GC-FAME (Gas Chromatograph Fatty Acid Methyl Ester)
Also known as Cellular Fatty Acid Analysis, allows for bacterial or yeast identification based on the specific fatty acid composition of the cell wall. Fatty acids are extracted from cultured samples and are separated by gas chromatography. A computer generated, unique profile pattern of the extracted fatty acids (9 to 20 carbon chains long) is compared through pattern recognition programs, to our microbial databases of well over 2800 species and subspecies. Our reports include the fatty acid profiles, a listing of the best database matches, along with an assigned statistical probability value indicating the confidence level of the match.
Biolog® MicroLog
This automated metabolic characterization system uses a 96-well microtiter plate arrayed with different carbon sources and the microorganism of interest. Each microorganism has unique capacities to oxidize the various carbon sources. Tetrazolium dye in each of the wells turn darker shades of purple as the carbon sources are oxidized by the microorganism. The test yields a characteristic pattern of positive (purple) and negative wells which provide a metabolic signature of the inoculated organism. The system`s computer the pattern signature with the database of interest to determine an identification. Databases include aerobes, anaerobes, gram positive & negative bacteria, yeast, actinomycetes, and lactic acid bacteria.
RiboPrinter™ (Qualicon™, a DuPont subsidiary)
The RiboPrinter™ system characterizes bacteria by creating a fingerprint pattern of an isolate`s genetic material. Pure colonies are isolated from culture, inactivated by heat and loaded into the sample carrier. DNA is extracted from the cell lysate and is digested to completion using an EcoR1 restriction enzyme. The DNA fragments are separated according to molecular size by gel electrophoresis and are the transferred to moving Nylon membranes. After denaturation, each membrane is hybridized with a chemically labeled rRNA operon probe from E. coli. The membrane is washed and treated with a blocking buffer and an antisulfonated, DNA antibody/alkaline phosphatase conjugate. Unbound conjugate is removed and a chemiluminescent substrate applied. This step makes each electrophoresis band containing genetic information from the rRNA genes visible to a custom CCD camera in the RiboPrinter™ system. The camera detects the light intensity of the targeted bands and converts the patterns from luminescent DNA fragments to digital information. Using algorithms, a pattern is extracted from the image data and is compared to other known patterns stored in the database to characterize and identify each sample.
MicroSeq™ 16S rRNA Gene Sequencing (PE Applied Biosystems)
Genomic DNA is extracted directly from dead or alive bacterial colonies grown under any conditions. The 16S rRNA gene is amplified using universal primers and thermalcyclers. The amplified 16S rRNA gene product is sequenced using dye terminator cycle sequencing chemistry. The sequence reactions are analyzed using automated DNA sequencers and software. Unknown bacterial samples are identified using the MicroSeq™ microbial identification software and 16S rRNA gene sequence database, containing over 1,500 entries. Dependant upon the level of confidence, speed and cost desired, either the entire 1540 base pair 16S rRNA gene, or a smaller portion of the gene can be (500 bp) sequenced. A printout of the actual sequence data and the identification is included in each report.
