be employed as a single-molecule DNA sequencer for massive parallelization and real-time spectral discrimination once the dark current of the TPD CMOS is further suppressed and the NPL fabrication parameters are optimized, such as lower temperature and lower autofluorescence thin film deposition, better ODs of the filters, and adding a light collection component for a higher photon collection efficiency.

Methods/Experimental Materials. Fluorescent PSBs, PSF-200NM, 0.19 μm, Yellow-Green 1 (ex/em 480/501), Orange (ex/em 475/540), and Red (abbreviated as PSB (RED), ex/em 538/584), were purchased from Magsphere Inc. (USA). PSBs Dragon Green (ex/em 480/520, Cat. #CP01F, Streptavidin Coated Microsphere Dragon Green 0.22 μm) and Yellow-Green 2 (abbreviated as PSB (YG2), ex/em 441/486, Cat. #09834, Fluoresbrite® YG Carboxylate Microspheres 0.20 µm) were purchased from Bangs Laboratories, Inc., USA and Polyscience, Inc., USA. Fluorescent dye Alexa Fluor™ 488 NHS Ester (Succinimidyl Ester) (abbreviated as Alexa 488, Cat. #A20000) was purchased from InvitrogenTM, Thermo Fisher Scientific, USA. Fluorescent dyes Atto 514-NHS ester (abbreviated as Atto 514, Cat. #67455) and Atto 532-NHS ester (abbreviated as Atto 532, Cat. #88793) were purchased from SIGMA, USA. Quantum dots QD625 (Cat. #Q22063) were purchased from InvitrogenTM, Thermo Fisher Scientific, USA.

Filter transmission measurement. An interference filter was deposited by ion-beam-assisted electron beam evaporation deposition (IAD) on the monitoring 2” glass wafers accompanying the NPLintegrated 8” TPD CMOS wafer. A color filter was spin-coated on a monitoring 4” glass wafer with a thickness of 0.5-μm and postbaked under the same conditions applied to the NPL-integrated TPD

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CMOS wafers. The transmittance of the monitoring samples was measured from 400 nm to 700 nm (Supplementary Fig. S2). In this study, all measurements were performed at room temperature unless indicated otherwise.

TPD responsivity. All TPD CMOS sensors were diced and wire-bond packaged on a pin gray array (PGA) 257-pin board by Integrated Service Technology (iST) Inc., Taiwan. The design of the sensor board and the software for accessing signals of the triple-junction detector array were customized by Personal Genomics, Inc., Taiwan.39 The sensor board and the wired-bond packaged TPD CMOS sensor with and without NPLs are shown in Supplementary Fig. S3(a)-(c). The response curve was measured by an exposed TPD CMOS sensor in a uniform light environment with calibrated photon intensities and wavelengths. The setup used a spot light source (Hamamatsu LC8, L9588-03, Japan) through a monochromator (Spectral Products, CM110 1/8m, USA) to generate light from 400 nm to 950 nm at 10 nm intervals, and the light was passed through a photometric integrating sphere with an outlet connected to a calibrated photodiode module (Hamamatsu, S3072, Japan) on a picoammeter (Keithley, 6485, USA). A C feedback program controlled the photon illumination density per millisecond when the TPD CMOS sensor was exposed to the main opening of the integrating sphere. For each condition, the response curve was measured and plotted as the maximum, minimum, and average responses from 8 center pixels (from R128C124 to R128C131) on a randomly selected TPD CMOS sensor with or without the fabrication processes. The TPD plus color filter represents the fabrication conditions in Supplementary Fig. S1(d). TPD plus interference filter and TPD plus color and interference filters indicate that the interference filter-coated glass was placed on the normal TPD and the TPD with color filter embedded in SiO2, respectively.

TPD signal and noise characteristics. The dark noise peak-to-peak variation was indexed by subtracting the minimum value from the maximum value of 100 data points per integration time and

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calculated by averaging 16 pixels (from R100C100 to R103C103 for the normal TPD and R156C106 to R159C109 for the NPL-integrated TPD) per die and 4 dies per condition for the normal TPD CMOS or the NPL-integrated TPD CMOS with a ø2.0-μm pinhole and color filter. Before data collection, each die was run for 15-20 min at 20°C to reach thermal equilibrium. The dynamic range was calculated based on five integration time regions of 25-100 ms (by averaging the data from 25 ms, 50 ms, 75 ms, and 100 ms), 101-200 ms (by averaging the data from 150 ms and 200 ms), 201-500 ms (by averaging the data from 300 ms and 500 ms), 501-1000 ms (by averaging the data from 750 ms and 1000 ms), and longer than 1000 ms (by averaging the data from 1250 ms and 1500 ms). The saturation voltage of each junction was 1334 mV; thus, the dynamic range could be obtained by the saturation value divided by the dark noise peak-to-peak variation.

Grating waveguide performance. For planar waveguide coupling, a 473-nm line laser (MBL-FN-473, 5-degree fan angle, power stability