Prep of Pure Culture and Gram Stain

Question 1

How is a pure culture isolated from a mixed culture? Why is it important?

Answer 1

1. flame the inoculating loop 2. take a sample of each isolated colony from each bacteria by touching the loop on the top of the bacteria. 3. transfer each sample to a separate sterile soy agar plate using the three-phase pattern of streaking important so each bacteria can grow separately without cross-contamination

Question 2

How is a slide prepared?

Answer 2

1. clean the slide with soap and water, then rinse with 95%alcohol 2. label slide on the frosts part. place circle in middle of slide 3. small drop of distilled water should be placed over the circled area. 4. After aseptically removing material from a culture it is them mixed with the drop of water on the slide. 5. dry the slide by holding it above the flame 6. once dried, quickly pass slide it 2-3 times through a flame so the bacteria adhere. 7. let cool

Question 3

Gram stain procedure and color changes

Answer 3

1. crystal violet stain; 1 min. 2. rinse with water. 3. iodine (rusty) stain; 1 min 4. rinse again. 5. alcohol stain; 10 sec. wash is colorless 6. rinse again. 7. safranin (red) stain; 1 min 8. rinse 9. blot dry in a blotter booklet

Question 4

DIfferent shapes of bacteria?

Answer 4

Question 5

Based on color determine whether a stain in gram + or gram -

Answer 5

• = stain pink (e.coli) rod shape; independent

+ = stain dark purple (m.luteus) sphere shape; cluster

Question 6

e. coli stain

Answer 6

Question 7

m.luteus stain

Answer 7

gram +

Question 8

diference between gram + and gram - cell wall?

Answer 8

A gram positive bacteria: thick layer of peptidoglycan that the stain can penetrate.

A gram negative bacteria: has an outer membrane covering a thin layer of peptidoglycan on the outside. The outer membrane prevents the initial stain from penetrating.

Question 9

Medical importance of gram stain?

Answer 9

determine the species

characteristics of disease

treatment of a bacteria