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® 20 E TM 07584J - En - 2010/05 20 100 / 20 160 07584J - en - 2010/05 ® TM IVD 20 E Identification system for Enterobacteriaceae and other non-fastidious Gram-negative rods SUMMARY AND EXPLANATION Material API 20 E is a standardized identification system for - Pipettes or PSIpettes Enterobacteriaceae and other non-fastidious, Gram- - Ampule protector negative rods which uses 21 miniaturized biochemical - Ampule rack tests and a database. The complete list of those - General microbiology laboratory equipment organisms that it is possible to identify with this system is given in the Identification Table at the end of this package POSSIBLE ADDITIONAL REAGENTS insert. - API OF Medium (Ref. 50 110) : Test for the determination of fermentative or oxidative PRINCIPLE metabolism. The API 20 E strip consists of 20 microtubes containing - API M Medium (Ref. 50 120) : dehydrated substrates. These tests are inoculated with Test for motility of facultative anaerobic bacteria. a bacterial suspension that reconstitutes the media. During incubation, metabolism produces color changes WARNINGS AND PRECAUTIONS that are either spontaneous or revealed by the addition of • For in vitro diagnostic use and microbiological reagents. control. The reactions are read according to the Reading Table • For professional use only. and the identification is obtained by referring to the • This kit contains products of animal origin. Certified Analytical Profile Index or using the identification software. knowledge of the origin and/or sanitary state of the animals does not totally guarantee the absence of CONTENT OF THE KIT transmissible pathogenic agents. It is therefore Kit for 25 tests (ref. 20 100) recommended that these products be treated as - 25 API 20 E strips potentially infectious, and handled observing the usual - 25 incubation boxes safety precautions (do not ingest or inhale). - 25 result sheets • All specimens, microbial cultures and inoculated - 1 clip seal products should be considered infectious and handled - 1 package insert appropriately. Aseptic technique and usual precautions for handling the bacterial group studied should be Kit for 100 tests (ref. 20 160) observed throughout this procedure. Refer to - 100 API 20 E strips (4x25 strips) "CLSI® M29-A, Protection of Laboratory Workers from - 100 incubation boxes Occupationally Acquired Infections; Approved Guideline - 100 result sheets - Current revision". For additional handling precautions, - 1 clip seal refer to "Biosafety in Microbiological and Biomedical - 1 package insert Laboratories - CDC/NIH - Latest edition", or to the regulations currently in use in each country. COMPOSITION OF THE STRIP • Do not use reagents past the expiry date. • Before use, check that the packaging of the various The composition of the API 20 E strip is given in the components is intact. Reading Table of this package insert. • Do not use strips which have been damaged : cupules deformed, desiccant sachet open, etc. REAGENTS AND MATERIAL REQUIRED BUT NOT • The performance data presented were obtained using PROVIDED the procedure indicated in this package insert. Any Reagents change or modification in the procedure may affect the - API NaCl 0.85 % Medium, 5 ml (Ref. 20 230) or results. API Suspension Medium, 5 ml (Ref. 20 150) • Interpretation of the test results should be made taking - API 20 E reagent kit (Ref. 20 120) or into consideration the patient history, the source of the individual reagents : TDA (Ref. 70 402) specimen, colonial and microscopic morphology of the JAMES (Ref. 70 542) strain and, if necessary, the results of any other tests VP 1 + VP 2 (Ref. 70 422) performed, particularly the antimicrobial susceptibility NIT 1 + NIT 2 (Ref. 70 442) patterns. - Zn reagent (Ref. 70 380) - Oxidase (Ref. 55 635*) * reference not sold in certain countries : use an equivalent reagent. - Mineral oil (Ref. 70 100) - API 20 E Analytical Profile Index (Ref. 20 190) or apiweb TM identification software (Ref. 40 011) (consult bioMérieux) bioMérieux SA English - 1 api® 20 E TM 07584J - en - 2010/05 STORAGE CONDITIONS Inoculation of the strip The strips are supplied in an aluminum pouch with • With the same pipette, distribute the bacterial desiccant sachets. suspension into the tubes of the strip (to avoid the Once opened (*), the pouch should be re-sealed using the formation of bubbles at the base of the tubes, tilt the clip seal (included in the kit) to preserve the remaining strip slightly forward and place the tip of the pipette or strips with the desiccant sachets : place the open end of PSIpette against the side of the cupule): the pouch along the seal and carefully clamp between - For the CIT , VP and GEL tests, fill both tube and the two parts. The strips may then be kept for up to cupule, 10 months after the pouch has been opened, at 2-8°C - For the other tests, fill only the tubes (and not the (or until the expiry date indicated on the packaging, if this cupules), comes before). - for