Thallium Overview

ION Biosciences Brilliant Thallium Assay is a total assay solution for multi-well plate-based, high-throughput measurements of Tl+ flux through K+, Na+, non-selective cation channels, and some Na+ or K+ transporters. The ION Biosciences Brilliant Thallium Assay is also useful for a wide range of effectors of ion channels and transporters including G protein-coupled receptors, lipid and protein kinase, for example. In multi-well, plate-based formats, the Brilliant Thallium Assay can be used to discover and characterize the effects of tens to many tens-of-thousands of compounds and environmental factors on effectors of Tl+ flux. Over the past 15 years, fluorescence-based measures of Tl+ flux have been widely utilized to discovery small molecule modulators of a host of Ion channels, transporters, GPCRS and other targets of interest for both drug discovery and basic research.  Inside the ION Brilliant Assay package, you will find all the reagents necessary for preparation of the Brilliant Thallium Assay for use as a washed or no wash assay for adherent and non-adherent cells. The addition of a probenecid solution and an extracellular background masking solution offers users the ultimate in compatibility for cells types which are difficult to load with fluorescent Tl+ indicators (e.g. Chinese Hamster Ovary, CHO cells) and when performing assays in complete, serum-containing cell culture medium is desired.

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Buy No Wash Thallium Assay

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No Wash Thallium Assay Product Specifications

Molecular Weight

840 g/mol

Odor

None

Toxicology Data

Not Known

Handling and Storage

Store at -20°C. Protect from light and moisture

 
Package Contents
Description
Storage Temperature

Reagent A

WaveFront Thallium Indicator

-20°C

Reagent B

DySolv

room temperature

Reagent C

Assay Buffer

4°C

Reagent D

Probenecid Solution

4°C

Reagent E

TRS Solution

4°C

Reagent F

100 mM Thallium Solution

room temperature

Reagent G

Chloride-free Stimulus Solution

4°C

Reagent H

High Potassium, Chloride-free Stimulus Solution

4°C

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No Wash Thallium Assay Single Microplate Kit Basic Protocol (Adherent Cells)

  1. Prepare Brilliant Thallium Indicator Solution
    1. Add 50 μL DySolv (Reagent B) to the tube containing Brilliant Thallium Indicator (Reagent A)
    2. Vortex until Reagent A is fully dissolved
    3. Transfer the Reagent A solution to the bottle containing Assay Buffer (Reagent C) and vortex briefly
    4. OPTIONAL1 Add 100 μL Probenecid (Reagent D) to the bottle containing the Reagent C solution and vortex briefly
    5. OPTIONAL2 Add 400 μL TRS (Reagent E) to the bottle containing the Reagent C solution and vortex briefly
  2. Prepare Thallium Stimulus Solution3
    1. In a separate container, add an appropriate volume of 100 mM Thallium Stimulus Solution (Reagent F) to an appropriate volume of Chloride-free Stimulus Solution (Reagent G).
  3. Assay
    1. Remove cell culture medium from cell-containing plate
    2. Add Brilliant Thallium Indicator Solution (80 μL/well for 96-well plates, 20 μL/well for 384-well plates)
    3. Protect from direct light. Incubate for 1 hour at room temperature.
    4. Collect data4 using an appropriate fluorescent plate reader with liquid handling capabilities (WaveFront Panoptic, Hamamatsu FDSS, Molecular Devices FLIPR, Molecular Devices Flexstation) equipped with filters designed for fluorescein-type fluors (excitation ~490/emission ~520 nm). After a minimum of 10 seconds of data collection, add an appropriate volume of Thallium Stimulus Solution from step 2 using the plate reader’s liquid handling capabilities.

1 The use of Probenecid is indicated for some cell types to counteract these cells’ propensity to extrude Brilliant Thallium and other similar indicator dyes resulting in poor dye retention.

2 The TRS reagent is useful for suppressing background resulting from fluorescent compounds in the solutions surrounding the cells. Use of TRS may result in improved signal-to-noise.

3 The concentration of thallium in the stimulus buffer will vary by application. Investigators are encouraged to explore final thallium concentrations ranging from 0.5 to 4.5 mM to achieve the best results. Some targets, typically voltage-gated channels, may benefit from the use of High Potassium Chloride-free Stimulus Solution (Reagent H). Investigators are encouraged to explore mixtures of different proportions of Chloride-free Stimulus Solution and High Potassium, Chloride-free Stimulus Solution to achieve the best results.

4 Data collection typically includes the addition of test compounds or additional reagents to the assay plate during the course of data acquisition. Volumes and concentrations of test compounds and additional reagents vary. Commonly used addition volumes are 5, 10, 20 μL for 384-well plates and 20, 40, and 80 μL/well for 96-well plates.

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No Wash Thallium Assay Ten Microplate Kit Basic Protocol (Adherent Cells)

  1. Prepare Brilliant Thallium Indicator Solution
    1. Add 500 μL DySolv (Reagent B) to the tube containing Brilliant Thallium Indicator (Reagent A)
    2. Vortex until Reagent A is fully dissolved
    3. Transfer the Reagent A solution to the bottle containing Assay Buffer (Reagent C) and vortex briefly
    4. OPTIONAL1 Add 1000 μL Probenecid (Reagent D) to the bottle containing the Reagent C solution and vortex briefly
    5. OPTIONAL2 Add 4 mL TRS (Reagent E) to the bottle containing the Reagent C solution and vortex briefly
  2. Prepare Thallium Stimulus Solution3
    1. In a separate container, add an appropriate volume of 100 mM Thallium Stimulus Solution (Reagent F) to an appropriate volume of Chloride-free Stimulus Solution (Reagent G).
  3. Assay
    1. Remove cell culture medium from cell-containing plate
    2. Add Brilliant Thallium Indicator Solution (80 μL/well for 96-well plates, 20 μL/well for 384-well plates)
    3. Protect from direct light. Incubate for 1 hour at room temperature.
    4. Collect data4 using an appropriate fluorescent plate reader with liquid handling capabilities (WaveFront Panoptic, Hamamatsu FDSS, Molecular Devices FLIPR, Molecular Devices Flexstation) equipped with filters designed for fluorescein-type fluors (excitation ~490/emission ~520 nm). After a minimum of 10 seconds of data collection, add an appropriate volume of Thallium Stimulus Solution from step 2 using the plate reader’s liquid handling capabilities.

1 The use of Probenecid is indicated for some cell types to counteract these cells’ propensity to extrude Brilliant Thallium and other similar indicator dyes resulting in poor dye retention.

2 The TRS reagent is useful for suppressing background resulting from fluorescent compounds in the solutions surrounding the cells. Use of TRS may result in improved signal-to-noise.

3 The concentration of thallium in the stimulus buffer will vary by application. Investigators are encouraged to explore final thallium concentrations ranging from 0.5 to 4.5 mM to achieve the best results. Some targets, typically voltage-gated channels, may benefit from the use of High Potassium Chloride-free Stimulus Solution (Reagent H). Investigators are encouraged to explore mixtures of different proportions of Chloride-free Stimulus Solution and High Potassium, Chloride-free Stimulus Solution to achieve the best results.

4 Data collection typically includes the addition of test compounds or additional reagents to the assay plate during the course of data acquisition. Volumes and concentrations of test compounds and additional reagents vary. Commonly used addition volumes are 5, 10, 20 μL for 384-well plates and 20, 40, and 80 μL/well for 96-well plates.

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