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Conventional Single-Stranded Adeno-Associated Virus (ssAAV) Service
Self-Complementary Adeno-Associated Virus (scAAV) Service
AAV Capsid Evolution & Screening Service
AAV Biodistribution & Shedding Assay Service
AAV Manufacturing / Large Scale Production

AAV Service

AAV OverviewResearch-Grade ServicesCDMO ServicesContact UsFAQ

Adeno-Associated Virus (AAV) stands as a leading delivery vector in gene therapy due to its non-pathogenic nature, low immunogenicity, and long-term expression capabilities. These properties have made AAV essential in treating neurodegenerative diseases, hereditary retinal disorders, and cancer immunotherapy. However, significant challenges persist in AAV development, including capsid targeting optimization, ITR sequence stability issues, and empty capsid ratio control. These limitations affect both experimental reliability and clinical translation.

AAV Overview

AAV belongs to the Parvoviridae family and is a non-enveloped, single-stranded DNA virus with a diameter of approximately 26nm and a genome size of about 4.7kb. The genome is flanked by 145-nucleotide Inverted Terminal Repeat (ITR) sequences at both ends. These ITRs function as replication and packaging signals. The genome contains two main Open Reading Frames (ORFs): rep and cap. The rep gene encodes four proteins (Rep78, Rep68, Rep52, and Rep40) involved in viral replication and regulation. The cap gene encodes three capsid proteins (VP1, VP2, and VP3) that form the icosahedral capsid structure. AAV also encodes the Assembly-Activating Protein (AAP), which facilitates viral particle assembly.

Structure of rAAV

After entering cells, AAV exists as free circular double-stranded DNA (dsDNA), maintaining stable gene expression without integrating into the host genome. Over 200 AAV variants have been identified to date, with 13 major serotypes (AAV1-AAV13) derived from primates. Different serotypes recognize receptors on different cell types. As of the first half of 2024, 8 AAV-based gene therapy products have received global regulatory approval:

Approved ProductCompanyIndicationSerotypeApproval Date
Beqvez Pfizer Hemophilia B AAVrh74var 2024.04.26
Elevidys Sarepta Duchenne Muscular Dystrophy AAVrh74 2023.06.22
Hemgenix CLSB Hemophilia B AAV5 2022.11.22
Roctavian BioMarin Hemophilia A AAV5 2023.06.29
Upstaza PTC AADC Deficiency AAV2 2022.07.18
Zolgensma AveXis Spinal Muscular Atrophy AAV9 2020.05.18
Luxturna Spark Inherited Retinal Disease AAV2 2018.11.22
Glybera uniQure Lipoprotein Lipase Deficiency AAV12 2012.10.02

AAV Research-Grade Services

Creative Biogene delivers industry-leading AAV vector solutions, driving advancements in gene therapy research and clinical development. Our comprehensive AAV platform integrates cutting-edge molecular biology, bioinformatics, and manufacturing systems to tackle key challenges in AAV vector development.

AAV Vector Design and Production Service

With diverse serotypes and engineered variants, Creative Biogene ensures precise targeting across tissues and species. Our expertise in vector design, promoter selection, and AAV optimization enables high-efficiency transduction and customizable production. We provide flexible design strategies and scalable manufacturing to support seamless transitions from early research to preclinical development.

Creative Biogene's diverse serotype portfolio includes conventional serotypes (AAV1-9) and engineered variants (e.g., AAV-DJ, AAV-PHP.eB), enabling precise targeting for neurological, hepatic, cardiac, and other tissue-specific applications. Our optimized serotypes ensure efficient transduction, with select variants effectively crossing the blood-brain barrier.

We also offer tissue-specific promoters for driving gene expression in targeted cells. These promoters are available for different species, including mouse, rat, and human, ensuring versatility across a wide range of experimental setups. From neuron-specific to ubiquitous promoters, we integrate enhancing elements to significantly improve target gene expression levels.

