Human Enolase 2/Neuron-specific Enolase Quantikine ELISA Kit Summary

Product Summary

The Quantikine Human Enolase 2 Immunoassay is a 4.5 hour solid-phase ELISA designed to measure human Enolase 2 in cell culture supernates, cell lysates, serum, and plasma. It contains E. coli-expressed recombinant human Enolase 2 and has been shown to accurately quantitate the recombinant factor. Results obtained using natural human Enolase 2 showed linear curves that were parallel to the standard curves obtained using the Quantikine kit standards. These results indicate that this kit can be used to determine relative mass values for naturally occurring human Enolase 2.

Preparation and Storage

Background: Enolase 2/Neuron-specific Enolase

Enolase 2 (2-phospho-D-glycerate hydrolyase; also Neuron-specific Enolase, NSE, neural enolase and gamma-enolase) is a 46 kDa member of the Enolase family of enzymes. It is expressed in developing neurons and glia, is known to catalyze the generation of phosphoenolpyruvate, and is suggested to possess neurotrophic activity for neurons, likely through an extracellular mechanism. Human Enolase 2 is 434 amino acids (aa) in length. The enzymatic site spans most of the length of the molecule. Enolase 2 exists as both a noncovalently-linked homodimer, or heterodimer with alpha-enolase. Full-length human Enolase 2 shares 99% aa identity with both mouse and canine Enolase 2. It shares 83% aa identity with human enolases 1 and 3.

Assay Procedure

Refer to the product datasheet for the complete assay procedure.

Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate.

1.    Prepare all reagents, standard dilutions, and samples as directed in the product insert.

2.    Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.

3.    Add 50 μL of Assay Diluent to each well.

4.    Add 50 μL of Standard, control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours.

5.    Aspirate each well and wash, repeating the process 4 times for a total of 5 washes.

6.    Add 100 μL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours.

7.    Aspirate and wash 5 times.

8.    Add 100 μL Substrate Solution to each well.

9.    Add 100 μL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.