the tests ADH, LDC, ODC, H2S and URE, create (*) Recommended method for opening the pouches : cut anaerobiosis by overlaying with mineral oil. open the pouch just below the seal while holding the • Close the incubation box. pouch upright, in order to avoid damaging the desiccant • Incubate at 36°C ± 2°C for 18-24 hours. sachets. READING AND INTERPRETATION SPECIMENS (COLLECTION AND PREPARATION) Reading the strip API 20 E is not for use directly with clinical or other • After the incubation period, read the strip by referring to specimens. the Reading Table. The microorganisms to be identified must first be iso- • If 3 or more tests (GLU test + or –) are positive, record lated on a culture medium adapted to the culture of all the spontaneous reactions on the result sheet and Enterobacteriaceae and/or non-fastidious Gram-negative then reveal the tests which require the addition of rods, according to standard microbiological techniques. reagents : INSTRUCTIONS FOR USE - TDA Test : add 1 drop of TDA reagent. A reddish Oxidase test brown color indicates a positive reaction to be recorded on the result sheet. The oxidase test must be performed according to the - IND Test : add 1 drop of JAMES reagent. A pink color manufacturer's instructions for use. The result should be developed in the whole cupule indicates a positive recorded on the result sheet as it is an integral part of the reaction to be recorded on the result sheet. final profile (21st identification test). - VP Test : add 1 drop each of VP 1 and VP 2 reagents. Preparation of the strip Wait at least 10 minutes. A pink or red color indicates • Prepare an incubation box (tray and lid) and distribute a positive reaction to be recorded on the result sheet. about 5 ml of distilled water or demineralized water If a slightly pink color appears after 10 minutes, the [or any water without additives or chemicals which may reaction should be considered negative. NOTE : The indole production test must be performed release gases (e.g., Cl2, CO2, etc.)] into the honey- combed wells of the tray to create a humid atmosphere. last since this reaction releases gaseous products which • Record the strain reference on the elongated flap of the interfere with the interpretation of other tests on the tray. (Do not record the reference on the lid as it may be strip. The plastic incubation lid should not be replaced misplaced during the procedure). after the addition of the reagent. • Remove the strip from its packaging. • If the number of positive tests (including the GLU test) • Place the strip in the incubation box. before adding the reagents is less than 3 : NOTE : API 20 E should only be used with - Reincubate the strip for a further 24 hours (± 2 hours) Enterobacteriaceae and/or non-fastidious Gram-negative without adding any reagents. rods. Fastidious organisms having demanding nutritional - Reveal the tests requiring the addition of reagents requirements and requiring appropriate handling (see previous paragraph). precautions (i.e., Brucella and Francisella) are not - To complete the identification, it may be necessary to included in the API 20 E database. Alternative procedures perform supplementary tests (refer to Identification must be used to exclude or confirm their presence. paragraph). Preparation of the inoculum Interpretation • Open an ampule of API NaCl 0.85 % Medium (5 ml) or Identification is obtained with the numerical profile. an ampule of API Suspension Medium (5 ml) as • Determination of the numerical profile : indicated in the paragraph "Warnings and Precautions" On the result sheet, the tests are separated into groups of the package insert for these products, or use any tube of 3 and a value 1, 2 or 4 is indicated for each. By containing 5 ml of sterile saline or sterile distilled water, adding together the values corresponding to positive without additives. reactions within each group, a 7-digit profile number is • Using a pipette or PSIpette, remove a single well- obtained for the 20 tests of the API 20 E strip. The isolated colony from an isolation plate. It is oxidase reaction constitutes the 21st test and has a recommended to use young cultures (18-24 hours old). value of 4 if it is positive. • Carefully emulsify to achieve a homogeneous bacterial • Identification : suspension. This is performed using the database (V4.1) This suspension must be used immediately after * with the Analytical Profile Index : preparation. - Look up the numerical profile in the list of profiles. NOTE : most Vibrio species are halophilous. If a Vibrio is * with the apiweb TM identification software : suspected, suspend the bacteria in API NaCl 0.85 % - Enter the 7-digit numerical profile manually via the Medium. keyboard. bioMérieux SA English - 2 api® 20 E TM 07584J - en - 2010/05 In some cases, the 7-digit profile is not discriminatory - Motility (MOB) : Inoculate an ampule of API M Medium enough and the following supplementary tests need to be (see package insert).
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