面向单分子检测的低噪声可扩展模拟前端设计

      纳米孔即孔径在10^-9米数量级的小孔，其尺寸与核酸、蛋白质等单个小分子的直径相当。其传感原理为恒电压条件下测量单个被测物堵塞纳米孔引起的电流变化，在单分子检测中该堵塞变化明显。纳米孔检测的电流幅度为10 pA – 20 nA，需覆盖直流到一定的截止频率，截止频率一般为1 kHz – 10 MHz。微弱电流信号的传感对于读出电路提出了高增益和低噪声的挑战。

纳米孔离子电流，可通过基于跨阻放大电路的膜片钳技术进行可靠地观察，板级电路已具有高信噪比。Axopatch-200B作为膜片钳放大器金标准，具有优秀的噪声性能。但是台式机的设计限制了其高通量、便携式发展。随着集成电路技术与医疗健康技术的结合愈发紧密，电子设备的小型化为探测生物系统提供了新的技术路线。

本项工作基于0.18 μm CMOS工艺设计和验证了一款单分子检测用的模拟前端芯片，具有低噪声、高增益、高线性度和可靠性的特性。为了实现高线性度与高可靠性，本文采用了电阻反馈型跨阻放大器。为了实现亚GΩ数量级的跨阻增益，采用3 kΩ/□的多晶硅电阻，基于分段电阻技术和单端转双端输出方法实现了534 MΩ的等效跨阻增益。基于非单位增益缓冲器设计实现等效7.14 fF的高可靠性电容，将系统带宽拓展7倍。采用大尺寸PMOS输入对管（W/L=784 μm/0.24 μm）和源极退化技术降低系统噪声水平，实测等效输入电流噪声为1.05 pA_RMS (<1 kHz)，优于Axopatch-200B最高带宽挡位下实测14.1 pA_RMS (<1 kHz)的噪声水平，略优于商用测序仪MinION的噪声水平，可以进一步多通道推广。同时，单通道放大器成功验证了生物纳米孔对核酸单分子事件的探测。本文将系统拓展至四通道设计，片上预留电化学电极接口，使得读出电路可与纳米孔传感器垂直集成。片外实现补偿电路，将输出响应达到稳态的时间由9 ms减小至2 ms，提升了77.8%。

Nanopore, similar in size to single molecules like nucleic acid and protein, offers advantages for single-molecule detection. Sensing involves measuring current changes when nanopore is blocked under constant voltage, typically ranging from 10 pA to 20 nA at frequencies of 1 kHz to 10 MHz, posing challenges of high gain and low noise in readout circuit.

Patch-clamp technology offers a high signal-to-noise ratio method for observing nanopore ion currents. However, Axopatch-200B's desktop design limits its high-throughput and portable development. Integrated circuits technology drives electronic device miniaturization, enabling new approaches to investigating biological systems.

This work develops an analog front-end chip for single-molecule detection using 0.18 μm CMOS technology, featuring low noise, high gain, high linearity, and reliability. A resistor-feedback transimpedance amplifier is employed for high linearity and reliability. Achieving 534 MΩ equivalent transimpedance gain involves segmented resistor technique and a single-to-differential output method. A high-reliability capacitance equivalent to 7.14 fF is realized, expanding the system bandwidth by 7 times. Noise reduction is achieved with large-size PMOS input transistors and source degeneration technique, resulting in an input current noise of 1.05 pA_RMS (<1 kHz), superior to the Axopatch-200B's 14.1 pA_RMS (<1 kHz) noise level. The amplifier successfully detects biological events in nanopores and is expanded to a four-channel design with on-chip reserved electrochemical electrode interfaces for vertical integration with nanopore sensors. Off-chip compensation circuits reduce the output response settling time from 9 ms to 2 ms, a 77.8% improvement.

