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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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