Rapid Microbiological Methods and the PAT Initiative - Numerous new RMM systems are available to replace traditional testing methods - BioPharm International

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Rapid Microbiological Methods and the PAT Initiative
Numerous new RMM systems are available to replace traditional testing methods


BioPharm International
Volume 18, Issue 12

Microcalorimetry

Type of Technology: Viability-based, but requires catabolic activity.

Premise of Technology: The process of microbial catabolism results in heat that can be measured by micro-calorimetry. The sample is placed in a sealed ampoule with media inside a calorimeter. The instrumentation can be used to establish growth curves. When high levels of contamination are present, one may need to use flow calorimetry.7

Commercial Systems Available: A variety of systems is available.

Other: This process cannot be used to determine if a single contaminant is present, nor can it be used on samples with mixed contaminants.7

Nucleic Acid Probes

Type of Technology: Nucleic-acid-based.

Premise of Technology: Data available from nucleic acid sequencing are used to select a desired nucleic acid. The desired nucleic acids are extracted, immobilized to a solid phase, and hybridized to a labeled probe. Alternatively, the extracted nucleic acids can be labeled and hybridized to an immobilized probe.17

Commercial Systems Available: The systems available depend on the type of outcome desired. They include Gene-Trak Systems (Gene-Trak) and Gene-Probe Systems (Gene-Probe).

Other: This technology is frequently used for characterization or identification of microorganisms, and for pathogen detection.

Polymerase Chain Reaction

Type of Technology: Nucleic-acid-based.

Premise of Technology: PCR makes copies of nucleic acid fragments. Nucleic acid fragments are amplified using polymerization techniques. The fragment of interest is heat-denatured; the reaction vessel is then cooled, and the polymerase begins to create the complementary strand. Another denaturation step, followed by another polymerization step, doubles the amount of DNA. Several iterations of this process produce a massive amount of DNA. A variety of PCR methods may be used: reverse transcripterase PCR, nucleic acid sequence-based amplification, or transcription-mediated amplification.

Commercial Systems Available: BAX Microbial Identification System (Qualicon) and Probelia System (BioControl Systems).

Other: This technology is widely used in other sciences, such as anthropology and forensics.

Raman Spectroscopy

Type of Technology: Cell-component-based.

Premise of Technology: A Raman spectrophotometer can generate a spectrum unique to the microorganism. The spectra are stable across taxonomic groups. The patterns are compared to a database of spectra of known microorganisms.

Commercial Systems Available: Raman spectrophotometer.

Other: Studies performed in clinical settings indicated that identifications could be made from cultures incubated for approximately five hours. The test is non-destructive.

Ribotyping - Molecular Typing

Type of Technology: Nucleic-acid-based.

Premise of Technology: This technology uses restriction fragment length polymorphisms (RFLPs) of nucleic acids from bacterial genomes. The size-separated RFLPs are hybridized to a ribosomal ribonucleic acid (RNA) probe. A chemiluminescent substrate is applied. A camera is used to convert the luminescing RFLPs to digital information. The digital information is captured and the data extracted. A pattern is generated and compared to a database of known patterns for identification. The ribotype is a stable epidemiological marker and provides definitive taxonomic information.

Commercial Systems Available: MicroSeq 16S rDNA Bacterial Identification System (Applied Biosystems) and Riboprinter (DuPont Qualicon).

Other: Molecular typing is considered the "gold standard" for identifying microorganisms.


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