Human TIGIT DuoSet ELISA New

Catalog #: DY7898-05 Datasheet / COA / SDS

Catalog #	Availability	Size / Price	Qty
DY7898-05

Ancillary Products Available

DY008B: DuoSet ELISA Ancillary Reagent Kit 2

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All TIGIT ELISAs

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Procedure

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Summary Product Features Kit Content Other Reagents Required Data Examples Product Datasheets Preparation and Storage Background

Human TIGIT DuoSet ELISA Summary

Assay Type

Solid Phase Sandwich ELISA

Format

96-well strip plate

Sample Volume Required

100 µL

Assay Range

21.9 - 1,400 pg/mL

Sufficient Materials

For five 96-well plates*

Specificity

Please see the product datasheet

* Provided that the recommended microplates, buffers, diluents, substrates and solutions are used, and the assay is run as summarized in the Assay Procedure provided.

This DuoSet ELISA Development kit contains the basic components required for the development of sandwich ELISAs to measure natural and recombinant human TIGHT. The suggested diluent is suitable for the analysis of most cell culture supernate samples. Diluents for complex matrices, such as serum and plasma, should be evaluated prior to use in this DuoSet.

Product Features

Optimized capture and detection antibody pairings with recommended concentrations save lengthy development time
Development protocols are provided to guide further assay optimization
Assay can be customized to your specific needs
Economical alternative to complete kits

Kit Content

Capture Antibody
Detection Antibody
Recombinant Standard
Streptavidin conjugated to horseradish-peroxidase (Streptavidin-HRP)

Other Reagents Required

DuoSet Ancillary Reagent Kit 2 (5 plates): (Catalog # DY008 B) containing 96 well microplates, plate sealers, substrate solution, stop solution, plate coating buffer (PBS), wash buffer, and Reagent Diluent Concentrate 2.

The components listed above may be purchased separately:

PBS: (Catalog # DY006), or 137 mM NaCl, 2.7 mM KCl, 8.1 mM Na2HPO4, 1.5 mM KH2PO4, pH 7.2 - 7.4, 0.2 µm filtered

Wash Buffer: (Catalog # WA126), or equivalent

Reagent Diluent*

Blocking Buffer*

Substrate Solution: ELISA TMB Substrate (R&D Systems,

Catalog # DY999B)

Stop Solution: 2N H2SO4 (Catalog # DY994)

Microplates: R&D Systems (Catalog # DY990)

Plate Sealers: ELISA Plate Sealers (Catalog # DY992)

*For the Reagent Diluent and Blocking Buffer are recommended for a specific DuoSet ELISA Development Kit, see the product datasheet.

Scientific Data

N/A

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Human TIGIT DuoSet ELISA Standard Curve

Product Datasheets

Product Datasheet

COA

SDS

Preparation and Storage

Stability & Storage

Store the unopened product at 2 - 8 °C. Do not use past expiration date.

Background: TIGIT

TIGIT (T cell Immunoreceptor with Ig and ITIM domains), also called Vstm3, Vsig9, and WUCAM, is a transmembrane protein in the CD28 family of the Ig superfamily proteins. TIGIT is expressed on NK cells and subsets of activated, memory and regulatory T cells, and particularly on follicular helper T cells within secondary lymphoid organs. It binds to CD155/PVR/Necl-5 and Nectin-2/CD112/PVRL2 on dendritic cells (DC) and endothelium. Binding of TIGIT by DC induces IL-10 release and inhibits IL-12 production. Ligation of TIGIT on T cells downregulates TCR-mediated activation and subsequent proliferation, while NK cell TIGIT ligation blocks NK cell cytotoxicity. CD155 and Nectin-2 also interact with DNAM-1/CD226 and CD96/Tactile, and TIGIT binding to CD155 can antagonize the effects of DNAM-1. Soluble TIGIT is able to compete with DNAM-1 for CD155 binding and attenuates T cell responses, while mice lacking TIGIT show increased T cell responses and susceptibility to autoimmune challenges.

Long Name:

T Cell Immunoreceptor with Ig and ITIM Domains

Entrez Gene IDs:

201633 (Human); 100043314 (Mouse); 363784 (Rat); 102121588 (Cynomolgus Monkey)

Alternate Names:

T cell immunoreceptor with Ig and ITIM domains; TIGIT; VSIG9; VSTM3; WUCAM

Assay Procedure

GENERAL ELISA PROTOCOL

Plate Preparation

Dilute the Capture Antibody to the working concentration in PBS without carrier protein. Immediately coat a 96-well microplate with 100 μL per well of the diluted Capture Antibody. Seal the plate and incubate overnight at room temperature.
Aspirate each well and wash with Wash Buffer, repeating the process two times for a total of three washes. Wash by filling each well with Wash Buffer (400 μL) using a squirt bottle, manifold dispenser, or autowasher. Complete removal of liquid at each step is essential for good performance. After the last wash, remove any remaining Wash Buffer by aspirating or by inverting the plate and blotting it against clean paper towels.
Block plates by adding 300 μL Reagent Diluent to each well. Incubate at room temperature for a minimum of 1 hour.
Repeat the aspiration/wash as in step 2. The plates are now ready for sample addition.

Assay Procedure

Add 100 μL of sample or standards in Reagent Diluent, or an appropriate diluent, per well. Cover with an adhesive strip and incubate 2 hours at room temperature.
Repeat the aspiration/wash as in step 2 of Plate Preparation.
Add 100 μL of the Detection Antibody, diluted in Reagent Diluent, to each well. Cover with a new adhesive strip and incubate 2 hours at room temperature.
Repeat the aspiration/wash as in step 2 of Plate Preparation.
Add 100 μL of the working dilution of Streptavidin-HRP to each well. Cover the plate and incubate for 20 minutes at room temperature. Avoid placing the plate in direct light.
Repeat the aspiration/wash as in step 2.
Add 100 μL of Substrate Solution to each well. Incubate for 20 minutes at room temperature. Avoid placing the plate in direct light.
Add 50 μL of Stop Solution to each well. Gently tap the plate to ensure thorough mixing.
Determine the optical density of each well immediately, using a microplate reader set to 450 nm. If wavelength correction is available, set to 540 nm or 570 nm. If wavelength correction is not available, subtract readings at 540 nm or 570 nm from the readings at 450 nm. This subtraction will correct for optical imperfections in the plate. Readings made directly at 450 nm without correction may be higher and less accurate.