|
|
|
|
Lymphocyte
Cell Fusion
 |
Spleen cells from an immunized mouse are fused with a
non-antibody producing tumor cell line using polyethylene glycol. |
| Using HAT as a selective media, only those hybrids that
have both the nucleus from the spleen and from the tumor line will grow. |
| The resulting hybridoma has the antibody producing
capabilities of the spleen cell and the "immortality" of the tumor cell. |
| Hybridoma supernatants are screened for the presence of
antibody. |
| Those selected are cloned and expanded. Positive clones
may be expanded in tissue culture or grown as ascites tumors in mice. |
Antibody
Screening Assay
- Antigen is noncovalently attached to each well of a microtiter plate. Free antigen is
washed away, then non-specific sites on the plate are blocked using a gelatin buffer. Test
antisera or supernatants are added and any specific antibody binds to the solid phase
antigen. Washing removes unbound antibodies.
|
 |
- Enzyme-labeled anti-immunoglobulin is added. This binds to the bound antibody. Excess
conjugate is washed off and substrate is added.
|
- The action of bound enzyme on substrate produces a colored product, which is detected as
increased absorbance in a spectrophotometer.
|
Antibodies
Technology
|
Antibody technology has proven to show utility in many areas of Biological and
Biochemical Sciences due to both the qualitative and quantitative nature of antibodies.
 Typical uses for antibody reagents include:
ELISA assay development
Western blot detection
Immunofluorescent labeling
Immunohistochemical labeling
Immunoprecipitation
Affinity purification
Epitope mapping
Flow cytometry
Various bioassays
|
Immunological Resource Center
Liping Wang, Laboratory Supervisor
264D Burrill Hall, 407 South Goodwin Avenue, Urbana, IL 61801
Phone: (217) 244-0557 FAX: (217) 333-8286244-6697
Email: IRC@life.uiuc.edu
|
| Last edited:
05 Nov 2004
|
|
|
|
