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Microbial Limit Testing Technology Overview. Advisory Committee for Pharmaceutical Science July 19, 2001. Compendial Methods USP. Plate Counts- Colony Forming Units Growth of microorganisms on solid medium Count visible colonies MPN- Most Probable Number

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Microbial limit testing technology overview

Microbial Limit TestingTechnology Overview

Advisory Committee

for

Pharmaceutical Science

July 19, 2001


Compendial methods usp
Compendial MethodsUSP

  • Plate Counts- Colony Forming Units

    • Growth of microorganisms on solid medium

    • Count visible colonies

  • MPN- Most Probable Number

    • Multiple serial dilutions of sample

    • Based on statistical distribution of organisms in a sample

ACPS


Compendial methods

ADVANTAGES

Simple

Counts only viable organisms

DISADVANTAGES

Incubation time (48-72 hours)

Not all organisms will grow on a single medium

Compendial Methods

ACPS


Microbial viability
Microbial Viability

  • Only interested in viable microorganisms

  • “New” methods must only count viable organisms

  • Viability indicators needed

ACPS


Esterase detection chemscan chemunex
Esterase DetectionChemScan®, Chemunex

  • Enzyme ubiquitous in microorganisms

  • Chem-Chrome, a non-fluorescent fluorescein derivative cleaved by esterase to fluorescein

ACPS


Chemscan procedure
ChemScan® Procedure

  • Filter Sample

  • Expose membrane to reagent

  • Membrane analyzed by laser-scanning

ACPS


Atp bioluminescence microstar tm microcount tm millipore
ATP BioluminescenceMicroStar(TM) & MicroCount(TM), Millipore

  • ATP- Primary energy source for all organisms

  • Luciferin/luciferase (reagent) reacts with ATP to produce light

ACPS


Microstar tm procedure
MicroStar(TM) Procedure

  • Filter Sample

  • Place membrane on solid medium for brief incubation

  • Disrupt cells to release ATP

  • Add bioluminescence reagent to membrane

  • Detect light with CCD and computer analysis

ACPS


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