Amplite® Fluorimetric Lipase Activity Assay Kit

The Amplite® Fluorimetric Lipase Activity Assay Kit provides a simple and quick protocol for measuring lipase activity. This assay leverages a coupled enzymatic reaction that converts a substrate into a fluorescent product (Ex/Em=540/610 nm), with fluorescence intensity proportional to the lipase activity present in the sample. This kit is suitable for detecting lipases from various biological samples like cells, supernatants, tissue extracts, or serum, and is adaptable for high-throughput screening applications. Lipases belong to a class of enzymes that catalyze the hydrolysis of ester bonds in lipids, such as triglycerides, phospholipids, and cholesterol esters. These enzymes are essential for the digestion, absorption, and metabolism of dietary fats in the body, playing crucial a role in lipid metabolism and cell signaling. Lipases are produced by various organs and tissues, including the pancreas, liver, intestine, and adipose tissue. Accurate measurement of lipase activity is critical in screening and diagnosis of diseases like pancreatitis, cystic fibrosis, celiac disease, Crohn's disease, and hyperlipidemia.

Example protocol

AT A GLANCE

Protocol Summary

Prepare test samples and the lipase standards (50 μL).
Add the lipase working solution (50 µL).
Incubate at 37 °C for 10-30 minutes.
Monitor the fluorescence intensity at Ex/Em = 540/610 nm (Cutoff = 570 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

Lipase Standard Stock Solution

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

For convenience, use the Serial Dilution Planner:
https://www.aatbio.com/tools/serial-dilution/11322

Lipase Standard

Add 20 μL of Lipase Standard Stock Solution to 480 μL of Lipase Assay Buffer (Component A) to create a 40 μg/mL lipase standard solution (STD7). Take 250 μL of the STD7 solution and perform 2X serial dilutions in Lipase Assay Buffer (Component A) to produce a series of diluted lipase standards, labeled STD6 to STD1.

PREPARATION OF WORKING SOLUTION

Lipase 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.

Test Samples

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 fluorescence intensity at Ex/Em = 540/610 nm (Cutoff = 570 nm).

References

View all 50 references: Citation Explorer

The effect of konjac glucomannan on enzyme kinetics and fluorescence spectrometry of digestive enzymes: An in vitro research from the perspective of macromolecule crowding.
Authors: Chen, Wenjing and Li, Sha and Albahi, Amgad and Ye, Shuxin and Li, Jing and Li, Bin
Journal: Food research international (Ottawa, Ont.) (2024): 114247

Green-emitting carbon dots as a "turn on" fluorescence bio-probe for highly sensitive and selective detection of lipase in human serum.
Authors: Al-Mashriqi, Haitham Saad and Sanga, Pascaline and Chen, Jia and Li, Xin and Xiao, Jing and Li, Yan and Qiu, Hongdeng
Journal: Analytical and bioanalytical chemistry (2024): 971-981

A pH-Sensitive Double Chromophore Fluorescent Dye for Live-Tracking of Lipophagy.
Authors: Engelhardt, Pascal M and Veronese, Matteo and Eryiğit, Alpay A and Das, Anushka and Kaczmarek, Alexander T and Rugarli, Elena I and Schmalz, Hans-Günther
Journal: Chemistry (Weinheim an der Bergstrasse, Germany) (2024): e202400808

Bioconversion of waste glycerol into viscosinamide by Pseudomonas fluorescens DR54 and its activity evaluation.
Authors: Jama, Dominika and Łaba, Wojciech and Kruszelnicki, Mateusz and Polowczyk, Izabela and Lazar, Zbigniew and Janek, Tomasz
Journal: Scientific reports (2024): 1531

A turn-on fluorescence sensor for detection of heparinase with heparin templated aggregation of tetracationic porphyrin derivative.
Authors: Pandey, Shrishti P and Singh, Prabhat K and Jha, Pamela and Jobby, Renitta
Journal: International journal of biological macromolecules (2023): 125934

Page updated on May 19, 2025

Ordering information

Price
Unit size
Catalog Number	11322
Quantity

Add to cart

Additional ordering information

Telephone	1-800-990-8053
Fax	1-800-609-2943
Email	sales@aatbio.com
International	See distributors
Bulk request	Inquire
Custom size	Inquire
Technical Support	Contact us
Purchase order	Send to sales@aatbio.com
Shipping	Standard overnight for United States, inquire for international