Leucine Aminopeptidase (LAP) Test

Last updated on June 23rd, 2021

Leucine Aminopeptidase (LAP) test is used to identify catalase-negative gram-positive cocci. Specifically, it differentiates Aerococcus and Leuconostoc from Streptococcus, Enterococcus, Lactococcus, and Pediococcus.

The LAP disk is a rapid test for the detection of the enzyme leucine aminopeptidase. Leucine-beta-naphthylamide impregnated disk serves as a substrate for the detection of leucine aminopeptidase. Following hydrolysis of the substrate by enzyme, the resulting beta- naphthylamine produces a red color upon the addition of cinnamaldehyde reagent.

Quality control

Perform QC on each new lot or shipment of LAP disks and color reagent prior to using them. Since the test is performed infrequently and most Gram-positive cocci are LAP positive, verify potency by testing the positive control each time a negative test is obtained by using test organisms.

  1. Enterococcus faecalis ATCC 29212—LAP positive
  2. Aerococcus viridans ATCC 11563—LAP negative

Method

  1. Before incubation, slightly dampen the LAP disk with reagent-grade water. Do not supersaturate the disk.
  2. Using a wooden applicator stick, rub a small amount of several colonies of an 18 to 24-hour pure culture onto a small area of the LAP disk.
  3. Incubate at room temperature for 5 minutes.
  4. After the incubation period, add 1 drop of cinnamaldehyde reagent.
LAP Test positive
Disk showing LAP test positive

Expected results

  1. Positive: Development of a deep red to reddish-purple color within 1 minute after adding cinnamaldehyde reagent. Weak reactions are pink.
  2. Negative: No color change or development of a slight yellow color.

LAP Test Results

LAP test is only one of a battery of useful tests for identifying catalase-negative, Gram-positive cocci (most Gram-positive cocci are LAP positive so not tested). LAP test result of some catalase-negative, Gram-positive cocci is as follows:

  • LAP negative: Leuconostoc
  • LAP positive: Streptococcus, Enterococcus, Lactococcus, and Pediococcus
  • LAP variable: Aerococcus

Limitations

  1. LAP test is one of the battery of useful tests for identifying catalase-negative, Gram-positive cocci and is most helpful when kit tests yield uncommon identifications or Gram stain results do not show cocci in chains.
  2. If the inoculum is insufficient, false-negative results may be seen.

References and further readings

  1. Clinical Microbiology Procedures Handbook, Fourth Edition. (2016). In Clinical Microbiology Procedures Handbook, Fourth Edition. American Society of Microbiology. https://doi.org/10.1128/9781555818814
  2. Color Atlas and Textbook of Diagnostic Microbiology, Koneman, 5th edition
About Acharya Tankeshwar 473 Articles
Hello, thank you for visiting my blog. I am Tankeshwar Acharya. Blogging is my passion. I am working as an Asst. Professor and Microbiologist at Department of Microbiology and Immunology, Patan Academy of Health Sciences, Nepal. If you want me to write about any posts that you found confusing/difficult, please mention in the comments below.