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Description
Caspase 4 Activity Detection KitProduct Specification Usage Determination of pNA standard curve: 1. The 10 mM pNA standard was diluted with reaction buffer to 200, 100, 50, 25, 12. 5, 6. 25, 3. 125 M. Set the zero tube without pNA. 2. Take 100l of each concentration and add it to a 96 well plate or 100l cuvette, and measure the absorbance OD value at 405nm. 3. The OD405 value of each concentration standard tube is the x axis, and the corresponding pNA concentration is the y axis.
Product Specification
| Usage | Determination of pNA standard curve: 1. The 10 mM pNA standard was diluted with reaction buffer to 200, 100, 50, 25, 12.5, 6.25, 3.125 μM. Set the zero tube without pNA. 2. Take 100μl of each concentration and add it to a 96-well plate or 100μl cuvette, and measure the absorbance OD value at 405nm. 3. The OD405 value of each concentration standard tube is the x-axis, and the corresponding pNA concentration is the y-axis. Use Excel to make a standard curve of pNA concentration versus OD405 value. Detection of Caspase 4 enzyme activity: 1. Take out an appropriate amount of Ac-LEVD-pNA (2mM) and place it on an ice bath for later use. 2. According to the 100ul reaction system: blank control group: 40μl of detection buffer, 0μl of sample to be tested, 50μl of lysate, 10μl of Ac-LEVD-pNA (2mM); Sample group: 40μl of detection buffer, 50μl of sample to be tested, 0μl of lysate, 10μl3 of Ac-LEVD-pNA (2mM). Add Ac-LEVD-pNA (2mM) and mix well. Be careful to avoid bubbles during mixing. Incubate at 37 º C for 60-120 minutes. A405 can be measured when the color change is obvious. If the color change is not obvious, the incubation time can be appropriately extended, or even overnight. 4. The A405 of the sample minus the A405 of the blank control is the absorbance produced by pNA catalyzed by caspase 4 in the sample. By comparing the standard curves obtained in the above steps, it is possible to calculate the amount of pNA catalytically produced in the sample. |
| Theory | Based on Caspase 4, it can catalyze the substrate Leu Glu Val Asp p nitroanilide (Ac LEVD pNA) to produce yellow pNA (p-nitroaniline), which can be used to detect the activity of Caspase 4 by measuring absorbance. pNA has a maximum absorbance at 405 nm. Caspase activity was obtained by measuring pNA using visible light colorimetry. Suitable for detecting Caspase 4 activity in mammalian tissues and cells. |
| Experiment Reagent Instrument | Microplate reader or spectrophotometer |
| Synonym | caspase-4,caspase 4 |
| Description | caspase 4 Activity Detection Kit is a kit that detects caspase 4 enzyme activity or purified caspase 4 enzyme activity in cell or tissue lysates by spectrophotometry. |
| Storage Temp. | When stored at-20 °C, Ac-LEVD-pNA and pNA need to be stored in the dark. |
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