With CRO as the core business, Zoonbio has been working in the field of antibody discovery for many years, with mature and perfect technology. In 2012, the company established Alpaca single domain antibody research center and launched nano antibody development service in 2014. Zoonbio can provide one-stop services such as antigen preparation, alpaca immunity, PBMC separation, cDNA preparation, antibody library construction and screening, nanobody expression and purification, antibody humanization, etc.

nano antibody

nano antibody

nano antibody

You can sample at the breeding base, supervise immunization and check the status of experimental animals in real time.

Nanobody Service Steps and Delivery Standards

Step Cycle Delivery Materials
1.Animal immunity
A.5 times of animal immunity
B.Titer detection
C.PBMC separation
D.RNA extraction
E.cDNA preparation
92 days 5 times of immunopotency test data
cDNA after 5 immunization
2.Construction of phage antibody library
A.VHH fragments of the cDNA prepared by PBMC were amplified to construct bacterial library and phage library and determine titer.

B.Antibody library quality:
(1)The insertion rate was more than 95%. 48 clones were randomly selected and sequenced without repeat sequence
(2)The capacity of antibody library reaches 109
4-6 weeks Quality inspection report
Antibody library
3.Antibody library screening
A.According to the actual situation of the project, select different screening schemes, such as competitive screening, solid-phase screening, liquid-phase screening, negative screening and other screening methods.

B.After each round of screening, 24 clones were selected for identification.
6-8 weeks more than 5 specific clones
4.Expression and purification of nanbody
A.The nanbody gene was cloned into eukaryotic or prokaryotic expression vector. 2 weeks High purity nanbody 100-500ug
5.Affinity determination
Determination of antibody affinity by Biacore/
Determination of antibody affinity by Nicoya LSPR
Determination of antibody affinity by Fortebio octet
1 week
1 week
1 week
Affinity measurement data
6.Humanization of nanbody(Analysis of antibody variable region and humanization design of antibody by self-developed software absys.)
A.Identify FR, CDR, key residues, PTMs

B.Through homology comparison, the consistency and similarity of variable region, key residues, PTMs, stability and other factors are considered,the optimal human antibody germline FR was selected as the template, and recombined with Alpaca CDR to design the initial humanized antibody.

C.Analyze the sequence and structure of the antibody, determine the key residues that need to reverse mutation, and design different numbers of reverse mutation VH.

D.The humanized gene was cloned into eukaryotic expression vector and transferred to 293 for transient expression to obtain 100-500ug purified antibody.

E.Binding activity and affinity test.
6-8 weeks At least one humanized antibody(The affinity and expression amount are similar to that of the original nanbody)

Related services:

Recombinant Antibody Expression Service

Western Blotting Service

Antibody Modification Technology Service