Onar® EUB:
Eubacteria Detection Kit for conventional PCR

Cat.-No. 12-1025              25 Tests
Cat.-No. 12-1050              50 Tests
Cat.-No. 12-1100              100 Tests
Cat.-No. 12-1250              250 Tests

 

Description

Onar®EUB is the first assay employing PCR technology for highly sensitive detection of eubacteria in various in situ biologicals including cell cultures and virus stocks.

Sensitive Detection of 50 copies per sample volume, corresponding to ~200 fg eubacterial DNA (e.g. Bacillus subtilis). Specific Detection of more than 45 eubacteria genera: Actinomyces, Bacillus, Entero coccus, Escherichia coli, Fusobacterium, Klebsiella, Lactobacillus, Micrococcus, Mycobacterium, Peptostreptococcus, Pseudomonas, Porphyromonas, Prevo tella, Salmonella, Serratia, Staphylococ cus, Streptococcus, etc. Reliable and rapid The kit contains positive control DNA, and an internal control to eliminate the possibility of false-negative results. Hands-on time less than 40 min. Convenient The reagents are pre-aliquoted for 25 re actions for convenient periodic or routine testing. Simple Easy analysis by comparison with the positive control. Yes or no result.

Points to Consider
Bacteria are frequently found as contaminants in cell cultures. Studies have identified an overall 6 % incidence of bacterial contamination of cell cultures examined. Thus, many cell cultures lack visual signs of bacterial contamination, generally indicated by discoloration of the fluid. Moreover, it has been demonstrated that standard antibiotics not only are ineffective against resistant bacterial infection but also have a strong impact on cell growth & differentiation.

Test Principle
Onar®EUB utilizes the polymerase chain reaction (PCR). The primer set is specific to a highly consered region within the 16S ribosomal RNA gene in the eubacteria genome. This allows for detection of all eubacteria species tested thus far and usually encountered as airborne contaminants in cell cultures. Eukaryo tic DNA is not amplified by Onar®EUB. Onar®EUB also provides internal control DNA to assure successfully executed reactions.

Onar®EUB should be performed only with EUB-Polymerase.
In principle the kit can be performed with any polymerase. However, polymerases of different manufacturers can be contaminated with bacterial DNA leading to false-positive results. Therefore we strictly recommend the use of only DNA-free EUB Polymerase (Cat.-No. 54-0100).

Kit Components

• primer set and nucleotides at opti-mized concentrations
• positive control for easy result verifi-cation
• internal control to ensure accurate testing
• instruction manual and storage box

Onar®EUB kit components are lyophilized and at +2 to +8 °C maintainable for at least 12 months.
Primer and nucleotides are aliquoted (á 25 tests) to guarantee consistent quality.
After re-hydratisation the reagents must be stored at -18 °C.
Note: Please use an appropriate DNA extraction kit (e.g. Minerva Biolabs Cat.-No. 56-1100) for the DNA purification step prior to PCR analysis.

Test Evaluation
The amplified PCR products are visualized by standard gel electrophoresis. A sample containing eubacterial DNA will produce a band around 468 bp band, with the exception of Micrococcus luteus, which produces a band at 448 bp. This band is absent in reactions performed with non contaminated samples and in the negative control (no-template control). The internal control DNA should be present in every reaction (210 bp band), except in samples of a strong contamination. In the event of a PCR inhibition, no bands are present.

Onar®EUB is intended for research only.

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