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Home / Assay Kits / Concentration Measurement Assays

MicroMolar ADP Assay Kit

BHE16300242

This product includes 350 ul of 10 x Buffer, 30 ul of 100 x MAD reagent 1, 30 ul of 100 x MAD reagent 2 and 300 ul of 10 x fluorescence dye. The assay reagents are sufficient for measurement of 100 samples of ADP using a standard black 384-well plate.

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Product Manual Datasheet
Protocols Description PROTOCOL
The following assay protocol is based on using a 384-well plate (Matrix 4318 plate) for the measurement. The sample volume is 15 µl and the final assay volume is 30 µl. For 96-well plate assays, the sample volume is 50 µl and the final assay volume is 100 µl. For assays using cuvette, the sample volume is 500 µl and the final assay volume is 1000 µl.
 
STANDARD CURVE
1. Sample preparation:
Prepare 15 µl of ADP solutions in a 384-well black plate (Matrix 4318 plate) with a two-fold serial dilution from 0.005 mM to zero in water. Freshly prepare the following premix for each 10 assays: 136.8 µl of H2O, 36 µl of 10 x Buffer, 3.6 µl of 100 x MAD Reagent 1, 3.6 µl of 100 x MAD Reagent 2.
 
The 1 x Buffer is composed of 50 mM Tris-HCl, pH 8.0, 3 mM MgCl2, 0.2 mM EDTA, 0.5 mM DTT, 50 mM NaCl, 0.003% Brij-35.
 
2. Detection:
Mix 15 µl of the ADP samples with 15 µl of the premix for 45 min. Dilute 33µl of the 10 x Fluorescence dye with 297 µl of water (10-fold dilution). Add 30 µl of 1 x Fluorescence dye and read the fluorescence intensity at 535 nm with excitation at 485 nm in 5 min.
 
3. Data Analysis:
Plot the fluorescence intensity Fc and the ADP concentration [ADP] to generate the linear standard curve.
 
                                                                   Fc = a [ADP] + b
 
Where the Fc values are from experimental data, the a and b values are from the linear fitting between the Fc values and the ADP concentrations.
 
UNKNOWN SAMPLES
Follow the same procedure to measure the fluorescence intensity Fc values from the unknown samples. Calculate the ADP concentrations in the unknown samples using the Fc values from the unknown samples and the a and b values from the standard curve.
 
                                                                    [ADP] = (Fc – b) / a
 
Combination SKU Supplier No. Size Your price Quantity
BHE16300242-1 MAD100K 100 assays $392.60
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BHE16300242-2 MAD500K 500 assays $1495.62
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