Human LIF Quantikine ELISA Kit Summary

Product Summary

The Quantikine^® Human LIF Immunoassay is a 4.5 hours solid phase ELISA designed to measure human LIF in cell culture supernates, serum, and plasma. It contains HEK293-expressed recombinant human LIF and antibodies raised against the recombinant protein. Results obtained for naturally occurring human LIF showed linear curves that were parallel to the standard curves obtained using the recombinant Quantikine^® kit standards. These results indicate that this kit can be used to determine relative mass values for natural human LIF.

Preparation and Storage

Background: LIF

Based on its helical structure, LIF (Leukemia Inhibitory Factor) is considered a member of the Interleukin-6 family of cytokines. Functionally, it has been implicated in a many physiological processes including development, hematopoiesis, bone metabolism, and inflammation. Some cell types known to express LIF include activated T cells, monocytes, astrocytes, osteoblasts, keratinocytes, regenerating skeletal muscle, mast cells, and fibroblasts. The activities of LIF are mediated through a high-affinity heterodimeric receptor complex consisting of two membrane glycoproteins: an alpha subunit (LIF R alpha, also known as LIF R beta and CD118) that binds LIF with low affinity and the 130 kDa (gp130) subunit that does not bind LIF by itself, but is required for high-affinity binding of LIF by the complex.

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.