The Biosensis Rat 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-2214) and full-length proNGF (BEK-2236).

Both kits are sandwich ELISAs, useful for the quantification of mature NGF and proNGF in less than 4 hours in cell culture supernatants and tissue homogenates only if used as directed, with a simplified protocol and no loss of sensitivity or specificity. Please refer to the most current kit protocols for BEK-2214 (Mature BDNF Rapid ELISA) and BEK-2236 (proNGF Rapid ELISA), for specific use instructions for each substrate application.

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-NGF 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 rat NGF protein as standard, and thus the Mature NGF/proNGF Combo kit is designed to measure the rat NGF isoforms. However, the rat mature NGF ELISA kit will cross-react with mouse and human mature NGF protein. The proNGF ELISA kit contains a recombinant mouse proNGF standard expressed in E.coli. Mouse proNGF and rat proNGF share 96% sequence homology, and internal validation data has demonstrated the ability of the proNGF ELISA to detect proNGF in rat PC12 cell lysates and rat brain tissue homogenate. However, the proNGF ELISA shows only little cross-reactivity (20%) with human proNGF.

This Combo kit is capable of distinguishing and independently quantifying the mature NGF and full-length NGF isoforms. Internal validation data demonstrates that the mature NGF ELISA assay antibodies preferentially detect the mature form of NGF. Cross-reaction of full-length proNGF was < 0.1% when assayed at 25 ng/mL in assay buffer, and not detectable when assayed across the mature NGF calibration range. The antibodies used in the proNGF ELISA kit bind epitopes within the pro-domain (capture) and mature domain (detection) of the protein, thus the proNGF ELISA assay does not detect the pro-domain peptide. No cross-reactivity is observed in the proNGF ELISA kit with mature mouse NGF and full-length proBDNF when tested in assay buffer.

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.

Nerve growth factor (NGF) is synthesized as a precursor (proNGF) which may be released and have physiological functions to cause cell death. It binds neurotrophin receptor p75 and sortilin and may also be important for the development of nervous system. proNGF is synthesized in target tissues and glia, transported retrogradely and may be released.

Mature NGF/proNGF Combo (BEK-2214/2236)

See BEK-2214 & BEK-2236 for specific details

Rat

Mature NGF (3.9 – 250 pg/mL) proNGF (0.156 – 10 ng/mL)

Typical limit of detection (LOD) for rat mature NGF is 2 pg/mL determined as 150% of the blank value. Typical limit of detection (LOD) for rat proNGF is less than 50 pg/mL, determined as blank value plus 3x standard deviation of blank OD (n=10).

Mature NGF ELISA: Detects rat NGF, and cross-reacts with human and mouse mature NGF. This mature NGF ELISA preferentially detects mature NGF over full-length proNGF. proNGF ELISA: Detects mouse and rat proNGF only, and shows only little reactivity (20%) with human proNGF. The proNGF ELISA detects the full-length form of proNGF only, and does not quantify mature NGF or the pro-domain peptide.
Cross-reactivity of NGF isoforms:
Mature NGF ELISA: The antibodies used in this ELISA kit bind epitopes within the mature domain of the protein. No cross-reactivity was observed with brain derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and NT-4/5 tested at 25 ng/mL in assay buffer. Mature NGF (27 kDa) and full-length proNGF (50 kDa) were assayed in parallel at equimolar protein concentrations across the mature NGF ELISA calibration range (3.9-250 pg/mL; 0.14-9.2 pmol/L). OD readings for proNGF were indistinguishable from the assay's blank OD readings. proNGF ELISA: Does not cross-react with proBDNF and mature NGF. Mature NGF spiked into brain homogenate does not interfere with proNGF quantification.

Rat

ELISA

Colorimetric

Culture Supernatant, Tissue Homogenates

Beta-nerve growth factor; Ngfb; NGF; pro-beta nerve growth factor; proNGF

P25427

For research use only.

United States

12 months after date of receipt if unopened.

Store at 2-8°C

Shipped on ice packs