Amplite® Fluorimetric Lead (II) Ion Quantitation Kit
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Telephone | 1-800-990-8053 |
Fax | 1-800-609-2943 |
sales@aatbio.com | |
International | See distributors |
Bulk request | Inquire |
Custom size | Inquire |
Shipping | Standard overnight for United States, inquire for international |
Intended use | Research Use Only (RUO) |
Overview | SDSProtocol |
Platform
Fluorescence microplate reader
Excitation | 490 nm |
Emission | 530 nm |
Cutoff | 515 nm |
Recommended plate | Solid black |
Components
Example protocol
AT A GLANCE
Protocol summary
- Prepare and add lead standards or test samples (50 uL)
- Prepare and add Lead Green working solution (50 uL)
- Incubate at room temperature for 10-30 minutes
- Monitor fluorescence intensity at Ex/Em = 490/530 nm
Important notes
Thaw all the kit components at room temperature before starting the experiment.
PREPARATION OF STOCK SOLUTION
Lead Green stock solution (200X):
Add 50 uL of DMSO (Component D) into the vial of Lead Green (Component A) to make Lead Green stock solution (200X). Note: Store the unused Lead Green stock solution at -20 °C in single use aliquots.
PREPARATION OF STANDARD SOLUTION
For convenience, use the Serial Dilution Planner: https://www.aatbio.com/tools/serial-dilution/19007
Add 10 uL of 100 mM Lead (II) Standard solution (Component C) to 990 uL H2O to generate 1 mM Lead standard solution. Take 1 mM Lead (II) standard solution to perform 1:3 serial dilutions by H2O to get serially diluted Lead (II) standards ranging from 0 to 1 mM.
PREPARATION OF WORKING SOLUTION
Lead Green working solution:
Add 25 uL Lead Green stock solution (200X) into 5 mL of Assay Buffer (Component B) to make a total volume of 5.025 mL. Note: Keep away from Light. Note: Lead Green working solution should be used promptly.
SAMPLE EXPERIMENTAL PROTOCOL
Table 1. Layout of lead standards and test samples in a solid black 96-well microplate. LS = Lead standard (LS1-LS7, 1000 to 1.4 uM); BL = blank control; TS = test sample.
BL | BL | TS | TS |
LS1 | LS1 | ... | ... |
LS2 | LS2 | ... | ... |
LS3 | LS3 | ||
LS4 | LS4 | ||
LS5 | LS5 | ||
LS6 | LS6 | ||
LS7 | LS7 |
Table 2. Reagent composition for each well.
Well | Volume | Reagent |
LS1-LS7 | 50 uL | serial dilution (1000 to 1.34 uM) |
BL | 50 uL | H2O |
TS | 50 uL | test sample |
- Prepare Lead Standards (LS), blank controls (BL), and test samples (TS) according to the layout provided in Tables 1 and 2. For a 384-well plate, use 25 µL of reagent per well instead of 50 µL.
- Add 50 uL of Lead Green working solution to each well of Lead Standard, blank control, and test samples to make the total Lead assay volume of 100 uL/well. For a 384-well plate, add 25 uL of working solution into each well instead for a total volume of 50 uL/well.
- Incubate the reaction at room temperature for 10 to 30 minutes, protected from light.
- Monitor the fluorescence intensity with a fluorescence plate reader at Ex/Em = 490/530 nm.
Images
Citations
Authors: Apte, A., Bradford, K., Dente, C., Smith, R. N.
Journal: J Trauma Acute Care Surg (2019): 707-716
Authors: Rana, M. N., Tangpong, J., Rahman, M. M.
Journal: Toxicol Rep (2018): 704-713
Authors: Kushwaha, A., Hans, N., Kumar, S., Rani, R.
Journal: Ecotoxicol Environ Saf (2018): 1035-1045
Authors: Grasso, I. A., Blattner, M. R., Short, T., Downs, J. W.
Journal: Mil Med (2017): e1843-e1848
Authors: Bustamante, N. D., Macias-Konstantopoulos, W. L.
Journal: J Emerg Med (2016): 45-9
Authors: Begly, J. P., Lajam, C. M.
Journal: Bull Hosp Jt Dis (2013) (2016): 229-33
Authors: Ashraf, U., Kanu, A. S., Mo, Z., Hussain, S., Anjum, S. A., Khan, I., Abbas, R. N., Tang, X.
Journal: Environ Sci Pollut Res Int (2015): 18318-32
Authors: Wani, A. L., Ara, A., Usmani, J. A.
Journal: Interdiscip Toxicol (2015): 55-64
Authors: Karrari, P., Mehrpour, O., Abdollahi, M.
Journal: Daru (2012): 2
Authors: Flora, G., Gupta, D., Tiwari, A.
Journal: Interdiscip Toxicol (2012): 47-58
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