Background: Compared with traditional antibodies, camel derived antibodies with higher affinity, are, easy to be recombined, expressed, humanized and combined with other target antibodies.。 They have great application value and development prospects in disease diagnosis and treatment. Sanyou proudly launched MIT Alpaca monoclonal antibody customized service with all above functions.
Methods: The preparation technology of MIT Alpaca monoclonal antibody integrated four technologies: Alpaca immunity, construction of array phage antibody library, high-throughput screening of magnetic beads and recombinant expression of CHO eukaryotic cells, and matched with comprehensive physical, chemical and biochemical analysis for screening.
Advantages: Alpaca monoclonal antibodies with good specificity, high affinity, clear sequence and verified by eukaryotic expression can be obtained in only 3-6 weeks (from the end of Alpaca immunization).
Cases: Since the launch of the service in 2016, by December 2021, Sanyou biology has completed hundreds of projects through this platform; For each target, 40-60 lead antibody clones with unique sequence, good specificity and verified affinity can be obtained.
Service Overviews
Background: Compared with traditional antibodies, camel derived antibodies with higher affinity, are,easy to be recombined, expressed, humanized and combined with other target antibodies.。 They have great application value and development prospects in disease diagnosis and treatment. Sanyou proudly launched MIT Alpaca monoclonal antibody customized service with all above functions.
Methods: The preparation technology of MIT Alpaca monoclonal antibody integrated four technologies: Alpaca immunity, construction of array phage antibody library, high-throughput screening of magnetic beads and recombinant expression of CHO eukaryotic cells, and matched with comprehensive physical, chemical and biochemical analysis for screening.
Advantages: Alpaca monoclonal antibodies with good specificity, high affinity, clear sequence and verified by eukaryotic expression can be obtained in only 3-6 weeks (from the end of Alpaca immunization).
Cases: Since the launch of the service in 2016, by December 2021, Sanyou biology has completed hundreds of projects through this platform; For each target, 40-60 lead antibody clones with unique sequence, good specificity and verified affinity can be obtained.
Service Contents
Service name
Service Details
Client Provides
Deliverables and Standards
Time
Customized service of MIT Alpaca monoclonal antibody
Alpaca immunity
Construction of immune Library
High throughput screening Eukaryotic expression verification
Target name, or target antigen, or cell line
Deliverables:
1. Dozens of lead antibody molecules
2. Provide antibody samples as required
Standards:
1. Antibody affinity ranking
2. The preferred molecule is verified by affinity kinetics or FACS
About 4 months
Humanization of Camel derived antibody
Humanized design Eukaryotic expression verification
Camel derived antibody sequence, target protein and control antibody,
identification cell line, etc
Humanized sequence, protein, plasmid and test report
4-6 weeks
Service Highlights
1. Clear Antibody Sequence and Reduce Redundant Screening Work
This service utilizes genetic engineering and phage display technology to obtain thegene sequence of antibody directly, so as to reduce unnecessary screening work.
2. Deliver Tens to Hundreds of Lead Antibody Molecules
This service is adopting the multi group primer array library construction method and multi type screening library strategy, and dozens to hundreds of candidate molecules can be obtained from one target.
3. Full Coverage of Difficult Targets and Hidden Epitopes
A large number of antibody molecules can be obtained for difficult targets such as G protein coupled receptors, ion channels and multiple transmembrane proteins, covering most of the protein epitopes.
4. Eukaryotic Expression, Cell Level Verification, Ensuring the Druggability of Delivered Molecules
The molecules obtained from this service will be constructed and expressred at the eukaryotic level, and the physical and chemical properties and cell functions of the antibody will be comprehensively tested to ensure that the delivered lead antibody has good druggability.
Service Features
1. Dozens of Lead Antibodies
A large number of nano antibody lead molecules can be obtained through the customized service of Sanyou MIT Alpaca monoclonal antibody. Based on the verification of MIT Alpaca library with 10 targets, 475 antibody clones with unique sequences were obtained, and the median number of clones was 38.
Fig. 1 Statistics of Binder amount/project
2. High Affinity of Lead Antibody
The nanoantibody candidate molecules screened by this service have high affinity and can reach the pM level. The affinity testing of the full-length IgG constructed from the single domain antibody clone is as follows. The antibodies in Fig. 2 and Fig. 3 come from two different projects. The figures show that multiple molecules obtained from these two projects have significantly better or equivalent affinity than the control.
