HBV超灵敏检测技术研究进展及输血应用前景

doi:10.3969/j.issn.1671-2587.2025.05.015

临床输血与检验 ›› 2025, Vol. 27 ›› Issue (5): 681-688.DOI: 10.3969/j.issn.1671-2587.2025.05.015

HBV超灵敏检测技术研究进展及输血应用前景

易萌¹, 钟晓龙², 王洋¹, 杜晓林¹, 樊斌¹, 段晓琼¹

¹中国医学科学院北京协和医学院输血研究所,四川成都 610052;
²绵阳市第三人民医院（四川省精神卫生中心）,四川绵阳 621000

收稿日期:2025-09-01 修回日期:2025-09-08 出版日期:2025-10-20 发布日期:2025-10-11
通讯作者: 段晓琼,主要从事肝炎病毒致病机制和输血相关新发再发病原体的流行及机制研究,（E-mail）xiaoqiongduan@163.com。共同通信作者：樊斌,主要从事输血相关传染病致病机制及流行病学研究,（E-mail）binguog@sina.com。
作者简介:易萌,主要从事输血传播病原体致病机制研究,（E-mail）yimeng@student.pumc.edu.cn。并列第一作者：钟晓龙,主要从事疑难血型及输血相关传染病研究,（E-mail）xiaolongzhong107@163.com。

Advances in Ultra-sensitive HBV Detection Technologies and Their Transfusion Applications

YI Meng¹, ZHONG Xiaolong², WANG Yang¹, DU Xiaolin¹, FAN Bin¹, DUAN Xiaoqiong¹

¹Institute of Blood Transfusion, Chinese Academy of Medical Sciences & Peking Union Medical College, Chengdu 610052;
²The Third Hospital of Mianyang, Sichuan Mental Health Center, Mianyang 621000

Received:2025-09-01 Revised:2025-09-08 Online:2025-10-20 Published:2025-10-11

摘要/Abstract

摘要： 隐匿性乙型肝炎病毒感染（OBI）因其极低病毒载量及肝内cccDNA持续潜伏特性,构成全球公共卫生安全的深层挑战。其防控长期受限于传统检测技术的灵敏度瓶颈,亟需通过技术革新实现突破。新兴检测技术在多角度实现突破：液滴数字PCR通过微区室化扩增实现超灵敏定量;CRISPR-Cas系统耦合等温扩增推动即时诊断发展;第三代测序技术开启病毒序列解析新维度;纳米材料介导的信号放大机制重塑基层筛查模式。本文对OBI最新诊断技术发展进行概述,对各个诊断技术的优劣势、输血相关应用前景等进行综合分析并提出建议,为推进我国输血相关OBI风险控制、开发适合我国国情的新检测技术提供科学依据。

关键词: OBI, 输血安全, 血液筛查, 超灵敏检测, ddPCR, CRISPR-Cas

Abstract: Occult hepatitis B virus infection (OBI), characterized by extremely low viral loads and the persistent latency of intrahepatic cccDNA, poses a profound challenge to global public health security. Its prevention and control have long been constrained by the sensitivity limitations of conventional diagnostic methods, highlighting the urgent need for technological breakthroughs. Emerging detection technologies are driving progress from multiple perspectives: droplet digital PCR enables ultra-sensitive quantification through micro-compartmentalized amplification; CRISPR-Cas systems coupled with isothermal amplification are advancing the development of point-of-care diagnostics; third-generation sequencing technologies open new dimensions for viral genome analysis; and nanomaterial-mediated signal amplification mechanisms are reshaping screening models at the primary-care level. This review summarizes recent advances in OBI diagnostic technologies, provides a comprehensive analysis of their advantages, limitations, and transfusion-related applications, and offers recommendations to support improved OBI risk control in blood transfusion and the development of novel diagnostic strategies tailored to China's national context.

Key words: Occult hepatitis B virus infection (OBI), Transfusion safety, Blood screening, Ultra-sensitive detection, Droplet digital PCR (ddPCR), CRISPR-Cas

中图分类号:

R457

易萌, 钟晓龙, 王洋, 杜晓林, 樊斌, 段晓琼. HBV超灵敏检测技术研究进展及输血应用前景[J]. 临床输血与检验, 2025, 27(5): 681-688.

