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Amplite® Colorimetric Lipase Activity Assay Kit
Example protocol
AT A GLANCE
Prepare test samples and the lipase standards (50 μL).
Add the lipase working solution (50 µL).
Incubate at 37 °C for 10-30 minutes.
Measure the absorbance at 586 nm.
PREPARATION OF STOCK SOLUTIONS
Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles
Reconstitute Lipase Standard (Component D) by adding 100 µL of ddH2O to achieve a concentration of 1 mg/mL. Mix thoroughly by pipetting up and down several times.
Note: The Lipase Standard Stock Solution can be stored at -20 °C, protected from light and should be used within 1 month after reconstitution.
PREPARATION OF STANDARD SOLUTIONS
https://www.aatbio.com/tools/serial-dilution/11315
PREPARATION OF WORKING SOLUTION
Add 200 µL of Lipase Substrate (Component C) to 5 mL of Lipase Substrate Buffer (Component B). This 5 mL solution is sufficient for 100 tests. Prepare the required amount of Lipase Working Solution proportionally based on the number of tests you need.
Tissues and cells can be homogenized in the Lipase Assay Buffer (Component A). To remove insoluble material, centrifuge the sample at 13,000xg for 10 minutes. Serum samples can be added directly to the wells. Adjust the samples to a final volume of 50 µL using Lipase Assay Buffer.
SAMPLE EXPERIMENTAL PROTOCOL
Table 1. Layout of lipase standards and test samples in a 96-well clear bottom microplate. (STD = Lipase Standards (STD1-STD7, 0.625~40 µg/ml), BL= Blank Control, TS=Test Samples)
BL | BL | TS | ... |
STD 1 | STD 1 | ... | ... |
STD 2 | STD 2 | ... | ... |
STD 3 | STD 3 | ||
STD 4 | STD 4 | ||
STD 5 | STD 5 | ||
STD 6 | STD 6 | ||
STD 7 | STD 7 |
Table 2. Reagent composition for each well.
Well | Volume | Reagent |
Lipase STD 1- STD 7 | 50 µL | Serial Dilutions (0.625-40 µg/mL) |
BL | 50 µL | Assay Buffer |
TS | 50 µL | Test Sample |
Prepare lipase standards (STD1-7), blank controls (BL), and test samples (TS) as outlined in Tables 1 and 2. For a 384-well plate, use 25 µL of reagent per well.
Add 50 µL of Lipase Working Solution to each well containing the blank control, Lipase Standards, and test samples (TS). If using a 384-well plate, add 25 µL of Lipase Working Solution to each well instead.
Incubate at 37 °C for 10-30 minutes, protected from light.
Monitor the absorbance at 586 nm.
References
Authors: Putra, Sharifah Shahira Syed and Chew, Chia Yong and Hayyan, Adeeb and Elgharbawy, Amal A M and Taskin-Tok, Tugba and Hayyan, Maan and Ngoh, Gek Cheng and Saleh, Jehad and Al Abdulmonem, Waleed and Alghsham, Ruqaih S and Nor, Mohd Roslan Mohd and Aldaihani, Ahmad G H and Basirun, Wan Jefrey
Journal: International journal of biological macromolecules (2024): 132245
Authors: Singh, Alka and Abhilasha, Kandahalli Venkataranganayaka and Acharya, Kathya R and Liu, Haibo and Nirala, Niraj K and Parthibane, Velayoudame and Kunduri, Govind and Abimannan, Thiruvaimozhi and Tantalla, Jacob and Zhu, Lihua Julie and Acharya, Jairaj K and Acharya, Usha R
Journal: Nature communications (2024): 4410
Authors: Xie, Xiaochun and Liu, Yang and Yang, Qi and Ma, Xiaojie and Lu, Yingying and Hu, Yuepeng and Zhang, Guofu and Ke, Lu and Tong, Zhihui and Liu, Yuxiu and Xue, Jing and Lu, Guotao and Li, Weiqin
Journal: The American journal of pathology (2024)
Authors: Kim, Jin-Kyung and Hwang, Sun Young and Kim, Se Eun and Lee, Gahyun and Ji, Soungjin and Kim, Jungho and Kim, Yongbaek
Journal: Journal of veterinary science (2024): e48
Authors: Rezaie, Hosna and Kajani, Abolghasem Abbasi and Jafarian, Faranak and Asgari, Sina and Taheri-Kafrani, Asghar and Bordbar, Abdol-Khalegh
Journal: Journal of biotechnology (2024): 23-31
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