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Generation and use of an antigen-specific hybrid to study B-cell function
R B Bankert1, E A Repasky, P K Mazzaferro
1Department of Molecular Immunology, Roswell Park Memorial Institute, Buffalo, NY 14263.
The Journal of Molecular and Cellular Immunology : JMCI
|January 1, 1987
Summary
B-lymphocytes re-express membrane-associated immunoglobulin (mIg) within six hours after antigen exposure, independent of new receptor synthesis. This suggests B-cell receptors are re-utilized or a cytoplasmic pool exists.
Area of Science:
- Immunology
- Cell Biology
- Biochemistry
Background:
- Crosslinking of membrane-associated immunoglobulin (mIg) on B-lymphocytes triggers significant cellular events, including receptor internalization and re-expression.
- Understanding the kinetics and mechanisms of B-cell receptor (BCR) re-expression is crucial for deciphering B-cell activation and antigen processing.
- Previous studies often used heterogeneous cell populations and anti-immunoglobulin antibodies, potentially limiting insights into antigen-specific responses.
Purpose of the Study:
- To determine the time course of mIg re-expression following antigen-induced clearance of the B-cell receptor.
- To investigate whether mIg re-expression is dependent on the synthesis of new immunoglobulin molecules.
- To characterize an antigen-specific B-cell hybrid for studying mIg dynamics.
Main Methods:
- Development and characterization of the antigen-specific B-cell hybridoma 2C3E1, targeting phthalate.
- Biochemical confirmation of membrane-associated immunoglobulin (mIg) expression.
- Induction of receptor clearance using phthalate-keyhole limpet hemocyanin (KLH) and monitoring of mIg re-expression kinetics.
Main Results:
- Re-expression of antigen-specific receptors was detected within one hour and completed by six hours after antigen exposure (first or second pulse).
- This re-expression did not require the synthesis of new immunoglobulin receptors.
- The hybridoma also produced secreted immunoglobulin (sIg), unexpectedly found associated with the cell surface, potentially via vesicles.
Conclusions:
- B-cell receptors are re-utilized following antigen-induced clearance, as re-expression occurs without new synthesis.
- A presynthesized pool of mIg may exist within the cytoplasm of B-lymphocytes, facilitating rapid receptor replenishment.
- The presence of surface-associated sIg, possibly linked to vesicular transport, warrants further investigation into Ig secretion mechanisms.