YI Meng, ZHONG Xiaolong, WANG Yang, DU Xiaolin, FAN Bin, DUAN Xiaoqiong. Advances in Ultra-sensitive HBV Detection Technologies and Their Transfusion Applications[J]. JOURNAL OF CLINICAL TRANSFUSION AND LABORATORY MEDICINE, 2025, 27(5): 681-688.

参考文献

[1] JENG W J,PAPATHEODORIDIS G V,LOK A S F. Hepatitis B[J]. Lancet,2023,401(10381):1039-1052.
[2] HSU Y C,NGUYEN M H.Curing chronic hepatitis B virus infection[J]. Lancet Infect Dis,2023,23(4):392-393.
[3] HSU Y C,HUANG D Q,NGUYEN M H.Global burden of hepatitis B virus: current status,missed opportunities and a call for action[J]. Nat Rev Gastroenterol Hepatol, 2023,20(8):524-537.
[4] SAITTA C,POLLICINO T,RAIMONDO G.Occult hepatitis B virus infection: an update[J]. Viruses,2022, 14(7). DOI:10.3390/v14071504.
[5] DE ALMEIDA N A A,DE PAULA V S. Occult Hepatitis B virus (HBV) infection and challenges for hepatitis elimination: A literature review[J]. J Appl Microbiol, 2022,132(3):1616-1635.
[6] WU T,KWOK R M,TRAN T T.Isolated anti-HBc: the relevance of hepatitis B core antibody-a review of new issues[J]. Am J Gastroenterol,2017,112(12):1780-1788.
[7] FU M X,SIMMONDS P,ANDERSSON M,et al.Biomarkers of transfusion transmitted occult hepatitis B virus infection: Where are we and what next?[J]. Rev Med Virol,2024,34(2):e2525.
[8] HE P,ZHANG P X,FANG Y P,et al.The role of HBV cccDNA in occult hepatitis B virus infection[J]. Mol Cell Biochem,2023,478(10):2297-2307.
[9] BUCIO-ORTIZ L,ENRIQUEZ-NAVARRO K, MALDONADO-RODRÍGUEZ A,et al. Occult hepatitis B virus infection in hepatic diseases and its significance for the WHO's elimination plan of viral hepatitis[J]. Pathogens,2024,13(8):662.
[10] CANDOTTI D,ASSENNATO S M,LAPERCHE S, et al.Multiple HBV transfusion transmissions from undetected occult infections: revising the minimal infectious dose[J]. Gut,2019,68(2):313-321.
[11] KLEIN D.Quantification using real-time PCR technology: applications and limitations[J]. Trends Mol Med,2002,8(6):257-260.
[12] IM Y R,JAGDISH R,LEITH D,et al.Prevalence of occult hepatitis B virus infection in adults: a systematic review and meta-analysis[J]. Lancet Gastroenterol Hepatol,2022,7(10):932-942.
[13] WANG C W,XUE R R,WANG X R,et al.High-sensitivity HBV DNA test for the diagnosis of occult HBV infection: commonly used but not reliable[J]. Front Cell Infect Microbiol,2023,13:1186877.
[14] CAI J Y,WU W M,WU J L,et al.Prevalence and clinical characteristics of hepatitis B surface antigen-negative/hepatitis B core antibody-positive patients with detectable serum hepatitis B virus DNA[J]. Ann Transl Med,2022,10(1):25.
[15] MOROZOV S,BATSKIKH S.Reactivation of hepatitis B virus infection—an important aspect of multifaceted problem[J]. World J Gastroenterol,2024,30(26):3193-3197.
[16] LIU S,ZHOU B,VALDES J D,et al.Serum hepatitis B virus RNA: a new potential biomarker for chronic hepatitis B virus infection[J]. Hepatology,2019,69(4): 1816-1827.
[17] SAMADI E,MIRSHAHABI H,MOTAMED N,et al.Prevalence of occult hepatitis B virus infection in hemodialysis patients using nested PCR[J]. Rep Biochem Mol Biol,2020,9(1):82-88.
[18] ROTH W K.History and future of nucleic acid amplification technology blood donor testing[J]. Transfus Med Hemother,2019,46(2):67-75.
[19] YE X L,LIU L H,CHEN L N,et al.High-frequency notable HBV mutations identified in blood donors with occult hepatitis B infection from Heyuan City of Southern China[J]. Front Immunol,2022,13:754383.
