Experiment 1: Microscopy, Cell Size Measurement & Counting Cells

Question 1

List the specimens used in the experiment

Answer 1

Bacteria: Diplococcus pneumoniae
Yeast: Activated baker’s yeast cell suspension
Plant: Hydrilla leaf cells
Animal: Human cheek cells
Protists/Single-celled eukaryote: Hay infusion

Question 2

What was the dye used in preparing the wet mounts for yeast cells and human cheek cells?

Answer 2

Methylene blue

Question 3

Enumerate the steps in focusing the microscope

Answer 3

1. Clean the mirrors and lenses with lens paper (kim wipes)
2. Bring the objective lens as far away from the stage as possible
3. Mount the specimen
4. Focus the object by first bringing the stage as close as possible to the objective lens using the coarse adjustment knob (look from the side) then look into the eyepiece and bring the stage down until the object is focused. Use the fine adjustment knob to make the focus more clear.
5. Rotate the objective lens to achieve a greater magnification. DO NOT use coarse adjustment knob to focus object
6. Use the oil immersion objective to focus on bacteria specimens.

Question 4

A small disc of glass on which a scale of 100 divisions is engraved

Answer 4

Ocular micrometer

Question 5

Ocular units

Answer 5

arbitrary: represent different lengths depending on the distance between the ocular and objective lenses and the magnification of the lens

Question 6

The standard used in calibration of the ocular micrometer

Answer 6

Stage micrometer

Question 7

What is one space in the stage micrometer equivalent to?

Answer 7

10micrometers = 0.01mm

Question 8

Linear magnification

Answer 8

achieved using the ocular (enlarges object) and objective (magnifies further the magnified image of the object)

product of the magnification of the ocular lens and the objective lens

Question 9

Magnification of illustration

Answer 9

size of the drawing divided by the actual size of the object

Question 10

What were the conditions for counting yeast cells using a hemocytometer

Answer 10

1.5mL tube
200 microL of yeast cell suspension
200 microL of methylene blue stain

incubated for 1 minute

counted within 10 minutes of placing the stain

Question 11

What is the total volume in each corner of the square of the hemocytometer?

Answer 11

0.1 mm^3

Question 12

Mean no. of cells/0.1mm^3 =

Answer 12

total no. cells in all 4 corners/4

Question 13

Total no. of cells/mL =

Answer 13

(mean no. of cells/0.1mm^3) x 10^4 x dilution factor

Question 14

Total no. of cells =

Answer 14

(Total no. of cells/mL) x Volume of solution (mL)

Question 15

% viable cells =

Answer 15

(Total no. viable cells)/(Total no. of cells)

Question 16

How can viable and non-viable cells be distinguished from one another?

Answer 16

Viable: unstained

Non-viable: stained

Question 17

Calibration constant (c) =

Answer 17

(No. of stage micrometer units x 10 micrometers/space)/No. of ocular micrometer units

Question 18

What are the limits of microscopy?

Answer 18

Varying types of microscopes
Specimen preparation
Microscope performance

Question 19

Why is it necessary to calibrate the ocular micrometer?

Answer 19

The ocular units of an ocular micrometer are arbitrary and dependent on the distance between the ocular and objective lens and the magnification of the lenses

Different measurements may be produced when switching between objective lenses thus the ocular micrometer must be calibrated to each objective lens of a specific microscope to be able to produce accurate measurements