NanoMolar Nickel / Cobalt Assay Kit

Mfr.No.	NMA1000
Description	Nickel (Ni++) and cobalt (Co++) are essential metal ions in biological systems. Many enzymes such as methionine aminopeptidase and glucose isomerase contain cobalt. Some other enzymes such as ureases from bactaria and plants use nickel as a cofactor. Synthesis of Ni / Co enzymes and coenzyme B12 requires high-affinity uptake of the metal ions from natural environments. In bacteria, Ni and Co uptake is mediated by secondary transporters and ATP-binding cassette systems. Understanding the differences between cobalt and nickel transporters might lead to drug development for gastritis and peptic ulceration. The NanoMolar Nickel / Cobalt Assay Kit is for measurement of nanomolar concentrations of nickel or cobalt. The assay is based on the principle that binding the fluorescence dye NMA with nickel or cobalt ions results in decrease of the fluorescence intensity (emission 535 nm, excitation 485 nm). Other metal ions including Mg2+, Ca2+ , Zn2+, Mn2+ , Al3+ , Cu 2+ and Ag+ give much lower assay sensitivity. Chelators such EDTA and thiol compounds bind strongly some of the metal ions and should be avoided in the assay. The assay kit can be used for high-throughput measurements of nickel or cobalt concentrations in biological samples or environmental water samples. The kit can also be used for biochemical assays of enzyme assays associated with nickel or cobalt metabolism. The kit includes 300 µl of 100 x NMA dye. It is for 1000 assays using 384-well plates (30 µl of sample volume) or 500 assays using 96-well plates (60 µl of sample volume). Cuvettes may also be used for measurements.
Concentration measurement	Metal ions

Product Manual	Datasheet
Protocols Description	PROTOCOL Standard curve 1. Sample preparation: Prepare the metal (nickel sulfate or cobalt chloride) solutions at a series of concentrations ranging from 1 µM to zero in 10 mM HEPES, pH 7.5. Dilute the 100 x NMA dye 100-fold with water to make the 1 x NMA dye. 2. Detection: Mix 30 µl of the sample with 30 µl of the 1x NMA dye solution for 10 min. Read the fluorescence intensity (Fc) at 535 nm (excitation 485 nm). The fluorescence for the buffer or water without metal is Fo. 3. Data Analysis: Calculate the Fo / Fc values and plot the correlation between the Fo / Fc values and the metal ion concentrations [M] (Ni++ or Co++). Fo / Fc = a [M] + b Where the Fo / Fc and [M] values are from experimental data, the a and b values are from the linear fitting between the Fo / Fc values and the metal concentrations. UNKNOWN SAMPLES Follow the same procedure to measure the fluorescence intensity Fc values from the unknown samples and calculate the Fo / Fc values. Calculate the metal ion concentrations [M] in the unknown samples using the Fo / Fc values from the unknown samples and the a and b values from the standard curve. [M] = (Fo / Fc – b) / a