领潮生物科技有限公司

Through the world-wide genome sequencing efforts on human, pathogenic microorganisms and parasites, many potential new protein sequences are available, and one of the easiest ways to approach their function is to obtain monoclonal antibodies against these unknown proteins using synthetic peptides or recombinant proteins derived from the DNA sequence.

Linc-Bio has gained extensive experience in this field and we offer a custom service, starting from the customer's antigen or a synthetic peptide, up to the final generation of monoclonal secreting cell lines. Moreover, mg to gram quantities of monoclonal antibodies can be produced with customer's hybridomas in tissue culture or as ascites.

The nature of the antigen, as well as its purity and the quantity available are important parameters in hybridoma development.

1. An amount of at least 5 mg of the antigen (proteins with purity>80% SDS) will be required.

3. Crude antigens as bacterium, virus or its proteins (without infectivity)can also be employed in proprietary immunization procedures if the supply of antigen is limited. However, a small amount of homogeneous antigen is critical for use in the screening.

SUGGESTED PROTOCOL AND TIME SCHEDULE
Week	Procedure	Specification
First 4-6	Immunization	5 BALB/c mice are immunized and serum antibody titers are monitored by ELISA.Detection of specific antibodies in the serum completes this phase.
Next 3	Fusion	A PEG mediated fusion is performed between the immune splenocytes and the HGPRT deficient NSO/2 myeloma cells. The viable hybridomas are selected using HAT medium.Identification of viable colonies after fusion completes this phase.
Next 2	Screening	Supernatants from all viable hybridoma are subjected to multiple screenings by ELISA. Samples from positive supernatant are provided to the customer for confirmatory screening. Identification of positive hybridoma completes this phase.
Next 4	Cloning	Cloning is performed by limiting dilution. Antibody specificity is determined by ELISA and the best clone is selected. The hybridoma is stored under liquid nitrogen. The customer is provided with at least two clones of hybridoma. Cloning of hybridoma completes this phase.
Next 2-4	Ascites fluid production	The selected hybridoma is expanded. The customer is provided with 30 ml of ascitic fluid and 100 ml of tissue culture supernatant.
The duration of a typical project is 4 to 5 months.

PRICE Specific price quotation and schedules for individual projects are prepared upon request.

Linc-Bio offers a custom service, starting from the design of peptides, synthesis of the peptides, coupling to carrier proteins up to the antibody production and affinity purification of the antiserum. antiserum. We recommended that at least two animals (two rabbits or three rats) be used for each antigen, due to variability in individual immune response.

1. An amount of at least 5 mg of the antigen (proteins with purity>80% SDS) will be required.

PROTOCOL The suggested protocol shown in the Table, it has proven effective with a wide variety of antigens. Developed at Linc-Bio, it uses an intradermal or subcutaneous injections for initial immunizations, followed by a timed series of intramuscular injections and serum samplings.

SUGGESTED PROTOCOL AND TIME SCHEDULE
Day	Procedure	Specification
1	Physical examination Antigen formulation. Preimmune serum sample.
2	Initial Immunization.	Antigen emulsified in Freund's Complete Adyuvant.
20	First boost.	Antigen emulsified in Freund's Incomplete Adyuvant.
30	First bleed; ELISA.
40	Second boost.	Antigen emulsified in Freund's Incomplete Adyuvant.
50	Second bleed; ELISA.	Aliquots of the sera are sent to the customer.
60	Third boost.	Antigen emulsified in Freund's Incomplete Adyuvant.
70	Exsanguinate; ELISA.	Aliquots of the sera are sent to the customer. There is the option to continue or stop the project. Finally, the customer is provided with 50 to 60 ml of immune sera.
The duration of a typical project is 2 to 3 months.

ANIMALS	Antigen needed	Provided	Duration	Price
Rabbits	20mg	10mg SAS cut PcAbs	2 months	1000 USD
Rats	10mg	1mg SAS cut PcAbs	3 months	660 USD