Hyaluronan Quantikine ELISA Kit Summary

Product Summary

The Quantikine Hyaluronan Immunoassay is a 4.5 hour solid-phase ELISA designed to measure at least 35 kDa Hyaluronan in human, mouse, rat, canine, and porcine cell culture supernates, tissue homogenates, serum, and plasma samples. It contains Streptococcus pyogenes-derived Hyaluronan and NS0-expressed human Aggrecan. This immunoassay has been shown to accurately quantitate Hyaluronan.

Preparation and Storage

Background: Hyaluronan

Hyaluronan, also known as hyaluronic acid (HA) or sodium hyaluronate, is a naturally occurring linear polymer. Hyaluronan is a glycosaminoglycan (GAG) that is ubiquitously present in the extracellular matrix of all vertebrates and is also present in the capsule of some strains of Streptococci. Mammalian hyaluronan is synthesized by one of three distinct hyaluronan synthases (HAS1, 2, and 3), which produce HA polymers with different chain lengths and differ in their rates of synthesis. Whereas high molecular weight HA (> 500 kDa) is anti-angiogenic, anti-inflammatory and immunosuppressive, low molecular weight HA (10 - 500 kDa) is highly angiogenic and pro-inflammatory. HA oligomers are anti-apoptotic and upregulate heat shock protein expression. Functionally, hyaluronan molecules are important for the maintenance of a highly hydrated extracellular matrix in tissues, which is involved in cell adhesion and supports cell migration.

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.