A simple panning method for the selection of cell surface antigen transfectants.

Area of Science:

• Immunology
• Molecular Biology
• Cell Biology

Background:

• Mouse Ltk- cells (H-2k) were used as a recipient cell line.
• Transfection with DNA from H-2d-positive cells can confer H-2d antigen expression.

Purpose of the Study:

• To isolate mouse Ltk- cells that express H-2d antigens after DNA transfection.
• To develop an efficient method for selecting transfected cells reactive with specific monoclonal antibodies.

Main Methods:

• Cells were co-transfected with total cell DNA and a plasmid containing the thymidine kinase (tk) gene.
• Selection of tk+ transfectants was performed in hypoxanthine/aminopterin/thymidine (HAT) medium.
• An indirect panning method using anti-H-2d monoclonal antibodies (McAb) and anti-immunoglobulin (anti-Ig) coated dishes was employed for cell isolation.

Main Results:

• The indirect panning method successfully isolated transfected cells reactive with anti-Ld/Rd or anti-Kd McAb.
• The two-step panning procedure required only one or two cycles for isolation.
• The method's advantage lies in the purification of only the anti-immunoglobulin reagent.

Conclusions:

• This study presents an effective and efficient indirect panning technique for isolating transfected cells expressing specific cell surface antigens.
• The method simplifies the selection process for genetically modified cells in immunology research.