gram stain and acid-fast stain

What type of staining is gram staining

differential staining

The primary stain used in gram staining is

crystal violet

The counter stain in gram staining is

safranin

what two types of cells will be identified by their response in gram staining?

gram-positive and gram-negative

What type of response does gram positive have?

appear purple (it retains the crystal violet-iodine complex even after the de-colorization with alcohol or acetone

What type of response does gram negative have?

appear pinkish red (alcohol or acetone removes the crystal-violet iodine)

What had the gram-negative result?

Pseudomonas aeruginosa

What had the gram-positive result

Staphlococcus aureus

Why is the counter-stain used?

Gram-negative does not retain the primary stain crystal-violet so a counter stain has to be used after the De-colorization step or the would not be visualized by the microscope.

What mordant is used

grams iodine

Why is grams iodine used as a mordant?

It complexes with the crystal violet and forms and insoluble complex in gram positive stains.

Steps for Gram Staining

1. primary stain-crystal violet

2. wash

3. mordant-grams iodine

4. de-colorization-alcohol

5. wash

6. counter stain-grams safranin

7. wash

8. blot dry

Is the dye-mordant complex removed from the gram-positive bacteria?

No

Is the dye-mordant complex removed from the gram-negative bacteria?

yes, it is leached away

Why doesn't the counter stain safranin change the violet color of the gram positive bacteria?

because the crystal-violet is a much more intense stain than safranin.

What affect does penicillin have on a gram positive bacteria?

penicillin will inhibit peptidoglycan synthesis.

What affect does penicillin have on gram negative bacteria?

None, except for gonorrhea

Should the slides be heat fixed before staining?

Yes

Gram positive bacteria's wall is

made up of a thick layer of peptidoglycan

gram negatives wall consist

of an outer membrane that covers a much thinner layer of peptidoglycan

What is the primary stain in acid-fast staining?

red/basic fuchsin

What is the counter stain in acid-fast staining?

blue/methylene blue

What is mycolic acid?

a waxy cell wall lipid. Composed of fatty acids and fatty alcohols that have hydrocarbon chains up to 80 carbons in length. The waxy microbes stick together, helps resist phagocytosis.

Are cell walls with high lipid content easily stained?

No, the mycolic acid in their cell walls prevents the penetration of most stains.

An example of a (+) acid fast bacilli

tuberculosis (Mycobacterium tuberculosis) and leprosy (Mycobacterium leprae)

First step in acid-fast staining

The primary stain (red/basic fuschin-contains phenol ) is placed on the heat-fixed smear and sits for 5-6 minutes (don't allow it to dry)

What color are results for (+) acid-fast bacteria

pink to red

What color are results for (-) acid fast bacteria

appear blue

Example of (-) acid stain bacteria

staphlococcus

Steps for acid-fast staining

1. primary stain-red basic fusion (5-6 min)

2. wash and shake

3. 2 drops ACID-ALCOHOL

4. rinse with water

5. counterstain

6. wash

7. blot dry

What type of is acid-fast staining

Its differential staining, with mixed smear. Both bacteria are mixed on the same slide.

What type of results do you get with differential staining - qualitative or quantitative

Quantitative

When you see on a lab test that a patient has a positive acid fast test what should you immediately suspect?

Tuberculosis