[1] MANRAO E A, DERRINGTON I M, PAVLENOK M, et al. Nucleotide discrimination with DNA immobilized in the MSPA nanopore[J]. PLoS ONE, 2011, 6(10):e25723.
[2] WORKMAN R E, TANG A D, TANG P S, et al. Nanopore native RNA sequencing of a human poly(A) transcriptome[J]. Nature Methods, 2019, 16(12): 1297-1305.
[3] ROBERTSON J W F, REINER J E. The Utility of Nanopore Technology for Protein and Peptide Sensing[M]//Proteomics. Wiley-VCH Verlag, 2018.
[4] PIGUET F, OULDALI H, PASTORIZA-GALLEGO M, et al. Identification of single amino acid differences in uniformly charged homopolymeric peptides with aerolysin nanopore[J]. Nature Communications, 2018, 9(1):996.
[5] LUCAS F L R, VERSLOOT R C A, YAKOVLIEVA L, et al. Protein identification by nanopore peptide profiling[J]. Nature Communications, 2021, 12(1):5795.
[6] CELAYA G, PERALES-CALVO J, et al. Label-Free, Multiplexed, Single-Molecule Analysis of Protein-DNA Complexes with Nanopores[J]. (2017):5815-5825.
[7] DERRINGTON I M, CRAIG J M, STAVA E, et al. Subangstrom single-molecule measurements of motor proteins using a nanopore[J]. Nature Biotechnology, 2015, 33(10): 1073-1075.
[8] ROSENSTEIN J. The promise of nanopore technology: Nanopore DNA sequencing represents a fundamental change in the way that genomic information is read, with potentially big savings[J]. IEEE Pulse, 2014, 5(4): 52-54.
[9] ROSENSTEIN J K, RAMAKRISHNAN S, ROSEMAN J, et al. Single ion channel recordings with CMOS-anchored lipid membranes[J]. Nano Letters, 2013, 13(6): 2682-2686.
[10] Axopatch 200B Patch Clamp Theory and Operation[R]. 1997.
[11] AWAN M A, WANG B, QUADIR N A, et al. Review and analysis of CMOS current readout circuits for biosensing applications[C]//Proceedings - IEEE International Symposium on Circuits and Systems: 2021-May. Institute of Electrical and Electronics Engineers Inc., 2021.
[12] DOOHWAN JUNG, SAGAR RAMESH KUMASHI, JONGSEOK PARK, et al. A CMOS Multimodality In-Pixel Electrochemical and Impedance Cellular Sensing Array for Massively Paralleled Synthetic Exoelectrogen Characterization[C]//2020 IEEE International Solid-State Circuits Conference. IEEE, 2020.
[13] YUN-SHIANG SHU, ZHI-XIN CHEN, YU-HONG LIN, et al. A 4.5mm2 Multimodal Biosensing SoC for PPG, ECG, BIOZ and GSR Acquisition in Consumer Wearable Devices[C]//2020 IEEE International Solid-State Circuits Conference. IEEE, 2020.
[14] LOPEZ C M, CHUN H S, BERTI L, et al. A 16384-electrode 1024-channel multimodal CMOS MEA for high-throughput intracellular action potential measurements and impedance spectroscopy in drug-screening applications[C]//2018 IEEE International Solid-State Circuits Conference: Vol. 61. Institute of Electrical and Electronics Engineers Inc., 2018: 464-466.
[15] ZHONG C B, MA H, WANG J J, et al. An ultra-low noise amplifier array system for high throughput single entity analysis[J]. Faraday Discussions, 2022, 233,33-43.
[16] YUN J, CHOI H, KIM J. Low-noise wide-bandwidth DNA readout instrument for nanopore applications[J]. Electronics Letters, 2017, 53(11): 706-708.
[17] KIM J, DUNBAR W B. High-precision low-power DNA readout interface chip for multichannel nanopore applications[J]. Sensors and Actuators, B: Chemical, 2016, 234: 273-277.
[18] KIM J, PEDROTTI K, DUNBAR W B. An area-efficient low-noise CMOS DNA detection sensor for multichannel nanopore applications[J]. Sensors and Actuators, B: Chemical, 2013, 176: 1051-1055.
[19] FANG S, YIN B, XIE W, et al. Low-noise and high-speed trans-impedance amplifier for nanopore sensor[J]. Review of Scientific Instruments, 2023, 94(7).
[20] YAN H, ZHANG Z, WENG T, et al. Recognition of bimolecular logic operation pattern based on a solid-state nanopore[M]//Sensors (Switzerland). MDPI AG, 2021: 1-11.