From standard single-stranded AAV (ssAAV) vectors to rapid-expression self-complementary AAV (scAAV) systems, our services are tailored to your specific insert size and expression requirements, with optimized delivery timelines to meet your research schedule.

How to Choose the AAV Type to Fits Your Needs?

Conventional serotypes

Engineered variants

Tissue-specific promoters

SerotypePrimary ReceptorSecondary ReceptorAffinity Tissues/Organs/Systems
AAV1Sialic acidAAVRSkeletal muscle, heart, mouse glial cells, ependymal cells, endothelial cells
AAV2HSPGFGF-R1, Integrin αVβ5/αVβ1, HGFR, LamRKidney, liver, retina, central nervous system
AAV3HSPGFGF-R1Human and non-human primate liver cells, mouse cochlear hair cells
AAV4Sialic acid—Liver, mammalian central nervous system, mouse kidney, lung, heart
AAV5Sialic acidPDGFRMouse retina, respiratory epithelial cells, liver, vascular endothelial cells, smooth muscle, non-human primate neurons
AAV6Sialic acid / HSPGEGFRCanine skeletal muscle, cardiac muscle, mouse respiratory epithelial cells, liver, skeletal muscle, cardiac muscle
AAV7——Mouse skeletal muscle, liver, central nervous system, non-human primate central nervous system
AAV8LamR—Canine liver, mouse skeletal muscle, heart, pancreas, kidney, liver
AAV9Integrin, LamRMouse central nervous system, retina, skeletal muscle, liver, pancreas, testes
SerotypePrimary Application TissueSerotype Description
AAV-DJBroad tissue applicabilityThis serotype is an optimized blend of 8 natural serotypes, showing effective infection in a wide range of human tissues and organs, with a strong hepatic cell tropism.
AAV-DJ/8 MyoAAVMuscle tissueDerived from AAV-DJ, this variant deletes the heparin-binding domain to reduce liver tropism interference, enhancing infection efficiency in non-hepatic cells with good specificity.
AAV-MGMicrogliaShows good specificity for microglial cells in the brain.
AAV-PHP.eBCentral nervous systemCrosses the blood-brain barrier, enhancing tropism for the central nervous system.
AAV-BI30Central nervous systemSpecifically and efficiently transduces endothelial cells throughout the central nervous system.
AAV-PHP.SPeripheral nervous systemEffectively infects the entire peripheral nervous system.
AAV2.7m8Retina, Inner earEffectively transduces retinal cells and cochlear hair cells in the inner ear.
AAV2-QuadYFRetina, Endothelial cellsEfficiently transduces retinal cells and vascular endothelial cells.
AAV2-retroSpinal nervesRetrograde transduction without crossing synapses.
SystemPromoterSpeciesTarget Cells
Nervous SystemTHRat, Mouse, HumanDopaminergic Neurons
hSynRat, Mouse, HumanNeurons
CaMKIIaRat, Mouse, HumanNeurons
ChATRat, Mouse, HumanCholinergic Neurons
TUBA1ARat, Mouse, HumanNeurons
GAD65Rat, Mouse, HumanGABAergic Neurons
GFAPRat, Mouse, HumanAstrocytes
MAGRat, MouseOligodendrocytes
NSEMouseMultiple Neurons
NesMouseNeural Stem Cells, Progenitor Cells
CnpMouseOligodendrocytes, Schwann Cells
Cardiovascular SystemTNNT2Mouse, HumanCardiomyocytes
Muscle SystemαMHCMouseCardiomyocytes
Hcn4MouseEmbryonic Cardiomyocytes
CD68 (short)Rat, Mouse, HumanMacrophages
F4/80Mouse, HumanMacrophages
Tie1MouseEndothelial Cells
TIE2Mouse, HumanEndothelial Cells
SM22aMouse, HumanVascular Smooth Muscle Cells
MHCK7MouseStriated Muscle Cells
MyogMouseMyogenic Cells
ACTA1MouseMyogenic Cells
LiverALBMouse, HumanHepatocytes
AFPMouse, HumanHepatocytes
TTRHumanHepatocytes
LungSP-CMouse, HumanAlveolar Type II Cells
Adipose TissueFABP4 (aP2)Mouse, HumanAdipocytes
RetinaRpe65Mouse, HumanRetinal Pigment Epithelium
hRHOMouse, HumanPhotoreceptor Cells
hBEST1MouseRetinal Pigment Epithelium Cells

Quality Control & Standards

1. Multi-dimensional Analytical Methods

Content Determination

Standardized protein quantification ensures AAV protein content meets specifications.

