A fluorometric microassay for histamine release from human gingival mast cells

Area of Science:

• Immunology
• Cell Biology
• Biochemistry

Background:

• Mast cells are key immune effector cells involved in inflammatory responses.
• Histamine release from mast cells plays a significant role in various physiological and pathological processes, particularly in oral mucosal tissues.
• Accurate and efficient methods for quantifying histamine release are crucial for understanding mast cell function in disease.

Purpose of the Study:

• To develop and validate a rapid, inexpensive microassay for determining histamine release from human gingival mast cells.
• To establish the sensitivity and detection limits of the developed assay.
• To assess the utility of the microassay in studying mast cell activation in oral mucosal conditions.

Main Methods:

• A microassay was developed utilizing the fluorescent coupling of histamine with o-phthalaldehyde (OPT) at a high pH.
• The assay was performed in 384 black well microplates using a small sample volume (10 microl/well).
• Human mast cell line (HMC-1) and primary human gingival mast cells were stimulated with substance P, anti-IgE, and calcium ionophore A23187 to measure histamine release.

Main Results:

• The microassay demonstrated a histamine detection limit of 0.031 microg/ml.
• Calcium ionophore A23187 significantly increased histamine release from both HMC-1 cells and primary gingival mast cells (p < 0.05).
• The assay proved effective in quantifying histamine release from stimulated mast cells.

Conclusions:

• A novel, rapid, and cost-effective microassay for quantifying histamine release from human mast cells has been successfully developed.
• This assay is sensitive and suitable for use with limited sample volumes, making it practical for research.
• The microassay provides a valuable tool to advance the study of mast cell-mediated histamine release in the context of diseased oral mucosa.