Ex 4 gram stain
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Ex. 4: Gram Stain. Chromophores Basic vs. acidic dyes. Primary stain (stains all cells on slide) Decolorizing step (removes stain from certain types of cells) Counterstain (stains the decolorized cells). Differential Stain. Strongly advised:.

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Ex. 4: Gram Stain

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Ex 4 gram stain

Ex. 4: Gram Stain

  • Chromophores

  • Basic vs. acidic dyes


Differential stain

Primary stain (stains all cells on slide)

Decolorizing step (removes stain from certain types of cells)

Counterstain (stains the decolorized cells)

Differential Stain


Strongly advised

Strongly advised:

When doing any of the microbiology labs, make sure to carefully follow the procedure outlined in the Materials and Methods section . . . And any advice given by your instructor!

From now on: read the labs before coming to class!

Have fun!


Little finger technique

“Little Finger Technique”


How to label a slide

How to label a slide

Circles on back

Other info on front


Smear preparation

Smear Preparation

From Liquid Medium

From Solid

Medium

Transfer one loopful of H2O into circle area of slide.

Using loop, suspend very small quantity of colony of interest in that drop of water and spread the suspension evenly. It should only appear slightly milky. Do NOT use too many bacteria

Air-dry and heat-fix

  • Suspend bacterial sediment in culture tube by gently tapping on it.

  • Transfer one loopful of the bacterial suspension to slide and spread out (You may exceed the limits of the circle)

  • Air-dry and heat-fix


Gram stain is

Gram Stain is ….

…the most important bacterial stain!

Therefore:

Memorize steps as soon as possible


Ex 4 gram stain

Gram Staining Reagents

Decolorizing agent

Primary stain

Counterstain

Mordant


Gram stain mechanism slide from lecture

Gram Stain Mechanism(slide from lecture)

  • Crystal violet-iodine crystals form in cell.

  • Gram-positive

    • Alcohol dehydrates peptidoglycan

    • CV-I crystals do not leave

  • Gram-negative

    • Alcohol dissolves outer membrane and leaves holes in peptidoglycan.

    • CV-I washes out


Gram stain animations

Gram Stain Animations

ASM Microbe Library

YouTube presentation

MacGraw Hill Animation


Artifacts

Artifacts

Precipitated crystal violet stain may resemble cocci. Clues: dense clumping, small size of the gram-positive dots. Paucity of organisms elsewhere in the field.


Examples of gram stains

Examples of Gram Stains


Gram stain of urine

Gram Stain of Urine


Gram stain of urine1

Gram Stain of Urine


Ex 4 gram stain

The End


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