[20] WANG K N,SANG B L,HE L M,et al.Construction of dPCR and qPCR integrated system based on commercially available low-cost hardware[J]. Analyst, 2022, 147(15):3494-3503.
[21] ZHANG L X,PARVIN R,FAN Q H,et al.Emerging digital PCR technology in precision medicine[J]. Biosens Bioelectron,2022,211:114344.
[22] LIU X J,FENG J P,ZHANG Q H,et al.Analytical comparisons of SARS-COV-2 detection by qRT-PCR and ddPCR with multiple primer/probe sets[J]. Emerg Microbes Infect,2020,9(1):1175-1179.
[23] PIERMATTEO L,SCUTARI R,CHIRICHIELLO R, et al.Droplet digital PCR assay as an innovative and promising highly sensitive assay to unveil residual and cryptic HBV replication in peripheral compartment[J]. Methods,2022,201:74-81.
[24] HAYASHI S,ISOGAWA M,KAWASHIMA K,et al.Droplet digital PCR assay provides intrahepatic HBV cccDNA quantification tool for clinical application[J]. Sci Rep,2022,12(1):2133.
[25] LIMOTHAI U,CHUAYPEN N,POOVORAWAN K,et al.Reverse transcriptase droplet digital PCR vs reverse transcriptase quantitative real-time PCR for serum HBV RNA quantification[J]. J Med Virol,2020, 92(12):3365-3372.
[26] WANG K N,LI B,GUO Y,et al.An integrated digital PCR system with high universality and low cost for nucleic acid detection[J]. Front Bioeng Biotechnol,2022, 10:947895.
[27] TAYLOR S C,NADEAU K,ABBASI M,et al.The ultimate qPCR experiment: producing publication quality, reproducible data the first time[J]. Trends Biotechnol, 2019,37(7):761-774.
[28] WEI L Y,WANG Z L,SHE Y B,et al.CRISPR/cas multiplexed biosensing: advances,challenges,and perspectives[J]. Anal Chem,2025,97(23):11943-11958.
[29] PATCHSUNG M,JANTARUG K,PATTAMA A,et al.Clinical validation of a Cas13-based assay for the detection of SARS-CoV-2 RNA[J]. Nat Biomed Eng, 2020,4(12):1140-1149.
[30] GOOTENBERG J S,ABUDAYYEH O O,LEE J W,et al.Nucleic acid detection with CRISPR-Cas13a/C2c2[J]. Science,2017,356(6336):438-442.
[31] CHEN J S,MA E B,HARRINGTON L B,et al.CRISPR-Cas12a target binding unleashes indiscriminate single-stranded DNase activity[J]. Science,2018,360(6387): 436-439.
[32] DING R H,LONG J Z,YUAN M Z,et al.CRISPR/Cas12-based ultra-sensitive and specific point-of-care detection of HBV[J]. Int J Mol Sci,2021,22(9):4842.
[33] SHI K,XIE S Y,TIAN R Y,et al.A CRISPR-Cas autocatalysis-driven feedback amplification network for supersensitive DNA diagnostics[J]. Sci Adv,2021,7(5). DOI:10.1126/sciadv.abc7802.
[34] 公少华. 基于CRISPR-Cas12a的酶催化核酸即时检测方法研究[D]. 济南:山东师范大学,2023.
[35] WANG S,LI H,KOU Z,et al.Highly sensitive and specific detection of hepatitis B virus DNA and drug resistance mutations utilizing the PCR-based CRISPR-Cas13a system[J]. Clin Microbiol Infect,2021,27(3): 443-450.
[36] TIAN Y,FAN Z H,XU L,et al.CRISPR/Cas13a-assisted rapid and portable HBV DNA detection for low-level viremia patients[J]. Emerg Microbes Infect,2023,12(1): e2177088.
[37] GARCIA-GARCIA S,CORTESE M F,RODRÍGUEZ-ALGARRA F,et al. Next-generation sequencing for the diagnosis of hepatitis B: current status and future prospects[J]. Expert Rev Mol Diagn,2021,21(4):381-396.
[38] MCNAUGHTON A L,ROBERTS H E,BONSALL D, et al.Illumina and Nanopore methods for whole genome sequencing of hepatitis B virus (HBV)[J]. Sci Rep,2019, 9(1):7081.
[39] LEE G Y,PARK K,LEE Y S,et al.Molecular diagnosis of patients with hepatitis A virus infection using amplicon-based nanopore sequencing[J]. PLoS One,2023,18(7): e0288361.
[40] WU Y D,GUO W S,PENG W P,et al.Enhanced fluorescence ELISA based on HAT triggering fluorescence "turn-on" with enzyme—antibody dual labeled AuNP probes for ultrasensitive detection of AFP and HBsAg[J]. ACS Appl Mater Interfaces,2017,9(11): 9369-9377.
