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Location: Home > Custom Services > Antibody Discovery Services > Phage Display & Antibody Library Services

Phage Display & Antibody Library Services

Date: 2018-10-22 Author: Leading Biology Click: 1721

Phage Display Service

M13 single strand filamentous phage display system is the most widely used library display system. The phage display library is formed by fusing the protein, antibody or polypeptide with phage pIII protein to participate in phage package and display on the phage surface. Using phage display library to make 3-5 rounds of affinity screening for specific target molecules, phage that specifically binds to the target molecule could be enriched to obtain corresponding DNA sequence information.


Our current phage display technology service platform covers frequently-used library types such as immune antibody library and natural antibody library. We strive for fulfilling individual customer's needs by customizing a variety of library and screening services to fit the individual preference with flexibility and efficience.


Phage Display Service Profile

 

M13 single strand filamentous phage display system is the most widely used library display system. The phage display library is formed by fusing the protein, antibody or polypeptide with phage pIII protein to participate in phage package and display on the phage surface. Using phage display library to make 3-5 rounds of affinity screening for specific target molecules, phage that specifically binds to the target molecule could be enriched to obtain corresponding DNA sequence information.


Our current phage display technology service platform covers frequently-used library types such as immune antibody library and natural antibody library. We strive for fulfilling individual customer's needs by customizing a variety of library and screening services to fit the individual preference with flexibility and efficience.






Leading Biology platform system mainly includes pComb3 phage display system and pCANTAB5e phage display system.


pComb3 Phage Display Technology



(1) Obtain the gene library: Collect the immune cells from the target antibody host and extract the total RNA of the cells. Amplify VH-CH1 antibody gene fragment and VL-CL1 antibody gene fragment respectively with multiple pairs of primers after reverse transcription.

(2) Transform E.coli with antibody gene library: The amplified antibody gene library was ligated to pComb3 plasmid, and then electrotransform E.coli XL12 Blue. Identify the capacity of gene library and the insert rate of antibody gene.

(3) Phage Fab antibody library construction: The helper phage VCSM13 was added to E.coli which was transferred into antibody gene and grown to logarithmic phase in order to rescue the phage. After overnight culture, collect the supernatant by centrifugation and get the phage Fab antibody library.

(4) Antibody library titer determination: Gradient dilute the phage Fab library obtained above to perform plaque culture experiment. Observe the growth of plaque to determine the titer of phage Fab antibody library.

(5) Enrichment screening of phage antibody library: Add phage antibody to the target antigen coated enzyme-linked plate to incubate, then follow by washing and eluting. Neutralize the eluted phage antibody, infect the bacteria again and carry out the second round of elutriation. Three rounds of elutriation would be conducted and titer was measured after each round of elution. During the last screening, set an BSA-coated control group to determine whether a large amount of anti-BSA phage antibody was enriched.

Fig 1. Process of pComb3 phage Display Technology




Fig 2. Genetic Map of Phagemid Vector pComb3



Fig 3. Genetic Map of Helper Phage VCSM13

pCANTAB5e Phage Display Technology

(1) Obtain the ScFv gene library: Collect the immune cells from the target antibody host and extract the total RNA of the cells. Amplify VH antibody gene fragment and VL antibody gene fragment respectively with multiple pairs of primers after reverse transcription. Randomly splice the full set of VH and VL gene into ScFv gene library with Linker primers after VH and VL fragment purification and recycling.

(2) Transform E.coli with ScFv gene library: The full set ScFv gene fragment was ligated to pCANTAB5e vector, and then electrotransform E.coli TG1. Identify the capacity of ScFv gene library and the insert rate of gene.

(3) Phage ScFv antibody library construction: The helper phage M13K07 was added to E.coli which was transferred into ScFv antibody gene and grown to logarithmic phase in order to rescue the phage.After overnight culture, collect the supernatant by centrifugation and get the phage ScFv antibody library.

(4) Antibody library titer determination: Gradient dilute the phage ScFv antibody library obtained above to perform plaque culture experiment. Observe the growth of plaque to determine the titer of phage ScFv antibody library.

(5) Enrichment screening of phage antibody library: Add phage antibody to the target antigen coated enzyme-linked plate to incubate, then follow by washing and eluting. Neutralize the eluted phage antibody, infect the bacteria again and carry out the second round of elutriation. Three rounds of elutriation would be conducted and titer was measured after each round of elution. During the last screening, set an BSA-coated control group to determine whether a large amount of anti-BSA phage antibody was enriched.



