FlowLITE: A protocol to characterize and quantify total antibody isotypes in human plasma using flow cytometry

Area of Science:

• Immunology
• Biotechnology
• Analytical Chemistry

Background:

• Antibody characterization is crucial for understanding immune responses and disease.
• Existing methods for antibody isotype and light-chain quantification can be resource-intensive.

Purpose of the Study:

• To provide a detailed protocol for the Flow Cytometry Light-chain and Isotype Expression (FlowLITE) assay.
• To enable multiplexed quantification of antibody light-chain and isotype expression in human plasma.

Main Methods:

• The Flow Cytometry Light-chain and Isotype Expression (FlowLITE) assay utilizes bead-based flow cytometry.
• The protocol involves antibody titration, multiplexed assay bead preparation, sample staining, and data analysis.
• Quantification of antibody light chains and isotypes in small fluid volumes is achieved.

Main Results:

• The FlowLITE assay allows for multiplexed quantification of all antibody isotypes.
• The protocol details steps for characterizing antibody light-chain and isotype expression in human plasma.
• The method is applicable to various species and fluid specimens.

Conclusions:

• The Flow Cytometry Light-chain and Isotype Expression (FlowLITE) assay offers a high-throughput method for antibody analysis.
• This protocol facilitates detailed characterization of antibody expression profiles.
• The assay is valuable for immunological research and diagnostics.