1. Analytical profile index
API 20E

2. The microorganisms to be identified must first be isolated on a culture medium adapted to the culture of Enterobacteriaceae and/or non-fastidious Gram-negative rods, according to standard microbiological techniques.

3. API 20E system
The API 20E is a standardized identification system for Enterobacteriaceae depending on the metabolic activity of bacteria.
The API 20 E strip consists of 20 microtubes (each one of microtubes have cupule, and a small tube ) containing dehydrated substrates.

4. Metabolism of carbohydrates
The API system
View of API 20E just after seeding…
Nitrogen metabolisms
and specific enzymes
Metabolism of carbohydrates
24h / 37°C…..

5. Materials NaCl 0.85 % Medium, 5 ml or API Suspension Medium, 5 ml
API 20 E reagent kit
TDA
JAMES
VP 1 + VP 2
NIT 1 + NIT 2
Zn reagent
Oxidase
Mineral oil
TDA: detection of tryptophan deaminase ( FeCL3+ demineralized water).
JAMES: detection of indole (paradimethylaminobenzaldehyd).
VP 1 + VP 2: detection of acetion (KOH+ alpha naphthol).
NIT 1 + NIT 2: detection of nitrites reduction (H2SO4+dimethyl naphthyamine).
Oxidase: tetramethyl-p-phenylenediamine.

6. Preparation of the inoculum
Open an ampule of API NaCl 0.85 % Medium (5 ml) or use any tube containing 5 ml of sterile saline or sterile distilled water, without additives.
Use colony from an isolation plate. It is recommended to use young cultures (18-24 hours old).
Carefully emulsify to achieve a homogeneous bacterial suspension.
This suspension must be used immediately after preparation.

7. 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.
Vortex mix the saline suspension to get uniform dispersal
Inoculate all the tubes on the test strip with the pipette by depositing the suspension into the cupules as you tilt the API tray

8. The API system
Introducing the API 20E system for Enterobacteria identification…
5 mL of sterile water
1 bacterial colony
Fill microtubes gallery with the bacterial suspension

10. Important
Create anaerobiosis in the tests ADH, LDC, ODC, H2S and URE. by overlaying with mineral oil.
Since the media in |CIT|, |VP|, and |GEL| compartments require oxygen, completely fill both the cupule and tube of these compartments.
Fill only the tube (and not the cupule) of the other tests.
Close the incubation box. Incubate at 36°C ± 2°C for hours.
ONPG: Permase enzyme, used to enter lactose (lactose positive must be ONPG positive and not vice versa, like vibrio ONPG positive but lactose negative )

12. The API system
API 20 E after incubation…Positive results for all tests :
API 20 E after incubation…Negative results for all tests :

14. The API system Example of results for bacteria to test :
1-Reading results :
2-Entering results in the database software:

15. The API system The most used API system… API Strep API Staph API 20NE
 Identification of Streptococcus species
API Staph
 Identification of Staphylococcus species
API 20NE
 Identification of Non Enterobacteria (Pseudomonas for example)
API 20E
 Identification of Enterobacteria