Dr.ayham abulaila

API 20e

Dr.ayham abulaila

The API 20E System The API 20E System is miniaturized

version of conventional tests that is used for the identification of members of the family Enterobacteriaceae and other gram-negative bacteria.

This system utilizes a plastic strip with 20 separate compartments.

Each compartment consists of a depression, or cupule, and a small tube that contains a specific dehydrated medium

Dr.ayham abulaila

Dr.ayham abulaila

Inculcation To inoculate each compartment it is necessary to first

make up a saline suspension of the unknown organism; then, with syring, each compartment is filled with the bacterial suspension.

The cupule receives the suspension and allows it to flow into the tube of medium.

The dehydrated medium is reconstituted by the saline.

To provide anaerobic conditions for some of the compartments it is necessary to add sterile mineral oil to them.

After incubation for 18–24 hours, the reactions are recorded, test reagents are added to some compartments, and test results are tabulated.

Dr.ayham abulaila

Dr.ayham abulaila

Enterobacteriaceae Identification API 20E : FIRST PERIOD

Two things will be accomplished during this period:(1) the oxidase test will be performed if it

has not been previously performed (2) the API 20E test strip will be inoculated.

Dr.ayham abulaila

Materials: agar slant or plate culture of unknown test tube of 5 ml 0.85% sterile saline API 20E test strip API incubation tray and cover squeeze bottle of tap water test tube of 5 ml sterile mineral oil Pasteur pipettes (5 ml size) oxidase test reagent Whatman no. 2 filter paper empty Petri dish Vortex mixer

Dr.ayham abulaila

FIRST PERIOD

1. If you haven’t already done the oxidase test on your unknown, do so at this time.

It must be established that your unknown is definitely oxidase negative before using this system.

2. Prepare a saline suspension of your unknown by transferring organisms from the center of a well-established colony on an agar plate (or from aslant culture) to a tube of 0.85% saline solution.

Dr.ayham abulaila

Dr.ayham abulaila

FIRST PERIOD

3. Dispense about 5 ml of tap water into the tray with a squeeze bottle. Note that the bottom of the tray has numerous depressions to accept the water.

4. Vortex mix the saline suspension to get uniform dispersal

5. Inoculate all the tubes on the test strip with the pipette by depositing the suspension into the cupules as you tilt the API tray

Dr.ayham abulaila

Dr.ayham abulaila

Important: Slightly under fill ADH, LDC, ODC, H2S,and

URE. (Note that the labels for these compartments are underlined on the strip.) Underfilling these compartments leaves room for oil to be added and facilitates interpretation of the results.

Since the media in |CIT|, |VP|, and |GEL| compartments require oxygen, completely fill both the cupule and tube of these compartments. Note that the labels on these three compartments are bracketed as shown here.

Dr.ayham abulaila

Dr.ayham abulaila

Second PERIOD: reading the results

Each chemical test give specific color The result readied as number of 7

digits This No is matched with tables or with

computer program to identify the bacteria

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