Product Details
abID | ab83355 |
---|---|
Cat # +Size | K354-100 |
Size | 100 assays |
Detection Method | Absorbance (570 nm) or Fluorescence (Ex/Em 535/587 nm) |
Species Reactivity | All |
Applications | The assay can detect as low as 50 picomol (1 µM) of ATP in various samples. |
Features & Benefits | • Simple procedure; takes less than 1 hour • Fast and convenient • Kit is designed to be a robust method which utilizes the phosphorylation of glycerol to generate a product that is easily quantified. |
Kit Components | • ATP Assay Buffer • ATP Probe (in DMSO) • ATP Converter • Developer Mix (lyophilized) • ATP Standard (1 µmol; lyophilized) |
Storage Conditions | -20°C |
Shipping Conditions | Gel Pack |
USAGE | For Research Use Only! Not For Use in Humans. |
Details
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1. Protein Precipitation: For biological samples with protein concentration less than ~ 20 mg/ml (tissue homogenate, cell lysate, urine, etc.), take 500 µl of sample, mix with 100 µl of ice cold PCA in 1.5 ml microfuge tubes, vortex briefly to mix well, place on ice for 5 min. Centrifuge at 13,000 x g for 2 min. Accurately transfer 480 µl of the supernatant to a fresh tube. For serum and other very high protein concentration samples, take 400 µl of sample and mix with 100 µl of ice-cold PCA, place on ice for 5 min. Centrifuge at 13,000 x g for 2 min. Accurately transfer 380 µl of the supernatant to a fresh tube. Depending on the nature of the analyte, the samples in PCA may be frozen at -70°C for up to a month for storage at this stage.
2. Sample Neutralization:Add 20 µl of ice-cold Neutralization Solution (resuspend the fine precipitate) and mix to neutralize the sample and precipitate excess PCA. There may be some gas (CO2) evolution so vent the sample tube. Place on ice for 5 min. Spin briefly (~ 1 - 2 min). Samples are now deproteinized, neutralized, and PCA has been removed. The samples may now be used in a variety of assays directly.As you can see, the general ratio for 1 N PCA vs 6 N KOH is 5:1. But, I would recommend a pH check after neutralization to see if it is within the range of 6.5 to 8.0. If not, you might need to add more KOH.