Fig 4. Process of pCANTAB5e phage Display Technology



Fig 5. Genetic Map of Phagemid Vector pCANTAB5e



Fig 6. Genetic Map of Helper Phage M13K07


Phage Display Technology Service Items

(Phage Display Antibody Library Construction)

Service Description

Service Period

Price

cDNA Synthesis

Collect the immune cells, extract total RNA and synthesize cDNA via reverse transcription.

1 week

Inquiry

Plasmid Construction

Amplify the antibody gene fragment through PCR with cDNA above as a template, and ligat into vector (such as pComb3 or pCANTAB5e vector)

4 weeks

Transformation

Electrotransform constructed vector into E. coli strain.

1 week

Library Capacity Identification

(1) Gradient dilute the transformed strain after recovery. Spread on the plate, then calculate the number of positive clone.

(2) Choose 10 positive single clones, extract the plasmid and verify through enzyme digestion.

(3) Calculate the capacity of antibody library.

Antibody Library Production

Obtain the phage display antibody library via helper phage rescue.

1 week

Shipping

Low temperature (Dry ice)

N/A

Total: Inquiry

2-3 months

Note: Clients need to pay 80% advance payment after the project starts.

Phage Display Library Screening


Our current phage display technology service platform covers commonly used library types such as immune antibody library and natural antibody library. We will provide flexible and efficient screening services according to the self-own phage library products chosen by clients or various personalized library provided by Leading Biology.

 

The following experiment procedures would be carried out according to the samples provided by clients:

(1) Antibody library titer determination: Gradient dilute the phage ScFv antibody library obtained above to perform plaque culture experiment. Observe the growth of plaque to determine the titer of phage ScFv antibody library.

(2) Enrichment screening of phage antibody library: Add phage antibody to the target antigen coated enzyme-linked plate to incubate, then follow by washing and eluting. Neutralize the eluted phage antibody, infect the bacteria again and carry out the second round of elutriation. Three rounds of elutriation would be conducted and titer was measured after each round of elution. During the last screening, set an BSA-coated control group to determine whether a large amount of anti-BSA phage antibody was enriched.


Service Description

Service Period

Price

Antigen Analysis

Detect antigen quality by SDS-PAGE or other methods.

1 week

Inquiry

Fab/ScFv Antibody Library preparation in Advance

Source from humans or mice with a capacity of 108-109 (Provided by Leading Biology or clients).

1 week

Antibody Screening

Elutriate for 4 rounds. Antibody affinity exceeds 10-7.

2 weeks

Conjugated Phage Analysis (About 40 Monoclones)

Phage amplification.

1 week

ELISA detection.

Phage DNA extraction.

Phage DNA sequencing.

Antibody Analysis (5-10 Monoclones)

Choose 5-10 strains of phage with different sequence from the sequenced conjugated phages above to carry out antibody soluble expression and ELISA identification.

1 week

Delivery

Test report (Sequencing result, etc). Antibody gene fragment cDNA.

N/A

Total

Inquiry

2-3 months

Note: Clients need to pay 80% advance payment before each project starts; Pay the rest after the project is completed.

Table 2. Phage Display Technology Service Items: Antibody Screening


Related Services

Premade Phage Display Antibody Libraries

Phage Display Library Construction

Phage Display & Antibody Library Services


Why Leading Biology?

With over 5 years of experience in custom antibodies, we offers numerous benefits for developing your custom antibody:


• Our industry leading titer guarantees of 1:50,000 eliminate the risk of not obtaining antibodies against peptide antigens.

• Our economies of scale allow us to pass cost savings to you and help maximize your budget.

• 100% of services are transparent throughout.

• Antibodies from our facility have been cited in many published research papers.

• By outsourcing antibody production and characterization needs to us, you can spend more time focused on your research.

• Our ideal location in California ensures that animals benefit from outdoor facilities and mild year-round temperatures.

• Confidentiality is a top priority for all projects. Antibodies that we manufacture belong to you and will not be commercialized.


Contact Information

Please obtain a quote before ordering, and refer to the quote number when you place an order.

Orders are typically confirmed within 12 hours.

Have a Question? Email us info@leadingbiology.com

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By Phone: 1-661-524(LBI)-0262 (USA)

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