Recombinant human β‐nerve growth factor (NGF): Biological activity and properties in an enzyme immunoassay

Abstract

Nerve growth factor (NGF) supports sympathetic and sensory neurons in the peripheral nervous system and also functions in the development and maintenance of cholinergic neurons in the basal forebrain. NGF distribution can be studied in the brain of the rat and mouse with the use of a sensitive two-site enzyme immunoassay (EIA) for mouse NGF. It would be of interest to measure the NGF protein also in the human brain, especially against the background that the cholinergic neurons are severely deteriorated in senile dementia of the Alzheimer type. The limited immunological cross-reactivity between NGFs from different species has previously hampered attempts to determine levels of the human NGF. We have now examined the biological activity and immunological properties of human recombinant NGF protein in medium conditioned by COS cells transfected with the human NGF gene. The human NGF behaved similar to mouse NGF in a sympathetic ganglion bioassay. The monoclonal antibody 27/21 to mouse NGF was shown to effectively block the activity of both the human recombinant NGF and mouse native NGF. A two-site EIA using monoclonal antibody 27/21 was optimized. Under the conditions used, the EIA detected the human recombinant NGF with the same sensitivity (1 pg/ml) as shown for the mouse NGF. It should now be possible to test this EIA also on homogenized tissue to examine human NGF in brain samples from Alzheimer patients and age-matched controls.

β-Nerve Growth Factor Levels in Newborn Cord Sera

“…As shown in Figure 1, the dose-response curve for the standard purified mouse NGF showed a linear relationship. Thus, the sensitivity of this two-site EIA under these conditions was routinely approximately 2 pg/ mL and is similar to that reported by Soderstrom et al (8).…”

“…Their recorded P-NGF levels were as high as 292 pg/mL. DISCUSSION P-NGF from humans and mice have been characterized and found to be highly homologous (8). Each monomer of both types is composed of 1 18 amino acids, but the amino acid sequence differs (8).…”

“…DISCUSSION P-NGF from humans and mice have been characterized and found to be highly homologous (8). Each monomer of both types is composed of 1 18 amino acids, but the amino acid sequence differs (8). Until recently, because of the similarity between human and mouse NGF, MAb 2712 1 to mouse /3-NGF has been used in a two-site EIA to detect human NGF from various tissues (5)(6)(7)(8).…”

“…Substantial progress has been made in research on nerve H A . A two-site EIA for B-NGF, slightly modified from that ~wth factors since the initial discovery by Levi Montalcini and described by Soderstrom et al (8), was used for the present study. ~mburger of the first neurotrophin, NGF (1).…”

NGF Withdrawal Induces Apoptosis in CESS B Cell Line through p38 MAPK Activation and Bcl-2 Phosphorylation

“…NGF production was also assessed by ELISA as described by Söderström et al (16) by using a commercially available NGF mouse monoclonal antibody (mAb) 27/21 (Boehringer Mannheim, Mannheim, Germany). The sensitivity of the test was ranging between 8 and 20 pg/ml.…”