Impurity Detection

Monitor host cell proteins, DNA, endotoxins, and viral contaminants.

Physicochemical Properties

Assess pH, osmotic pressure, impurities, and particle size for stability.

Potency Testing

Perform genome titer (qPCR), capsid titer (ELISA), and biological activity tests.

Purity Analysis

Use SDS-PAGE and HPLC to evaluate capsid protein purity and empty capsid rates.

2. International Regulatory Standards

We have implemented a comprehensive quality management system aligned with global regulations, guaranteeing that our AAV vector products meet the stringent requirements for research and clinical applications. Our services comply with key regulations such as FDA 21 CFR 600-680, EMA/CAT/80183/2014, ICH guidelines, ISO 9001:2015, and NMPA guidelines.

How do We Support Your AAV Production for Research?

AAV Production Specification Table

Production TypeApplicationMinimum Titer (GC/mL)VolumeTurnaroundInquiry
Pilot-Scale ResearchCell culture>2×10¹¹100-500 uL6-12 daysInquiry
Medium-Scale ResearchCell culture & small animal studies>2×10¹¹500-1000 uL7-14 daysInquiry
Large-Scale ResearchAnimal studies & preclinical research>1×10¹2>1 mL14-21 daysInquiry

AAV Capsid Evolution and Screening Services

Creative Biogene's advanced AAV capsid engineering platform addresses critical industry challenges of limited tissue targeting and neutralizing antibody recognition in natural AAV serotypes. Through an integrated approach combining directed evolution with rational design, we develop highly diverse capsid libraries and conduct systematic screening to optimize three key vector attributes:

  • Enhanced tissue specificity
  • Improved immune evasion
  • Superior transduction efficiency

Our high-throughput platform can screen up to 10⁶ capsid variants simultaneously, supported by a comprehensive validation pipeline from murine models to non-human primates. Clients receive multidimensional analysis reports detailing targeting efficiency, immunogenicity profiles, and complete biodistribution data. Clinical outcomes demonstrate our technology's capacity to enhance liver targeting by 3-fold while reducing off-target tissue distribution by 60%, significantly expanding the therapeutic window for gene therapy applications.

AAV Biodistribution and Shedding Detection Services

To ensure full compliance with FDA and EMA regulatory requirements, our comprehensive AAV biodistribution and shedding detection services provide crucial insights into vector distribution patterns and environmental release risk assessment. Our analytical suite includes:

  • Precise vector genomic copy quantification in target and non-target organs using qPCR/ddPCR technologies
  • Longitudinal monitoring of vector shedding patterns across multiple biological matrices
  • High-resolution immunofluorescence staining for precise localization of transgene expression

We support diverse administration routes including intravenous, intracerebral, and intramuscular delivery, with cross-species validation capabilities spanning from rodent models to non-human primates. This comprehensive approach ensures complete regulatory-compliant data packages that seamlessly transition from preclinical animal studies to clinical applications.

AAV CDMO Services

Adeno-associated virus (AAV) vectors dominate gene therapy due to their safety and efficiency, with over 70% of gene therapies utilizing them. However, scalable, high-quality AAV production remains a key bottleneck as the market expands. Enhancing production capacity and quality control is crucial to meeting demand. As a professional CDMO, Creative Biogene has built a comprehensive AAV platform to tackle technical challenges in commercial production.

AAV High-Yield Platform

Creative Biogene offers industry-leading AAV vector production and purification services, utilizing third-generation AAV vector systems that ensure high titer and low empty capsid rates. Our technology platform combines advanced production processes and refined purification workflows to provide high-quality AAV vector solutions for clinical trials, and commercial applications.

