Product/Service

Monoclonal Antibody Production

Source: Spring Valley Laboratories, Inc.
Spring Valley's monoclonal development program is divided into phases to provide you flexibility and control of the progression of the project
New Page 5

 

Spring Valley's monoclonal development program is divided into phases to provide you flexibility and control of the progression of the project.

In addition to the typical project outlined here, other more or less complex protocols are possible, including unique immunization approaches, double, triple, and special screening assays, or utilization of other strains of mice. Please inquire if you have special needs.

Phase 1: Immunization

 (5 to 6 weeks)
This phase accomplishes immunization, the development of the screening assay, and the titration of the mouse sera to determine the best mouse candidate for fusion. At the conclusion of this phase, in consultation with the investigator, a decision is made regarding the next step. The usual protocol for this phase includes:

Typical protocol: immunization
  • Immunization of five mice (50 - 100 µg immunogen required/mouse)
  • Two boosts (50 - 100 µg immunogen required per mouse)
  • Serum collection
  • ELISA assay to determine titer (= 30-60 µg of immunogen required)

    • Phase 2: Fusion

     (2 to 3 weeks)
    The selected mouse or mice are given a final boost. Spleen cells are then isolated and fused to NS-1 myeloma cells using polyethylene glycol fusion. Supernatant fluid, from wells which show growth, are tested in the ELISA screening assay and the results are reported to the investigator.

    Typical protocol: fusion
  • Immunogen boost
  • Isolation of spleen cells
  • Fusion
  • Plating of fused cells
  • Feeding and culturing of hybridoma cells
  • ELISA screening of culture supernatant fluid (= 100-150 µg of immunogen required)

    • Phase 3: Cloning

     (3 to 4 weeks)
    In consultation, Spring Valley and the investigator will select hybridomas for cloning. The selected hybridomas are cloned by limiting dilution in 96 well plates to isolate clones of interest. ELISA results are reported to the investigator to determine further action. The investigator also has the option of determining immunoglobulin isotype.

    Typical protocol: cloning
  • Selecting hybridomas
  • Cloning selected hybridomas
  • Feeding and culturing hybridoma cells
  • ELISA screening of culture supernatant fluid(s) ((100-150 (g of immunogen required)

    • Phase 4: Ascites

     (2 to 3 weeks)
    In this phase the investigator selects as many clones as desired for ascites production. Ascites production is priced in five-mice units but can be expanded to produce as large a volume as required. Typically each mouse will produce between 3 and 5 ml of ascites depending upon the characteristics of the cell line.

    Typical protocol: ascites
  • Pristane priming of mice
  • Injecting hybridoma cells
  • Harvesting ascites

    • Additional Monoclonal Antibody Services

    Cell Line Maintenance

  • Cell line cloning
  • Cell line expansion
  • Supernatant fluid production
  • Cryopreservation
  • Liquid nitrogen storage

    • Ascites Production


    Ascites is produced in BALB/c mice. Each mouse is tapped a maximum of 3 times. Yields vary and are dependent upon the immunogen.

    5 mice: estimated yield 15 to 25 ml
    20 mice: estimated yield 60 to 100 ml
    50 mice: estimated yield 150 to 250 ml
    100 mice: estimated yield 300 to 500 ml
    250 mice: estimated yield 750 to 1250 ml

    In vitro Production of Monoclonal Antibodies


    Spring Valley can produce monoclonal antibodies from your hybridoma using a hollow fiber system, cell culture flasks, or CO2/O2 gas permeable bags as an alternative to ascites. The antibody yield of the in vitro production methods will vary with each cell line. Please call for a quote.

    Spring Valley Laboratories, Inc., P.O. Box 242, Woodbine, MD 21797. Tel: 410-795-2222; Fax: 410-795-2242.