Sera used for complement-mediated cytolysis can alter B cell function in vitro.

Area of Science:

• Immunology
• Cell Biology

Background:

• Nonspecific effects of reagents used for lymphocyte isolation are typically assessed by cell viability and proliferation.
• Monoclonal anti-human B cell antibodies are used to isolate B cell subpopulations.

Purpose of the Study:

• To investigate consistent changes in B cell function observed between donors during B cell subpopulation isolation using monoclonal antibodies.
• To identify the source of these alterations in B cell function.

Main Methods:

• Characterization of monoclonal anti-human B cell antibodies.
• Analysis of B cell function, viability, proliferation, and subpopulation distribution.
• Testing of commercial complement-containing sera from different vendors and animal species (rabbit, guinea pig).
• Stimulation of B lymphocytes with pokeweed mitogen or Staphylococcus aureus (Cowan I).

Main Results:

• Consistent alterations in B cell function were detected between donors, irrespective of the antibody used for depletion.
• The source of these alterations was identified as commercial complement-containing sera used for cytolysis.
• Different lots of serum from 3 vendors and 2 animal species altered B lymphocyte differentiation.
• Cell viability, lymphocyte subpopulation distribution, and proliferative responses remained unaffected.

Conclusions:

• Commercial complement-containing sera can introduce variability in B cell differentiation assays.
• Serum lot and source (animal species, vendor) significantly impact B lymphocyte differentiation.
• Researchers must account for the influence of complement-containing sera on B cell function in experimental design and interpretation.