[41] ZHANG H,SHI Y F,WU Z G,et al.Nanoparticle-based biosensor integrated with multiple cross-displacement amplification for visual and rapid identification of hepatitis B virus and hepatitis C virus[J]. Microbiol Spectr,2025,13(5):e0173824.
[42] CHEN C C,LAI Z L,WANG G J,et al.Polymerase chain reaction-free detection of hepatitis B virus DNA using a nanostructured impedance biosensor[J]. Biosens Bioelectron,2016,77:603-608.
[43] WU Y Q,ZENG L F,XIONG Y,et al.Fluorescence ELISA based on glucose oxidase-mediated fluorescence quenching of quantum dots for highly sensitive detection of Hepatitis B[J]. Talanta,2018,181:258-264.
[44] DELSHADI S,FRATZL M,RAMEL O,et al.Magnetically localized and wash-free fluorescence immunoassay (MLFIA): proof of concept and clinical applications[J]. Lab Chip,2023,23(4):645-658.
[45] JIANG L P,LI Y Y,XU Z,et al.Simultaneous electrochemical determination of two hepatitis B antigens using graphene-SnO₂ hybridized with sea urchin-like bimetallic nanoparticles[J]. Mikrochim Acta, 2021,188(4): 109.
[46] YANG Q S,WANG P,MA E H,et al.A sandwich-type electrochemical immunosensor based on Au@Pd nanodendrite functionalized MoO₂ nanosheet for highly sensitive detection of HBsAg[J]. Bioelectrochemistry, 2021,138:107713.
[47] CHEN R B,HU Y Q,CHEN M Z,et al.Naked-eye detection of hepatitis B surface antigen using gold nanoparticles aggregation and catalase-functionalized polystyrene nanospheres[J]. ACS Omega,2021,6(14): 9828-9833.
[48] CHEN X,SHI Y F,ZHAO Q,et al.One-step,rapid, nanoparticle-based biosensor platform for the simultaneous identification of hepatitis B virus and hepatitis C virus in clinical applications[J]. BMC Microbiol,2024,24(1): 455.
[49] AN J,YUAN M D,HAN Y Q,et al.Nano-immuno-conjugates inspired by hydrophilic perovskite fluorescent spheres and magnetic assisted for detection of hepatitis B surface antigen[J]. Mikrochim Acta,2024, 191(8):473.
[50] XU S S,WU X H,WU L,et al.Systematic optimization of universal real-time hypersensitive fast detection method for HBsAg in serum based on SERS[J]. Anal Chem, 2024,96(17):6784-6793.
[51] YONG S K,SHEN S K,CHIANG C W,et al.Silicon nanowire field-effect transistor as label-free detection of hepatitis B virus proteins with opposite net charges[J]. Biosensors,2021,11(11):442.
[52] YAN S X,KONG C P,CHENG J Z,et al.High sensitivity detection of Hepatitis B virus RNA based on 3D-DNA nanomachine and protein nanopore sensing[J]. Mol Biomed,2025,6(1):55.
[53] LI M H,TAO C Y,TANG Z Y,et al.Ultrasensitive clinical identification of hepatitis B surface antigen (HBsAg) by CRISPR-assisted nanopore sensing[J]. Biosens Bioelectron, 2025,286:117579.
[54] YARALı E,ERDEM A.Cobalt phthalocyanine-ionic liquid composite modified electrodes for the voltammetric detection of DNA hybridization related to hepatitis B virus[J]. Micromachines,2021,12(7):753.
[55] MAO J,WANG J X,CHEN H L,et al.Development of a sandwich-type electrochemical DNA sensor based on CeO₂/AuPt nanoprobes for highly sensitive detection of hepatitis B virus DNA[J]. Bioelectrochemistry,2025, 163:108901.
[56] LIU X X,LIU T,YUAN X L,et al.DNA locked-in double-faced-nanotube for ultrafast biosensing of hepatitis virus[J]. Small,2025,21(29):e2502223.

HBV超灵敏检测技术研究进展及输血应用前景

Advances in Ultra-sensitive HBV Detection Technologies and Their Transfusion Applications

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