Human Total PON1 DuoSet IC ELISA Summary

Product Summary

This DuoSet IC ELISA contains the basic components required for the development of sandwich ELISAs to measure total PON1 in cell lysates. An immobilized capture antibody specific for PON1 binds both phosphorylated and unphosphorylated PON1. After washing away unbound material, a biotinylated detection antibody is used to detect both phosphorylated and unphosphorylated protein, utilizing a standard Streptavidin-HRP format.

Preparation and Storage

Background: PON1

PON (paraoxonase/lactonase) family members PON1, PON2 and PON3 are calcium-dependent antioxidant enzymes. PON1 is secreted into the bloodstream associated with high-density lipoproteins (HDL). Serum PON1 concentrations vary widely among normal individuals, in part due to differential expression of some polymorphisms. The antioxidant activity of PON1 and other PON members may slow the initiation and progression of atherosclerosis.

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.