The Biosensis Human Mature NGF/proNGF Combo Rapid enzyme-linked immunosorbent assay (ELISA) Kit combines individual, but complementary ELISA kits for the two most important NGF isoforms: Mature NGF (BEK-2212) and full-length proNGF (BEK-2226).
Both kits are sandwich ELISAs, useful for the quantification of mature NGF and proNGF in less than 4 hours in cell culture supernatants, serum, and plasma (citrate) only if used as directed, with a simplified protocol and no loss of sensitivity or specificity. Researchers have used both kits for NGF/proNGF measurement in human urine, however, Biosensis has not yet internally validated the use of urine in both ELISA kits. End-users are strongly advised to perform essential validation experiments to assure accurate NGF/proNGF quantification in human urine. Please refer to the most current kit protocols for BEK-2212 (Mature BDNF Rapid ELISA) and BEK-2226 (proNGF Rapid ELISA), for specific use instructions for each substrate application, in particular blood samples.
The Mature NGF ELISA kit consists of a pre-coated mouse monoclonal anti-NGF capture antibody, a biotinylated anti-NGF detection antibody and horseradish peroxidase (HRP)-conjugated streptavidin. The proNGF ELISA kit consists of a pre-coated anti-proNGF capture antibody, a biotinylated anti-proNGF detection antibody and horseradish peroxidase (HRP)-conjugated streptavidin. The addition of a substrate (3,3',5,5'-tetramethylbenzidine, TMB) yields a coloured reaction product which is directly proportional to the concentration of mature NGF or proNGF present in samples and protein standards. A NGF and proNGF positive control (QC sample) is provided to assure consistent assay performance.
The Mature NGF ELISA kit employs a high-quality recombinant human NGF standard approved by the World Health Organization (WHO). The proNGF ELISA kit contains a recombinant human proNGF standard expressed in E.coli. Note that this Mature NGF/proNGF Combo kit is designed to measure the human protein forms, although due to sequence homology the Mature NGF ELISA kit will detect mouse and rat mature NGF. The proNGF ELISA kit does not cross-react with rodent forms of proNGF.
This Combo kit is capable of distinguishing and independently quantifying the mature NGF and full-length NGF isoforms. Internal validation data demonstrates <0.1% cross-reactivity (determined at 25 ng/mL, diluted in assay buffer) of human proNGF protein in the mature NGF ELISA, demonstrating the preferential quantification of mature NGF over full-length human proNGF. The absence of proNGF cross-reactivity in the mature NGF ELISA kit has been independently confirmed by
Mossa AH et al. (2020). The antibodies used in the proNGF ELISA kit bind epitopes within the pro-domain of the protein and therefore recognize proNGF and the pro-domain peptide, but do not cross-react with mature NGF!
Important: Accurate NGF quantification in human citrate-plasma requires the addition of Heterophilic Antibody Blocker BL-005-500. Accurate proNGF quantification in human serum requires the addition of Heterophilic Antibody Blocker BL-003-1000. Both Heterophilic Antibody Blockers are provided in the kit.
This ELISA kit has not been tested for other applications. It has been configured for research use only and is not to be used for diagnostic or clinical procedures.