Diverse Cell Line Options

Various 293 cell lines(293T, 293F, etc.) support early-stage research, development, and IND/BLA submissions.

Efficient Plasmid Optimization

Insertion of key regulatory elements into non-coding regions of multi-serotype RepCap plasmids boosts yields by 2 to 6 times.

Scalable Production

Suspension processes scale from SF125 to 200L, with upstream yields up to 1E16 vg and recovery exceeding 30%, meeting GMP requirements.

Strict Impurity Control

Optimized processes control impurities like AAV empty capsids, plasmid DNA, and HCD residues, enhancing product safety.

Creative Biogene offers tailored optimization strategies that span the entire AAV manufacturing workflow—from plasmid design to cell line adaptation, transfection efficiency, and purification consistency—ensuring high productivity, stability, and regulatory compliance.

Upstream process challenges and solutions

Downstream process challenges and solutions

Optimization FocusChallenges and DifficultiesCreative Biogene Solutions
GOI/RC/Helper TriplasmidTotal plasmid quantity and ratio impact titer and empty capsid rateSystematic plasmid ratio optimization to reduce empty capsid rate
Cell Line and MediaCell line selection, licensing costs, cell-media compatibilityIndependent development of high-yield cell lines and matched media
Virus Yield and Empty Capsid RateITR sequences, promoter, transgene, poly APlasmid backbone molecular optimization to enhance production
Cell Shear SensitivityHigh rotation speed, significant stirring paddle shear force, multiple bubbles, low viabilityOptimization of stirring and aeration strategies to reduce shear force
Transfection Complex MixingUneven incubation mixing, low transfection efficiencyStandardized transfection process to improve uniformity
Scale-up Process StabilityScaling strategies, equipment stability, viral production consistencyEstablishment of stable process scale-up parameters
Optimization FocusChallenges and DifficultiesCreative Biogene Solutions
Downstream RecoveryComplex processes, low recovery ratesOptimization of lysis, clarification, ultrafiltration, and chromatography processes
Production-Related Impurity ResidualsHost cell DNA, plasmid DNA, host cell proteins, ligands, nuclease residualsMulti-step purification processes for removing process-related impurities
Product-Related Impurity ResidualsIncomplete/mispackaged viral capsids, empty capsids, replication-competent AAVDevelopment of specific empty capsid removal technologies
Empty Capsid Removal and Scale-up StabilitySignificant empty capsid removal difficulties, poor stabilityIon exchange chromatography optimization to enhance empty capsid removal efficiency

AAV Master Seed and Formulation Quality Control

60+ test methods covering product-related impurities, process-related impurities, safety, infectious titer, target gene expression, characteristic sequence identification, purity, content, and potency.

CategoryDetection ItemsAnalytical MethodsStandards
IdentificationCapsid Protein IdentificationSDS-PAGE, CE-SDS, ELISAPositive (1:1:10)
Characteristic Sequence IdentificationFirst/Second-generation Sequencing, PCRConsistent with Theoretical Sequence
Quality AttributesGenomic TiterqPCR, ddPCRMeasured
Content and PotencyTCID50 or Alternative Infectious Titer Detection MethodsMeasured
Potency-Infectious TiterqPCR, ELISA, Western BlotPositive
Potency-Gene ExpressionCell-Based In Vitro Biological ActivityPositive
PurityProtein PuritySDS-PAGE, CE-SDS≥90%
Empty Capsid RateAUC (Gold Standard), AEX HPLC, Electron Microscopy≤10%
ImpuritiesHost Cell ProteinsELISA, HPLC-MS≤30ng/ml
Host Cell DNAqPCR, ddPCR<10ng/E12vg
Plasmid DNA ResidualsqPCR, ddPCR<10ng/E12vg
Affinity LigandsELISAInternal Standards
SafetyMycoplasmaCulture Method, qPCRNegative
Exogenous Viral FactorsIn Vitro Culture MethodNegative
EndotoxinLimulus Amebocyte Lysate Reagent<0.5EU/ml (Intrathecal Administration <0.2EU/(kg·h))
Replication-Competent AAVqPCR, Cell Culture Method≤1/10⁸ copies

Comprehensive Quality Management System

Creative Biogene implements a comprehensive quality management system covering personnel, process control, documentation, and risk management to ensure regulatory compliance and product consistency.

