1 / 13

Advanced Bacterial Counting Methods for Laboratory Analysis

Explore direct and indirect bacteria counting techniques including calibrated stained smears, membrane filtration, and electronic counter methods. Learn about dilution series, optical density measurement, and more for accurate bacterial quantification in lab settings.

vernonmcdonald
Download Presentation

Advanced Bacterial Counting Methods for Laboratory Analysis

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. Counting Bacteria • Filename: CelCount.ppt • Hugh B. Fackrell

  2. Presentation Outline • Direct Counts • Microscopic • Electronic counter • Dilution series • Membrane filtration • Indirect Counts • Optical density • Weight of culture • Chemical

  3. Direct Counts: Microscopic • Calibrated stained smear • Counting chamber • Epifluorescence Need high numbers of bacteria in the sample to be useful

  4. Calibrated Stained Smear

  5. Petroff- Hauser Counting Chamber

  6. Epifluorescence Microscopy • Nucleic acid specific dye • 3-6 tetramethyldiamino acridine • Binds between base pairs of DNA or RNA • Intercalation • Fluoresces when bound • DS-DNA -> Green • SS-DNA -> Red Orange • Degraded DNA -> Red Orange

  7. Epifluorescence • Approximate viable cells /total cells • Viable cells green • Dead cells orange • Count low numbers • cells fluorscent against a black background

  8. 1ml 1ml 1ml 1ml 1ml 1/1,000 1/10 1/100,000 1/100 1/10,000 Dilution Series: dilution 9ml broth

  9. 1ml 1ml 1ml 1ml 1ml 1/1,000 1/10,000 1/10 1/100 1/100,000 Dilution Series: plating

  10. 1/1,000 1/100 1/10 1/10,000 1/100,000 Dilution Series: Colony counts

  11. Membrane filtration • Membrane filter • fixed pores 0.45 m 0.22 m • bacteria trapped on filter • Filter known amount of fluid • ALL bacteria trapped on membrane • Place filter in Petri dish with appropriate medium • Incubate • Count number of colonies

  12. Membrane filtration • Number of Viable cells • Each colony from a single bacterium • Sensitive • eg filter entire bottle of pop • <1 bacterium / bottle • Growth media • selective or differential • Combine with Epifluorescence • Black polycarbonate filters

  13. Electronic Measurement • Turbidity • Broths with many bacteria become turbid • turbidity increases light scattering • increased absorbance in spectrophotometer • Cell counters • fluid pumped through a micro pipette • pumped past an electronic sensor • records number of cells

More Related