Introduction of LAP (Leucine Aminopeptidase) Test
Table of Contents
LAP stands for leucine aminopeptidase which is an enzyme useful for deﬁnitive identiﬁcation of catalase-negative, gram-positive cocci of Aerococcus and Leuconostoc (LAP test negative) from other organisms in the group like Streptococcus, Enterococcus, Lactococcus, and Pediococcus (almost always positive).
Principle of LAP test
Hydrolysis of the leucine-pnaphthylamide substrate by the enzyme LAP on a paper disk inoculated with the bacteria like Streptococcus, Enterococcus, Lactococcus, and Pediococcus releases leucine and free b-naphthylamide. The bnaphthylamide combines with cinnamaldehyde reagent to form a pink to cherry-red color.
Requirements for LAP (Leucine Aminopeptidase) Test
- Test organisms -Fresh growth less than 24 hours old of catalase-negative, gram-positive cocci. If growth is poor, a culture that has been incubated for up to 72 hours may be used.
- LAP Disk
- 0.01% p-Dimethylaminocinnamaldehyde (the same reagent as used in the PYR test)
- Sterile loop or stick
- Petri dish or slide
- Control strains
Positive Control (PC): Enterococcus faecalis ATCC 29212
Negative Control (NC): Aerococcus viridans ATCC 11563
Procedure of LAP Test
- Place the disk onto a petri dish or a slide.
- Moisten (do not saturate the disk) with a loopful of distilled water.
- Using a sterile stick or loop, smear with the suspected isolate.
- Incubate at room temperature for 5 minutes.
- Add a drop of cinnamaldehyde reagent and wait 2 minutes to observe the color.
Result and Interpretation of LAP (Leucine Aminopeptidase) Test
LAP test positive: Development of a deep red to reddish-purple after the addition of cinnamaldehyde reagent
LAP test Negative: No color change or development of a yellow color
Enterococcus faecalis ATCC 29212: Positive
Aerococcus viridans ATCC 11563: Negative
LAP (Leucine Aminopeptidase) Test-Positive and Negative Bacteria
Limitations of the LAP Test
- False negatives may result from insufﬁcient inoculum.
- This test is only one of a 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 chaining cocci.
- Cowan and Steel’s, manual for the identification of medical bacteria
- Lynne S. Garcia, Clinical Microbiology Procedures Handbook