Abstract
A four-antibody sandwich enzyme-linked immunosorbent assay (ELISA) for vascular endothelial growth factor (VEGF) for application in blood (serum and plasma) and tumor tissue extracts was set up within the framework of the EORTC Receptor and Biomarker Study Group (RBSG). Polyclonal antibodies against VEGF165 were raised in chickens and rabbits, and used in a previously described assay format. The assay was validated and characterized for use in serum, plasma and tumor tissue extracts. The resulting VEGF ELISA was found to be specific for VEGF165 and VEGF121, the main isoforms of VEGF. The assay showed good precision and parallelism in serial dilutions of samples. The assay was not susceptible to interference by heterophilic antibodies because avian antibodies (duck anti-chicken and chicken anti-VEGF) were used in the pre-analyte stage and mammalian antibodies (rabbit anti-VEGF and goat anti-rabbit) in the post-analyte stage. In conclusion, a sensitive, robust and specific VEGF ELISA has been developed. Research into the prognostic value of VEGF employing this assay is currently underway.
Publication types
-
Comparative Study
-
Evaluation Study
-
Validation Study
MeSH terms
-
Animals
-
Antibodies, Heterophile / immunology
-
Antibody Specificity
-
Artifacts
-
Biomarkers, Tumor / analysis*
-
Biomarkers, Tumor / blood
-
Breast Neoplasms / blood
-
Breast Neoplasms / chemistry*
-
Chickens / immunology
-
Cytosol / chemistry
-
Dimerization
-
Dose-Response Relationship, Immunologic
-
Ducks / immunology
-
Endothelial Growth Factors / analysis*
-
Endothelial Growth Factors / blood
-
Endothelial Growth Factors / immunology
-
Endothelial Growth Factors / isolation & purification
-
Enzyme-Linked Immunosorbent Assay*
-
Female
-
Goats / immunology
-
Humans
-
Immune Sera
-
Lymphokines / analysis*
-
Lymphokines / blood
-
Lymphokines / immunology
-
Lymphokines / isolation & purification
-
Neoplasm Proteins / analysis*
-
Neoplasm Proteins / blood
-
Protein Isoforms / analysis*
-
Protein Isoforms / blood
-
Rabbits / immunology
-
Recombinant Fusion Proteins / immunology
-
Recombinant Fusion Proteins / isolation & purification
-
Reproducibility of Results
-
Sensitivity and Specificity
-
Species Specificity
-
Vascular Endothelial Growth Factor A
-
Vascular Endothelial Growth Factors
Substances
-
Antibodies, Heterophile
-
Biomarkers, Tumor
-
Endothelial Growth Factors
-
Immune Sera
-
Lymphokines
-
Neoplasm Proteins
-
Protein Isoforms
-
Recombinant Fusion Proteins
-
VEGFA protein, human
-
Vascular Endothelial Growth Factor A
-
Vascular Endothelial Growth Factors