A cell surface ELISA in the mouse using only poly-L-lysine as cell fixative.

Area of Science:

• Immunology
• Cell Biology
• Biochemistry

Background:

• Enzyme-linked immunosorbent assays (ELISA) are widely used for antibody detection.
• Previous ELISAs often used human or tumor cells, limiting application to normal mouse lymphocytes.
• Normal mouse spleen cells offer access to genetic resources from diverse mouse strains.

Purpose of the Study:

• To develop a novel ELISA for analyzing antibodies to cell surface antigens on normal mouse lymphocytes.
• To optimize the preparation of cell-coated ELISA plates for improved cell surface determinant preservation.
• To establish a sensitive and stable assay for antibody quantification and screening.

Main Methods:

• Developed an ELISA using normal mouse spleen cells coated onto plates.
• Eliminated glutaraldehyde fixation, using poly-L-lysine for cell adherence and preservation.
• Detected biotinylated monoclonal antibodies at approximately 10 ng/well.
• Detected unlabeled antibodies in inhibition assays at approximately 10 ng/well.

Main Results:

• The optimized ELISA effectively uses normal mouse spleen cells.
• Eliminating glutaraldehyde preserved cell surface determinants.
• The assay demonstrated high sensitivity for antibody detection (approx. 10 ng/well).
• Cell-coated plates are stable and can be stored for months.

Conclusions:

• A robust and sensitive ELISA for detecting antibodies to cell surface antigens on normal mouse lymphocytes has been established.
• This method enables the utilization of genetic resources from various mouse strains.
• The assay is suitable for both antibody quantification and screening of monoclonal antibody fusions.