1.2.4 Determination of cell Numbers

Question 1

3 ways in determining cell numbers?

Answer 1

• Total cell count
• Viable count / Plate count
• Measuring the density or turbidity of cells

Question 2

Can be used to estimate the number of bacteria present in a specimen. It does not distinguish between live and dead cells

Answer 2

Total cell count (Direct counting)

Question 3

What microscope is used in Total cell count?

Answer 3

Phase-contrast micorscope

Question 4

What microscope is used in Total cell count?

Answer 4

Phase-contrast microscope

Question 5

Used to determine the number of colony-forming units per milliliter (CFU/mL of bacteria). This method provides a count of viable cells only.

Answer 5

Viable Count or Plate count

Question 6

How Plate count is done?

Answer 6

Serial dilution then plate each of diluted

Question 7

Common amount of colonies required in Plate count?

Answer 7

30-300 colonies

Question 8

The density of a bacterial broth culture in log phase can be correlated to CFU/mL of the culture. This method is used to prepare a standard inoculum for antimicrobial susceptibility testing

Answer 8

Density Measurement

Question 9

What equipment is use to determine the turbidity of a sample?

Answer 9

Spectrophotometer

Question 10

When is density measurement is standard?

Answer 10

Antimicrobial susceptibility testing

Question 11

Conventional method of Density measurement

Answer 11

Compare it to 0.5 Mcfarland solution

Question 12

How to measure Exponential phase or lag phase that is to be used in Density measurement?

Answer 12

Optical density 600 (0.02 - 0.05)

Question 13

2 ways in performing plate count

Answer 13

Spread-plate method
Pour-plate method

Question 14

Steps in Spread-plate method?

Answer 14

1. Sample is pipetted onto surface of agar plate (0.1 mL or less)
2. Sample is spread evenly over surface of agar using sterile glass spreader

Question 15

Steps in Pour-plate method

Answer 15

1. Sample is pipetted in to sterile plate (0.1 - 1 mL)
2. Sterile medium (Molten Agar 45 -50 C) is added and mixed well (Swirl) with inoculum
3. Solidify 15-20 mins
4. Incubate (35 +- 2) - 18 - 24 hrs

Question 16

Most valid count of colonies?

Answer 16

30 -300 colonies

Question 17

If there are too much colonies, what should be the interpretation?

Answer 17

TNTC (To numerous to count)

Question 18

Most common dilution to dilute the bacteria in Plate count?

Answer 18

10-Fold dilution

Question 19

How 10-fold dilution is done?

Answer 19

0.5 of sample with 4.5 mL of diluent or 1.0 ml of sample with 9.0 mL of diluent

Question 20

How 100-fold dilution is done?

Answer 20

0.05 mL can be mixed with 4.95 mL of diluent or 0.1 mL with 9.9 mL of diluent

Alternatively, a 10^-2 dilution can be achieved by making two successive 10 fold dilution`

Question 21

What is the diluent use for 10 fold dilution?

Answer 21

Phosphate buffer solution