Human TIM-3 Quantikine ELISA Kit Summary

Product Summary

The Quantikine Human TIM-3 Immunoassay is a 2.5 hour solid phase ELISA designed to measure TIM-3 levels in cell culture supernates, cell lysates, serum, plasma, and urine. It contains NS0-expressed recombinant human TIM-3 and antibodies raised against the recombinant protein. Results obtained for naturally occurring human TIM-3 showed linear curves that were parallel to the standard curves obtained using the Quantikine Human TIM-3 Immunoassay standards. These results indicate that this kit can be used to determine relative mass values for natural human TIM-3.

Preparation and Storage

Background: TIM-3

TIM-3 (T cell immunoglobulin and mucin domain-3), also known as HAVCR2 is an immunosuppressive protein that enhances tolerance and inhibits anti-tumor immunity. It is upregulated on several populations of activated myeloid cells (macrophage, monocyte, dendritic cell, microglia, mast cell) and T cells (Th1, CD8+, NK, Treg). TIM-3 ligation by Galectin-9 attenuates CD8+ and Th1 cell responses and promotes the activity of Treg and myeloid derived suppressor cells. Dendritic cell-expressed TIM-3 dampens inflammation by enabling the phagocytosis of apoptotic cells and the cross-presentation of apoptotic cell antigens. TIM-3 also binds the alarmin HMGB1, thereby preventing the activation of TLRs in response to released tumor cell DNA.

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.