QuantiChrom™ Leucine Aminopeptidase Assay Kit

Protocol SDS

Application

• For quantitative determination of LAP enzyme activity in biological samples, e.g. cells, tissues, and serum samples.

Key Features

• Sensitive and accurate. Use as little as 10 μL samples. Linear detection range in 96-well plate: 0.025 to 250 U/L.
• Fast and convenient. The procedure involves addition of a single working reagent and incubation for 20 min. The assay can be run at room temperature or 37°C.
• High-throughput. Homogeneous "mix-incubate-measure" type assay. Can be readily automated to assay thousands of samples per day.

Method

• OD405nm

Samples

• Cells, serum, and other biological samples

Species

• All

Procedures

• 20 min

Size

• 100 tests in 96-well plate

Detection Limit

• 0.025 U/L

Shelf Life

• 6 months

More Details

LEUCINE AMINOPEPTIDASE (LAP) (E.C 3.4.11.1) is an enzyme commonly found in liver cells and intestinal cells that hydrolyzes leucine from the N-terminus of proteins and polypeptides. The expression of LAP increases in damage and immune response, so increased LAP activity in serum samples can indicate potential liver damage, tumorigenesis, or inflammation.

BioAssay Systems' LAP activity assay provides a convenient colorimetric method to measure LAP activity in biological samples. In this assay, LAP hydrolyzes a dye-linked leucine substrate, resulting in a yellow, colored product, measurable at OD405nm. The increase in absorbance at 405 nm (ΔOD) in 20 minutes is directly proportional to the LAP activity.

Is leucine aminopeptidase enzyme included in the kit?

The customer will need to provide their own enzyme. Leucine Aminopeptidase, microsomal from porcine kidney (e.g Sigma, Cat# L6007) was used for the development of this kit.

I am using the enzyme you recommended to run this assay (Sigma, Cat# L6007). Did you prepare stock solutions in Assay Buffer or is there a better solvent to use?

We dissolved porcine LAP in 0.5M MgCl₂. According to BRENDA Enzyme Database, when using this solvent, porcine LAP should be stable indefinitely within pH 7-9.

When preparing Working Reagent, I noticed turbidity after the addition of substrate to Assay Buffer. Is this normal?

Yes, we have occasionally observed the same phenomenon; however, we found after thorough mixing by pipetting up and down at room temperature, the substrate fully dissolves. There has been no observable effect on the reaction from this. We recommend allowing all reagents to come to room temperature prior to preparing Working Reagent. This should help to limit possible turbidity when preparing the assay.

For more detailed product information and questions, please feel free to email or call us at 1-510-782-9988 x 2. For more general information regarding our assays, please refer to Technical Support.

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If you or your labs do not have the equipment or scientists necessary to run this assay, BioAssay Systems can perform the service for you.

– Fast turnaround
– Quality data
– Low cost

Please email or call 1-510-782-9988 x 2 to discuss your projects.