MicroMolar Cisplatin Assay Kit

Mfr.No.	CPT200
Description	Cisplatin [or cis-diamminedichloroplatinum, Pt(NH3)2Cl2] is the first member of platinum-containing anti-cancer drugs. It binds DNA and causes DNA crosslinking which ultimately triggers apoptosis (programmed cell death). The MicroMolar Cisplatin Assay Kit (Catalog number CPT200) is designed for high throughput measurement of micromolar concentrations of Cisplatin. The assay is based on the light absorbance at 535 nm. The assay kit can be used for assays of cisplatin in drug discovery, drug development, pharmaceutical samples and biological samples. The MicroMolar Cisplatin Assay Kit (Catalog number CPT200) includes 4 ml of Reagent A, 400 µl of 10 x Reagent B, and 2 ml of 10 x Reagent C. It is for 200 assays using 96-well plates. Cuvettes may also be used for measurements.
Concentration measurement	Drug molecules

Product Manual	Datasheet
Protocols Description	ASSAY PROTOCOL The following assay protocol is based on using a 96-well plate for the measurement. The sample volume is 100 µl and the final assay volume is 240 µl. The assay reactions and measurement are on a transparent 96-well plate. It is a high throughput assay format. For assays using cuvette, the sample volume is 400 µl and the final assay volume is 960 µl. The assay reactions are carried out in eppendorf tubes or test tubes. The samples are transferred into a cuvette for measurement of the light absorbance. STANDARD CURVE 1. Sample preparation: Prepare 100 µl of cisplatin solutions in the wells of a transparent 96-well plate with a two-fold serial dilution from 0.1 mg/ml to zero in water. For each 10 assays, dilute 20 µl of 10 x Reagent B with 180 µl of water to make 1 x Reagent B; dilute 100 µl of the 10 x Reagent C with 0.9 ml of water to make 1 x Reagent C. Note: If the 10 x Reagent B solution forms crystals during storage in a freezer, warm the solution with hands or water batch until the crystal in completely dissolved before preparing the 1 x Reagent B solution. 2. Assay: Into the wells with 100 µl of cisplatin solutions, add 20 µl of Reagent A and 20 µl of 1 x Reagent B. Incubate the mixtures at 65°C for 30 min. Add 100 µl of 1 x Reagent C into each well. Read the light absorbance at 535 nm (A₅₃₅) 3. Data Analysis: Plot the A₅₃₅values and the Cisplatin concentration [Cisplatin] to generate the linear standard curve. A₅₃₅ = a [Cisplatin] + b Where the A₅₃₅ values are from experimental data, the a and b values are from the linear fitting between the A₅₃₅ values and the Cisplatin concentrations. UNKNOWN SAMPLES Follow the same procedure to measure the light absorbance A₅₃₅values from the unknown samples. Calculate the Cisplatin concentrations in the unknown samples using the A₅₃₅ values from the unknown samples and the a and b values from the standard curve. [Cisplatin] = (A₅₃₅– b) / a