Personnel Management

  • Role Responsibility Documentation (SOP-QA-011)
  • Quality System Training Framework (SOP-QA-005)
  • Production Area Personnel Conduct (SOP-CM-001)

Process and Production Control

  • CQA/CMA/CPP Comprehensive Evaluation(SOP-QA-045)
  • Research Protocol and Report Management(SOP-PD-052)
  • Production Workflow Standardization(SOP-CM-013)

Quality Assurance

  • Data Integrity Assurance (SOP-QA-027)
  • Quality Risk Mitigation (SOP-QA-012)
  • Deviation Response Protocols (SOP-QA-013)
  • Change Control Mechanism (SOP-QA-014)
  • Corrective Action Management (SOP-QA-015)
  • Specification Deviation Investigation (SOP-QC-083)

Document Control

  • Document Lifecycle Management (SOP-QA-001)
  • Record Governance Framework (SOP-QA-007)
  • Archival System Protocols (SOP-QA-024)

Material Management

  • Supplier Quality Assurance (SOP-QA-017)
  • Material Categorization Protocols (SOP-QA-018)
  • Material Release Procedures (SOP-QA-019)

Facility and Equipment Management

  • Equipment Lifecycle Governance (SOP-EN-016)
  • Validation Management Protocols (SOP-VT-001)
  • Preventive Maintenance Strategy (SOP-EN-017)

Information Security Management System

Certified to ISO 27001, our information security system ensures data integrity and confidentiality through multi-level access control, real-time monitoring, and encrypted protection.

01

Network Security

  • Real-time Threat Monitoring
  • Redundant Data Protection Strategy
  • Firewall Defense Mechanism
  • Network Access Governance
  • Centralized Identity Authentication
  • Segmented Resource Isolation
03

Terminal Device Security

  • Unified Endpoint Management
  • Automated Critical Data Backup
  • Sensitive Data Encryption
  • Proactive Threat Mitigation
05

Information Security Management

  • Security Awareness Training
  • Legal Confidentiality Frameworks
  • Data Classification Protocols
  • Time-Sensitive
  • Information Control
02

Server Security

  • Intrusion Prevention Monitoring
  • Data Redundancy Preservation
  • Comprehensive Antivirus Defense
  • Emergency Remote Management
  • Granular Access Control
04

User Behavior Management

  • Role-Based Access Control
  • Sensitive Data Transmission Monitoring
  • Comprehensive Operational Logging
06

Compliance and Certification

  • ISO 27001 Information Security Standardization

How do We Support Your AAV Production at Every Scale?

AAV Production Specification Table

Production TypeApplicationMinimum Titer (GC/mL)VolumeTurnaroundInquiry
Pilot-Scale Ultra-PureCell culture & small animal studies>2×10¹²100-500 uL7-14 daysInquiry
Medium-Scale Ultra-PureAnimal studies & preclinical research>2×10¹²500-1000 uL7-14 daysInquiry
Large-Scale Ultra-PurePreclinical research & IND filing>1×10¹³>1 mL14-21 daysInquiry
Large-Scale cGMPCommercial production>2×10¹³CustomCustomInquiry

Comprehensive Support Services

To accelerate our clients' progress from experimental design to outcome conversion, we have established a full-dimensional support system:

1Expert Consultation: Virologists assist with serotype selection, promoter optimization, and animal model matching.

2Technical Guidance: Provide SOPs and guidance for complex injections, like stereotaxic and vitreous cavity injections.

3Real-Time Tracking: Clients can view cell culture data, purification progress, and quality reports through a digital platform.

