Labeled TEMPO-Oxidized Mannan Differentiates Binding Profiles within the Collectin Families

Area of Science:

• Biochemistry
• Immunology
• Analytical Chemistry

Background:

• Rapid sepsis diagnosis is crucial for improving patient outcomes.
• Detecting pathogen-associated molecular patterns (PAMPs) in blood offers a host-independent sepsis biomarker.
• Lectins play a role in innate immunity by recognizing molecular patterns.

Purpose of the Study:

• To investigate a novel concept for sepsis detection using fluorescent ligands and lectins.
• To develop a method for creating fluorescent ligands with precise avidity for lectin binding.
• To demonstrate the potential of this system for identifying high-avidity ligands and developing diagnostic tools.

Main Methods:

• Chemical modification of yeast mannan to create fluorescent ligands using TEMPO oxidation and carbodiimide coupling.
• Assessing the avidity changes of modified mannan with human collectins (mannan-binding lectin and surfactant protein D) and porcine SP-D.
• Exposing fluorescently labeled lectin-ligand complexes to different solutions to observe ligand displacement.

Main Results:

• Chemical modifications altered mannan avidity for human collectins but not porcine SP-D.
• Fluorescent derivatives were captured by human lectins, and fluorescence loss occurred upon exposure to higher-affinity ligands.
• Ligand displacement demonstrated two-stage competition in collectin carbohydrate recognition, dependent on avidity discrepancies.

Conclusions:

• Chemically modulated fluorescent ligands can be displaced from collectins by higher-affinity ligands, indicating a competition mechanism.
• This approach shows promise for developing diagnostic tools for sepsis detection.
• The system could be adapted for multiplex array assays to identify high-avidity ligands.