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Rabbit Monoclonal Antibody Production Services

Rabbit Monoclonal Antibody Development, Custom Rabbit Monoclonal Antibody Production

Catalog No. Product Name Size List Price (US$) Quantity
BS026 Custom Rabbit Monoclonal Antibody Production 1 package Request
Description

Syd Labs has successfully developed rabbit monoclonal antibodies using antigen specific single B cell-based antibody generation technologies. After identification and isolation of antigen specific memory B cells by flow cytometry-based single B cell sorting, the antibody variable regions (VH and VL) from single or clonal B cells are cloned and sequenced. Recombinant rabbit monoclonal antibodies are produced, purified, and tested for various applications, such as ELISA, Western blot, IP, IF, ICC, and IHC. 

The most comprehensive custom rabbit monoclonal antibody production services: custom production of rabbit monoclonal antibodies against non-modified, phospho-specific, methylation-specific, and acetylation-specific peptides and proteins; rabbit monoclonal antibodies for ELISA, Western blot, IP, IF, ICC, and IHC applications. Syd Labs team has successfully developed a large number of high specificity and high affinity rabbit monoclonal antibodies against a wide range of difficult targets such as small molecules, antibody idiotype, and antigens non-immunogenic in murine, camel or Ilama.

BS026: Custom Rabbit Monoclonal Antibody Production Service

Phase I: Immunization and boosts of 2 rabbits following the standard immunization protocol and ELISA screening of rabbit polyclonal sera. The animal positive respone to the antigen is critical to isolate high specificity and high affinity rabbit monoclonal antibodies. For some difficult targets, more rabbits and longer immunization are required.

Phase II: PBMC or splenocyte isolation from selected rabbit(s), detection and isolation of antigen specific memory B cells by flow cytometry-based single B cell sorting, culturing single B cells in individual wells to secret antibodies, high throughput ELISA screening to isolate antigen-positive B cells. Supernatant (culture media) from ELISA-positive clones is sent to clients for evaluation of specific applications. If clients have their own assays of antibody screening, supernatant from 96-well plates is sent without ELISA test.

Phase III: Cloning and sequencing the antibody variable regions (VH and VL) from antigen specific B cells selected by clients, and transient production of recombinant rabbit monoclonal antibodies in HEK 293 cells. Purified rabbit monoclonal antibodies are sent to clients for confirmation and test of specific applications.

You provide: Ten mg of peptide (purity > 85%, length > 12 amino acids) or 4 mg of soluble protein (concentration > 0.4 mg/ml, purity > 90%, molecular weight > 20 kDa) for immunization, and (optional) additional 5 mg for antigen affinity purification.

Deliverable:
* About 80 ul supernatant from ELISA positive cells for evaluation;
* About 100 ug purified rabbit recombinant monoclonal antibodies for evaluation;
* Amplified DNA fragments;
* Remaining antigens;
* Study report including ELISA results and antibody VH and VL sequences.

Rabbit antibody humanization service is available if requested.

Questions and Answers:

Question: Other companies use hybridoma technologies to develop rabbit monoclonal antibodies. What is advantage of your single B cell-based antibody technology?
Answer: The number of rabbit hybridoma cells for antibody screening from one animal is limited. It is also common for mouse hybridoma generation. Syd Labs can normally sort >10000 single B cells for culturing and screening. Up to 60% single B cells can be ELISA positive. In general, success rate of single B cell-based method of custom monoclonal antibody development is higher than hybridoma-based method, especially for those difficult to work targets. In addition, the companies that provide rabbit hybridoma generation services ask for royalty in addition to service fee. We charge similar amount of service fee to theirs but we do not ask for royalty fee. Our program of fee for service is welcome by diagnostics companies.

Question: Other companies use phage display technologies to screen rabbit monoclonal antibodies. What is advantage of your single B cell-based rabbit monoclonal antibody production protocol?
Answer: In general, the cost of phage display library construction and screening is higher than single B cell-based method. More importantly, clients can choose to sort and culture ~800, 1200, 1600, 2000 or more single B cells. The basic fee for ~800 single B cells is low compared to that for phage display library construction and screening. In addition, the pair(s) of VH and VL in the scFv-Fc library may not come from the nature; a large library capacity (10^9 and even higher) may come from a small number of cells, 10^5, most of which do not bind to the antigen at all.

Question: What is advantage of rabbit monoclonal antibodies to mouse monoclonal antibodies?
Answer: Compared with mouse antibodies, rabbit antibodies have 10 – 100 times higher affinity, in the range of 1 - 100 pM;  superior specificity in assays, such as improved assay sensitivity, increased assay accuracy with decreased false positive rate, lower assay noise on IHC with paraffin-embedded tissue samples; broader range of epitopes with less abundance and difficult epitopes, such as protein modifications, enzyme cleavage sites, or conformational epitopes, small molecules and haptens, including small molecule drugs, steroid hormones, lipids, and glycolipids; great stability and longer shelf life.

Question: What do I need to provide if I want to develop a rabbit monoclonal antibody against a small molecule chemical?
Answer: You need to have the molecule conjugated to KLH for animal immunization and biotin for single B cell sorting and antibody screening.

Question: Do you do DNA immunization for rabbit monoclonal antibody development? Or cells expressing a protein on membrane?
Answer: Yes for both.

Question: What is the turnaround time for a rabbit monoclonal antibody development project?
Answer: On average 18 weeks.

Syd Labs also provides other monoclonal antibody development services:

Human and monkey monoclonal antibody development services
Mouse monoclonal antibody development services
Rat monoclonal antibody development services
Camelid monoclonal antibody development services

References:

A Robust High Throughput Platform to Generate Functional Recombinant Monoclonal Antibodies Using Rabbit B Cells from Peripheral Blood. Seeber S, Ros F, Thorey I, Tiefenthaler G, Kaluza K, Lifke V, Fischer JA, Klostermann S, Endl J, Kopetzki E, Pashine A, Siewe B, Kaluza B, Platzer J, Offner S. PLoS One. 2014 Feb 4;9(2):e86184.

Generation of Recombinant Monoclonal Antibodies from Immunised Mice and Rabbits via Flow Cytometry and Sorting of Antigen-Specific IgG+ Memory B Cells. Starkie DO, Compson JE, Rapecki S, Lightwood DJ. PLoS One. 2016 Mar 29;11(3):e0152282.

From rabbit antibody repertoires to rabbit monoclonal antibodies. Weber J, Peng H, Rader C. Exp Mol Med. 2017 Mar 24;49(3):e305.

Stable long-term cultures of self-renewing B cells and their applications. Kwakkenbos MJ, van Helden PM, Beaumont T, Spits H. Immunol Rev. 2016 Mar;270(1):65-77.

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