4Automated Alerts: Email notifications for key milestones in the production process.

5Virus Activity Preservation: Offer solutions such as lyophilization protectant formulas.

6Efficacy Evaluation: Collaborate with partners to validate gene expression and monitor CAR-T cell dynamics in models.

7CMC Document Preparation: Support in creating CMC documents and designing stability studies.

8Faster Approval: Help expedite regulatory approval timelines.

Contact Us

Creative Biogene, with the core strategy of being "technology-driven and customer-centric," is dedicated to becoming a one-stop solution provider in the gene therapy field. Through continuous technological iterations, we help clients reduce research and development costs and accelerate clinical translation. Visit our website or contact customer service for exclusive technical solutions and limited-time offers!

FAQ

Q1: What factors affect AAV transduction efficiency?

A: AAV transduction efficiency is influenced by factors such as AAV serotype, injection method, infection period, and genome form. The AAV capsid characteristics affect its distribution in the body, and the choice of promoter influences expression patterns. Successful transduction depends on the virus's ability to bind to surface receptors, undergo endocytosis, and release its genome. Surface polysaccharides (e.g., sialic acid, galactose, heparan sulfate) and certain growth factor receptors and integrins are closely associated with AAV transduction.

Q2: How long does the expression period last after AAV virus injection?

A: AAV expression starts in animals within 1-2 weeks after injection and lasts for a long time. It is recommended to observe for the first time 3-4 weeks after infection in in vivo experiments. Expression duration may vary across different tissues and organs due to differences in cell division and renewal rates.

Q3: How to validate an effective interference target when constructing an interference AAV virus?

A: Validating effective interference targets can be done by synthesizing siRNA for the target gene, constructing shRNA plasmids, or packaging shRNA lentivirus to infect target cells. Alternatively, interference AAV virus can be packaged directly for in vivo validation. For target cell validation, transfection efficiency of siRNA and shRNA should be considered.

Q4: What is the maximum length of the target gene that can be constructed in an AAV virus?

A: The size of the target gene an AAV can carry depends on the vector elements, including the necessary promoter, target fragment, and fluorescent or tag elements. For overexpression AAV vectors, the external gene fragment should typically not exceed 2.8 kb; the best loading capacity for rAAV is under 5 kb. Interference sequences are generally short and are typically not restricted by vector capacity.

Q5: How to use AAV for in vitro and in vivo applications?

  • In vitro: Calculate virus dosage using Multiplicity of Infection (MOI) where MOI = required viral particles / number of infected cells. Generally, most cells require 1,000-10,000 MOI, with some needing up to 500,000 MOI. Adding 0.8 µM of camptothecin for 4 hours before infection can improve transduction efficiency.
  • In vivo: Select appropriate AAV serotype and injection method based on the tissue type. Common injection methods include tail vein injection, intrathecal injection, and cardiac injection, each requiring different viral dosages to ensure sufficient viral distribution and expression.

Q6: How to ensure effective animal experiments with AAV, which is more suitable than lentivirus or adenovirus?

A: Virus titer is crucial for animal experiments, as it determines the number of viral particles delivered to the tissues and the maximum volume. Referencing literature for the target tissue can be helpful, but pre-experiments should determine the optimal dosage. Factors like animal age, observation time, and injection method also influence experimental outcomes.

Q7: What is the difference between scAAV (double-stranded AAV) and ssAAV (single-stranded AAV)?

A: Single-stranded AAV must convert to double-stranded form inside the cell for gene transcription and translation, a process that is slow and stable expression typically occurs 2-4 weeks after injection. Double-stranded AAV has 6-15 times higher infection efficiency, with detectable expression within 3 days after injection, but has a smaller vector capacity.

Q8: What does the AAV titer unit vg/mL represent? What methods are used to detect AAV virus titer?

A: vg/mL stands for Vector Genomes/mL, equivalent to Genomic Copies per mL (gc/mL), representing the number of genome copies per mL of viral solution. RT-PCR is typically used to measure virus titer.

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