Custom Antibody
Services
Custom service offerings
• DNA sequence
• Plasmid
• Peptide
• Structure
• Protein
• Peptide
• Small molecule
• Cells
Antigen
Preparation
Immunization
Antigen and
screening materials
Antisera,
Purifled polyclonal
antibody
- Core service
- Input
- Additional service
- Output
• Spleen
• Lymphocytes
Fusion &
Screening
Subcloning
Expansion &
Delivery
Production &
Purification
Supernatants
Supernatants
Frozen cells
Supernatants,
Purified
monoclonal
antibody
Screening
Assays
Screening
Assays
Antibody
Cloning &
Engineering
Assay
Development
Immunogen design and preparation
• Peptide design, synthesis, and conjugation
• cDNA cloning and recombinant protein production in bacterial or mammalian systems
Hybridoma development
• Fusion and screening of supernatants by ELISA
• Differential screening for phospho-, point mutation-, cleavage- or isoform-specific projects
RabMAb production
• Small scale (<10 mg): hybridoma/Integra
• Mid scale (10 to 500 mg): recombinant/Transient
• Large scale (500 mg or more): recombinant/CHO stable cell lines
Stable cell line development
• Heavy and light chain cloning into single, proprietary vector
• Expression in suspension CHO cells
• Multiple rounds of subcloning for optimized production
Polyclonal antibody services
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Immunization strategy design
Antisera production and titer testing
Affinity purification and/or affinity depletion
Optional: Spleen harvest for RabMAb development
Humanization
• Novel patented humanization method
• Other humanization technologies also available
Assay development
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•
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Sandwich and competition ELISAs
Receptor-ligand binding, phosphorylation and functional cell-based assay
Tissue microarray IHC profiling
PK, PD, antigenicity assays
Why a custom rabbit monoclonal antibody
from Abcam?
Abcam’s custom antibody service utilizes RabMAb® technology, Abcam’s patented antibody technology for the development high quality rabbit monoclonals. Benefits of Abcam’s rabbit monoclonal antibody service includes:
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Novel epitope recognition
Picomolar affinity
High specificity
Excellent IHC staining results
Detection of subtle changes such as post-translational modifications
Human protein and mouse ortholog cross-reactivity
Maintain continuity in your antibody projects
Comparison of rabbit and
mouse monoclonal antibodies
Comparison
Antigen recognition
Rabbit monoclonals
Mouse monoclonals
• High success rates with a variety of
antigens, including small molecules
and peptides
• Small molecules and peptides often
non-immunogenic
• Good responses to rodent antigens
• Broad epitope recognition
• Less responsive to rodent antigens
• Limited epitope recognition
Affinity
Picomolar (KD = 10-12 M) possible
Typically nanomolar (KD = 10-9 M) or lower
Specificity
Higher
Lower
IHC / ICC usage
Excellent results in IHC and ICC
Not always suitable for IHC or ICC
CDX2
Paraffin-embedded human
colonic carcinoma tissue
stained with CDX2 rabbit monoclonal (ab76541) and Vendor
A’s CDX2 mouse monoclonal
(both at 1:1000 dilution factor)
Rabbit MAb
Mouse MAb
Rabbit MAb
Mouse MAb
Rabbit MAb
Mouse MAb
E-Cadherin
Paraffin-embedded human
breast carcinoma tissue
stained with E-Cadherin rabbit
monoclonal (ab40772) and
Vendor A’s E-Cadherin mouse
monoclonal (both at 1:50
dilution factor)
HER2 / ErbB2
Paraffin-embedded human
breast carcinoma tissue stained
with HER2 / ErbB2 rabbit monoclonal (ab134182) and Vendor
A’s HER2 / ErbB2 mouse
monoclonal (both at 1:500
dilution factor)
Antibody generation process
Polyclonal
Polyclonal
Antigen
Immunization
Polyclonal antisera
Purified polyclonal
antibodies
Detection
polyclonalresponse. The collected
Immunization
Polyclonal Antigen
antibodies: A rabbit
is injected withPolyclonal
antigenantisera
and mountsPurified
an immune
Detection antisera
antibodies
contain a mixture of antibodies and other components. The antibodies are
typically affinity-purified, and they recognize
multiple epitopes on the antigen.
Monoclonal
+
Monoclonal
Antigen
Immunization
Lymphocytes
+ Partner cells
Fusion mixture
Antigen
Immunization
Lymphocytes
Partner cells
Fusion mixture
Frozen hybridomas
Subclones
Multiclones
Frozen hybridomas
Subclones
Multiclones
Production
Purified monoclonal
antibodies
Detection
Production
Purified monoclonal
antibodies
Detection
Monoclonal antibodies: Following immunization with the antigen of choice, lymphocytes are isolated from the rabbit’s
spleen. Fusion with a myeloma cell line results in immortal hybridomas that secrete antibodies. These cells are plated, at
which point many wells contain multiple cell types or multiclones. The supernatants are then screened for the presence
of specific antibodies, and the best multiclones are diluted and replated to isolate single lines or subclones. After further
screening, the subclones are expanded and frozen for delivery. Monoclonal antibodies, which recognize a single epitope
on the antigen, can then be produced in bulk by the hybridomas or by exogenous cell lines.
Antibody generation timeline
2
1
4
3
Go/No-go
Go/No-go
Go/No-go
6
5
8
7
Month
Immunization
Order
Customer
Testing
Invoice
Fusion &
Screening
Customer
Testing
Subcloning
Invoice
Customer
Testing
Invoice
Expansion
& Delivery
Invoice
Polyclonal - 3 months
Monoclonal - 7 to 8 months
Legend:
- Process stages
- You may choose to proceed with the service (Go) or end it (No-go)
if you are already satisfied at this point
- Abcam sends processed antibody for customer testing
- You receive an invoice from us at these stages
“
“
In the last 6 years we have developed a number of very useful
rabbit monoclonal antibodies. For our research work such
sensitive and specific tools had not been available before.
Without the project planning and management of the group in
Vienna and without the excellent scientists in California this
success story would not have been possible.
- Dr. Pidder Jansen-Duerr,
Institute for Biomedical Ageing Research of the Austrian Academy of Sciences
Email: service@abcam.com
USA +1 (888) 772-2226
Canada +1 (877) 749-8807
Germany, Austria, and Switzerland
+43 (0) 2242 31006
UK and rest of Europe
+44 (0) 1223 696000
Japan
+81 (0)3 6231 0940
Hong Kong
(852) 2603-6823
China
+86 21 6129 2488
400 6896 692 (toll-free)
014_14_DG
Rest of world
+1 (888) 772-2226
Discover more at abcam.com/custom
Copyright
Our RabMAb technology is covered under the following U.S. Patents, No. 5,675,063 and 7,429,487