Fig. 2 Affinity ranking assay of SD10
Fig. 3 Affinity ranking assay of SD41
3. Parameters Related to Drug Developability Assessment
After the full-length molecular construction obtained from this service, theexpression amount of the antibody and the physical/chemical properties of the antibody were analyzed. The following Table shows a comprehensive evaluation system of drug developability assessment, including purity and concentration determination, primary structure analysis, affinity, and affinity kinetics.
Table 1 Drugdevelopability of Antibody from MITA
Category
Test content
Test method
Purity and concentration Testing
Purity identification
SDS-PAGE/SEC/CE-SDS
Purity and concentration Testing
Concentration identification
Protein A-HPLC/UV280
Primary structure analysis
Molecular weight analysis
LC/MS
Primary structure analysis
Electric point
iCIEF
Primary structure analysis
Hydrophobicity identification
HIC-HPLC
Primary structure analysis
Charge heterogeneity measurement
CEX
Primary structure analysis
Peptide map analysis
LC-UV-MS/MS
Primary structure analysis
N-sugar spectrum analysis
LC/MS
Affinity and Affinity Kinetics
Affinity detection
ELISA
Affinity and Affinity Kinetics
Affinity kinetic detection
BLI/SPR
Affinity and Affinity Kinetics
Cell binding test (if any)
FACS
Case Stastics
1. Cell Level Binding Analysis
Most of the nano antibody candidates screened throuhg this platform have cell level activity and blocking activity. Fig. 4 shows a case project using FACS to analyze the binding activity of antibodies to cell surface proteins. It can be seen that all lead antibody molecules show good cell level binding activity.
Fig. 4 Binding affinity determination by FACS
2. Validation of Efficacy in Vitro
The nanoantibody candidate molecules obtained through this service platform exhibits excellent efficacy in vitro, even better than the control antibody. In MLR experiment, CD4+T cells and DC cells with different donors were co-cultured. Anti-PD-L1 antibody can effectively block PD1/PD-L1 inhibition signal, activate T cells and stimulate the secretion of cytokines. As shown in Fig. 5, after treatment with PD-L1 candidate antibodies (A1, A2), the secretion of IL-2 increased significantly, indicating.
’
Fig. 5 In vitro evaluation of SD10 sdAb candidates by MLR
3. Validation of Efficacy in Vivo
The nano antibody candidate molecules screened through this service platform reflect excellent efficacy in vivo. The in vivo antitumor efficacy verification cases of target candidate molecules in different mouse CDX models are shown in Fig. 6, Fig. 7 and Fig. 8. The treatment group and control group were administered twice a week for three consecutive weeks. The results showed that each target candidate antibody showed the same antitumor effect as the control antibody at each dose, even better than the positive control.
Fig. 6 Tumor growth inhibition
Fig. 7 Tumor growth inhibition
Fig. 8 Tumor growth inhibition
4. The Progress of Representative Projects
Table 2 The Progress of Representative
Item No
Target
Indication
Project Difficulties
Molecular Form
Library Source
Progress
SYZJ002
NCOV
Novel coronavirus
Broad spectrum
neutralization activity
Double antibody
Alpaca immune bank
In IND
SY11-sAb
CL18
Digestive tract tumor
Multiple cross touch
Nano antibody
Alpaca immune bank
In IND
SYDX P003
PL1/H2/VF
Digestive tract tumor
ADCC
Tertiary antibody
Alpaca immune bank +Alpaca immune bank +Biosimilars
Production cell line
SYDX-B019
TR2/C4
Breast cancer,
lung cancer
Liver toxicity TNFR2
Double antibody
Alpaca immune bank
+ Alpaca immune bank
In vitro efficacy
SYDX B020
TR2/C18.2
Solid tumor
Liver toxicity TNFR2
Double antibody
Alpaca immune bank +Alpaca immune bank
In vitro efficacy
SYDX-B022
TR2/PL1
Solid tumor
Dual immunosuppression relief
Double antibody
Alpaca immune bank +Alpaca immune bank
In vitro efficacy
LK003
undisclosed
Solid tumor
Immune regulation related targets
Full length antibody
Alpaca immune bank
In vitro efficacy
SYDX B014
I17A/F
Rheumatoid arthritis
Neutralizing bispecific antibody
Double antibody
Alpaca immune bank +Alpaca immune bank
In vitro efficacy
DJ001
undisclosed
AMD,DME
Blocking antibody
Full length antibody
Alpaca immune bank
In vitro efficacy
DJ002
undisclosed
AMD,DME
Blocking antibody
Full length antibody
Alpaca immune bank
In vitro efficacy