A highly sensitive enzyme immunoassay for G-CSF in human plasma

Area of Science:

• Biochemistry
• Immunology
• Hematology

Background:

• Granulocyte colony-stimulating factor (G-CSF) is crucial for neutrophil production.
• Accurate measurement of plasma G-CSF is essential for understanding various physiological and pathological conditions.
• Existing methods for G-CSF quantification may lack sensitivity or specificity.

Purpose of the Study:

• To develop and validate a sensitive sandwich enzyme-linked immunosorbent assay (ELISA) for quantifying human plasma G-CSF.
• To assess the assay's performance characteristics, including sensitivity, specificity, and reproducibility.
• To determine baseline plasma G-CSF levels in a cohort of healthy volunteers.

Main Methods:

• A sandwich ELISA was developed using a mouse monoclonal antibody and a sheep polyclonal antibody conjugated with beta-D-galactosidase.
• Assay validation included assessment of intra- and inter-assay coefficients of variation and cross-reactivity with other cytokines.
• Plasma G-CSF levels were measured in 57 healthy volunteers, and the relationship with absolute neutrophil counts was analyzed.

Main Results:

• The developed ELISA demonstrated high sensitivity with a detection limit of 0.5 pg/mL.
• Intra-assay coefficients of variation ranged from 2.2% to 3.6%, and inter-assay variations were 8.3% to 10.2%.
• The assay showed no cross-reactivity with other tested human cytokines. Mean plasma G-CSF in healthy volunteers was 10.5 ± 4.5 pg/mL, with no significant correlation to neutrophil counts.

Conclusions:

• The developed sandwich ELISA is a sensitive and reproducible method for measuring human plasma G-CSF.
• This assay enables accurate quantification of G-CSF, facilitating further research into its biological roles.
• Baseline plasma G-CSF levels in healthy adults were established, providing a reference range for future studies.