[21] KIM D, BYUN S, PU Y, et al. Design of a Current Sensing System with TIA Gain of 160 dBΩ and Input-Referred Noise of 1.8 pArms for Biosensor[J]. Sensors, 2023, 23(6):3019.
[22] SHEKAR S, NIEDZWIECKI D J, CHIEN C C, et al. Measurement of DNA translocation dynamics in a solid-state nanopore at 100 ns temporal resolution[J]. Nano Letters, 2016, 16(7): 4483-4489.
[23] DJEKIC D, FANTNER G, LIPS K, et al. A 0.1% THD, 1-MΩ to 1-GΩ Tunable, Temperature-Compensated Transimpedance Amplifier Using a Multi-Element Pseudo-Resistor[J]. IEEE Journal of Solid-State Circuits, 2018, 53(7): 1913-1923.
[24] SINA PARSNEJAD, HAITAO LI, ANDREW J. MASON. Compact CMOS Amperometric Readout for Nanopore Arrays in High Throughput Lab-on-CMOS[C]//2016 IEEE International Symposium on Circuits and Systems (ISCAS). IEEE, 2016.
[25] MULBERRY G, WHITE K A, KIM B N. Analysis of Simple Half-Shared Transimpedance Amplifier for Picoampere Biosensor Measurements[J]. IEEE Transactions on Biomedical Circuits and Systems, 2019, 13(2): 387-395.
[26] TAHERZADEH-SANI M, HUSSAIN HUSSAINI S M, REZAEE-DEHSORKH H, et al. A 170-dBΩ CMOS TIA With 52-pA Input-Referred Noise and 1-MHz Bandwidth for Very Low Current Sensing[J]. IEEE Transactions on Very Large Scale Integration (VLSI) Systems, 2017, 25(5): 1756-1766.
[27] HSU C L, JIANG H, VENKATESH A G, et al. A Hybrid Semi-Digital Transimpedance Amplifier with Noise Cancellation Technique for Nanopore-Based DNA Sequencing[J]. IEEE Transactions on Biomedical Circuits and Systems, 2015, 9(5): 652-661.
[28] DAWJI Y, HABIBI M, GHAFAR-ZADEH E, et al. A Scalable Discrete-Time Integrated CMOS Readout Array for Nanopore Based DNA Sequencing[J]. IEEE Access, 2021, 9: 155543-155554.
[29] WANG Y, JIN G, TANG H, et al. A potentiostat readout array for nanopore-based DNA sequencing[J]. IEICE Electronics Express, 2024: 21.20240118.
[30] PANSODTEE P, SELBERG J, JIA M, et al. The multi-channel potentiostat: Development and evaluation of a scalable mini-potentiostat array for investigating electrochemical reaction mechanisms[J]. PLoS ONE, 2021, 16(9):e0257167
[31] LU S Y, SHAN S S, LU T H, et al. A Review of CMOS Electrochemical Readout Interface Designs for Biomedical Assays[M]//IEEE Sensors Journal. Institute of Electrical and Electronics Engineers Inc., 2021: 12469-12483.
[32] LIU X, FAN Q, HU X, et al. A Fast Current Sensing Front-End IC Design for Nanopore-Based DNA Sequencing[C]//Proceedings of the International Conference on Anti-Counterfeiting, Security and Identification, ASID: 2022-December. IEEE Computer Society, 2022: 81-85.
[33] CRESCENTINI M, THEI F, BENNATI M, et al. A distributed amplifier system for bilayer lipid membrane (BLM) arrays with noise and individual offset cancellation[J]. IEEE Transactions on Biomedical Circuits and Systems, 2015, 9(3): 334-344.
[34] CAIRNS-GIBSON D F, COCKROFT S L. Functionalised nanopores: Chemical and biological modifications[J]. Chemical Science, 2022, 13(7): 1869-1882.
[35] WU Y, GOODING J J. The application of single molecule nanopore sensing for quantitative analysis[M]//Chemical Society Reviews. Royal Society of Chemistry, 2022: 3862-3885.
[36] CHENJIE DONG, YIZHOU JIANG, KE JIANG, et al. A 37.37µW-per-Cell Multifunctional Automated Nanopore Sequencing CMOS Platform with 16*8 Biosensor Array[C]//2020 IEEE International Symposium on Circuits and Systems (ISCAS): proceedings : ISCAS 2020 : Virtual Conference, October 10-21, 2020.
[37] ROSENSTEIN J K, WANUNU M, MERCHANT C A, et al. Integrated nanopore sensing platform with sub-microsecond temporal resolution[J]. Nature Methods, 2012, 9(5): 487-492.
