Amplite® Colorimetric Zinc Ion Quantitation Kit

1. Prepare Zn²⁺ Standards or test samples (50 µL)
2. Add Zinc working solution (50 µL)
3. Incubate at room temperature for 5 - 10 minutes
4. Read absorbance ratio of A_610nm/A_470nm

Thaw all kit components at room temperature before starting the experiment.

Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles

Add 10 µL of 100 mM ZnCl₂ Standard solution (Component C) into 990 µL Assay Buffer (Component B) to get 1 mM ZnCl₂ standard solution.

Add 25 μL of Zn-620™ (Component A) into 5 mL Assay Buffer (Component B) to make Zn working solution.

Table 1. Layout of ZnCl₂ standards and test samples in a clear bottom 96-well microplate. Zn= Zinc Standards (Zn1 - Zn7, 100 to 1.56 µM), BL=Blank Control, TS=Test Samples.

Table 2. Reagent composition for each well.

1. Dilute the test sample to 1.56 - 100 µM range with Assay Buffer (Component B).
2. Prepare ZnCl₂ standards (Zn), blank controls (BL), and test samples (TS) according to the layout provided in Tables 1 and 2. For a 384-well plate, use 25 µL of reagent per well instead of 50 µL.
3. Add 50 µL of Zn working solution to each well of ZnCl₂ standard, blank control, and test samples to make the total ZnCl₂ assay volume of 100 µL/well. For a 384-well plate, add 25 µL of Zn working solution into each well instead, for a total volume of 50 µL/well.
4. Incubate the reaction for 5 - 10 minutes at room temperature, protected from light.
5. Monitor the absorbance ratio increase with a absorbance plate reader at A_610nm/A_470nm.