[38] W. H. COULTER. Means for counting particles suspended in a fluid[P]. 1953.
[39] DEAMER D, AKESON M, BRANTON D. Three decades of nanopore sequencing[M]//Nature Biotechnology. Nature Publishing Group, 2016: 518-524.
[40] KASIANOWICZ J J, BRANDIN E, BRANTON D, et al. Characterization of individual polynucleotide molecules using a membrane channel[R]//Biophysics: Vol. 93. 1996.
[41] DERRINGTON I M, BUTLER T Z, COLLINS M D, et al. Nanopore DNA sequencing with MspA[J]. Proceedings of the National Academy of Sciences of the United States of America, 2010, 107 (37): 16060-16065.
[42] MANRAO E A, DERRINGTON I M, LASZLO A H, et al. Reading DNA at single-nucleotide resolution with a mutant MspA nanopore and phi29 DNA polymerase[J]. Nature Biotechnology, 2012, 30(4): 349-353.
[43] VAN DER VERREN S E, VAN GERVEN N, JONCKHEERE W, et al. A dual-constriction biological nanopore resolves homonucleotide sequences with high fidelity[J]. Nature Biotechnology, 2020, 38(12): 1415-1420.
[44] YU J, CAO C, LONG Y T. Selective and Sensitive Detection of Methylcytosine by Aerolysin Nanopore under Serum Condition[J]. Analytical Chemistry, 2017, 89(21): 11685-11689.
[45] JIALI LI, DEREK STEIN, CIARAN MCMULLAN, et al. Ion-beam sculpting at nanometre length scales[J]. Nature, 2001, 412(6843): 166-169.
[46] C. DEKKER. Solid-state nanopores[J]. Nature Nanotechnology, 2007, 2(4): 209-215.
[47] YING C, ZHANG Y, FENG Y, et al. 3D nanopore shape control by current-stimulus dielectric breakdown[J]. Applied Physics Letters, 2016, 109(6): 063105.
[48] SCHOCH R B, HAN J, RENAUD P. Transport phenomena in nanofluidics[J]. Reviews of Modern Physics, 2008, 80(3): 839-883.
[49] AMAYREH M, BAAKEN G, BEHRENDS J C, et al. A Fully Integrated Current-Mode Continuous-Time Delta-Sigma Modulator for Biological Nanopore Read Out[J]. IEEE Transactions on Biomedical Circuits and Systems, 2019, 13(1): 225-236.
[50] JACOB K. ROSENSTEIN, KENNETH L. SHEPARD. Temporal Resolution of Nanopore Sensor Recordings[C]//2013 35th annual international conference of the IEEE Engineering in Medicine and Biology Society (EMBC) 3-7 July 2013, Osaka, Japan. 2013.
[51] LIU H, ZHOU Q, WANG W, et al. Solid-State Nanopore Array: Manufacturing and Applications[M]//Small. John Wiley and Sons Inc, 2023.
[52] WANUNU M, SUTIN J, MCNALLY B, et al. DNA translocation governed by interactions with solid-state nanopores[J]. Biophysical Journal, 2008, 95(10): 4716-4725.
[53] SMEETS R M M, KEYSER U F, DEKKER N H, et al. Noise in solid-state nanopores[J]. Proceedings of the National Academy of Sciences of the United States of America, 2008, 105 (2): 417-421.
[54] FRAGASSO A, SCHMID S, DEKKER C. Comparing Current Noise in Biological and Solid-State Nanopores[M]//ACS Nano 2020, 14, 2, 1338–1349.
[55] WEN C, ZENG S, ARSTILA K, et al. Generalized Noise Study of Solid-State Nanopores at Low Frequencies[J]. ACS Sensors, 2017, 2(2): 300-307.
[56] LUGER P. Shot noise in ion channels[R]//Btochimwa et Btophysica Acta: Vol. 413. 1975.
[57] H. Frohlich, Theory of Dielectrics: Dielectric Constant and Dielectric Loss. [M]Oxford University Press, 1949.
[58] YANAGI I, FUJISAKI K, HAMAMURA H, et al. Thickness-dependent dielectric breakdown and nanopore creation on sub-10-nm-thick SiN membranes in solution[J]. Journal of Applied Physics, 2017, 121(4).
[59] RAZAVI. B. Design of Analog CMOS Integrated Circuits[M] McGraw Hill, 2016.
[60] MAGIEROWSKI S, HUANG Y, WANG C, et al. Nanopore-CMOS interfaces for DNA sequencing[M]// Biosensors 2016, 6(3), 42.
[61] SAKMANN_NEHER. Single-Channel Recording[M]//Single-Channel Recording. Springer US, 1995.
[62] FLEMING S J. Probing nanopore-DNA interactions with MspA[D]. 2017.