Session 1 - Basics

Question 1

Anatomy of a microscope

Answer 1

A: Diopter Adjustment

B: Headpiece

C: Body

D: Condenser Adjustment

E: Coarse Focus Knob

F: Fine Focus Knob

G: Brightness Control

H: Lightness Control

I: Stage Control

J: Condenser

K: Iris Diaphragm

L: Stage

M: Objective Lenses

N: Nosepiece

O: Eyepieces

Question 2

Diopter Adjustment

Answer 2

Turn them to adjust the focal plane for each individual eye, as they are often not exactly the same. Once you got them to the same level, your brain doesn’t have to constantly try to compensate for the difference and you won’t get headaches.

Question 3

Headpiece

Answer 3

Connects the eyepieces to the objective lenses, also often here is the insertion to attach the viewing tube for the camera.

Question 4

Body

Answer 4

Houses lamp and brightness control, “backbone” that connects the the head- and nosepiece all the way to the bottom of the microscope.

Question 5

Condenser Adjustment

Answer 5

Adjusts the height of the condenser to focus the light correctly onto the slide with the sample by moving the condenser up and down (if not done correctly you might see grainy artifacts and the organisms will be not perfectly focused).

Question 6

Coarse Focus Knob

Answer 6

Used to coarsely adjust distance of the stage to the 4x objective lense to get them into focus and to align the other higher magnification lenses (parfocal)

Question 7

Fine Focus Knob

Answer 7

Used to finely adjust distance of the stage to the lens with higher magnification lenses (10x/40x) to be able to focus through the full depth of the sample

Question 8

Brightness slider

Answer 8

Adjusts brightness depending on the lenses you use and amount of illumination needed.

Question 9

Illumination

Answer 9

LED lamp that acts as the source of light.

Question 10

Stage Control

Answer 10

Two knobs to move stage in vertical and horizontal direction.

Question 11

Condenser

Answer 11

Gathers and focuses the light onto the sample, needs to be in the correct position to focus the light onto the sample correctly to achieve maximum clarity and sharpness.

Question 12

Iris Diaphragm

Answer 12

Ring aperture to adjust the amount of light coming through and therefore increasing the contrast due to shadowing. This is what makes most of the organisms and structures distinguishable to the human eye in the first place.

Question 13

Stage

Answer 13

This is where you position the sample slide with cover glass and hold it into place with the metal clip.

Question 14

Objective lenses

Answer 14

One of two magnifications taking place through these lenses: 4x / 10x / 40x

The higher the magnification of the lense the less light is reaching the eye, so you need to adjust the brightness when changing magnifications.

Question 15

Nosepiece

Answer 15

Rotating part that holds the objective lenses, rotate until it clicks to switch to a different magnification lense.

Question 16

Eyepiece

Answer 16

The lenses closest to the eyes. The coding on the side tells your the field of view and the magnification of the eyepieces. We use 10x. This is where you look through to see the sample.

Question 17

What does it mean when we say, these microscopes are ‘compound light microscopes’?

Answer 17

They have more than one lens: ocular (eye pieces) and objective lenses as well as its own light source.

Question 18

What does it mean when we say, the lenses are parfocal?

Answer 18

The lenses are all aligned and relatively little focusing needed as long as the 4X objective is focused correctly.

Question 19

How should you store your microscope when you are done using it for the day?

Answer 19

Always place a dust cover on top. Even if you’re just on coffee break.

Question 20

What cleaning procedure should you use to clean your slides and coverslips?

Answer 20

When taking the slide off the stage, put it on a flat surface. Then, taking a cloth or paper towel, gently wipe off the cover slip. That will also be how you initially clean off the drop. Then you breathe on the slide and wipe with a lens cloth. You may also decide to wash the slide in clean water or soapy water if you see residue. Use the same procedure with coverslips.
Lens cleaning solution acceptable too

Question 21

What is the function of the two jars ‘clean water’ and ‘dirty water’?

Answer 21

The ‘clean water’ jar contains water free of chlorine or chloramine that you can use in your test tubes to prepare a sample. It can also be used to get an extra rinse of your pipette and store pipettes after they have been rinsed thoroughly. The ‘dirty water’ jar is used to give the first rinse to your pipettes after they are in contact with your sample or dilution.

Question 22

How do you prevent cross contamination between samples?

Answer 22

Make sure that you keep your samples separate and clean your lab scoop between each sample. Also, use a clean pipette with each different sample.

Question 23

What equipment do you need to calibrate your pipette?

Answer 23

Volume method: dry test tube, pipette, clean water

Question 24

How should you hold your pipette when counting drops that you are adding to the test tube?

Answer 24

Upright and without touching the side of the test tube.

Question 25

Why is it important to hold the pipette upright and without touching the side?

Answer 25

To make sure that drops go to the bottom and do not adhere to the sides of the test tube, but primarily to ensure drops are of consistent size

Question 26

How would you describe where the water level in the test tube should be to read the volume?

Answer 26

The bottom of the meniscus “bowl” should be on the 1 ml line or the 2 ml line depending on the graduation mark you are measuring to.

Question 27

If you filled the test tube to 2 ml, how do you determine the number of drops per ml (drops/ml) for your calibrated pipette?

Answer 27

You need to divide the number by 2 in order to get the average drop in 1 ml

Question 28

Why is the correct calibration of the pipette necessary?

Answer 28

It is necessary in order for the sMApp to correctly calculate the organism numbers in the whole slide. calculates the amount of liquid that is in the drop of sample dilution you placed on your slide to correctly calculate the organism populations in the sample.

Question 29

Why must you always take a sample from your test tube using a calibrated pipette?

Answer 29

In order to get the most accurate numbers in the sMApp.

Question 30

What equipment is required to collect a field sample?

Answer 30

Apple corer, sealable plastic bag, permanent marker

Question 31

How many core-samples do you need to take for a field sample?

Answer 31

A minimum of 3 when collecting around 1 plant or one core sampling area

Question 32

Where in the soil do you collect your soil sample from?

Answer 32

You take your sample from within the first 3 inches of soil. This is done after pulling back any organic matter that is on top.

Question 33

Where, relative to a plant, do you collect your soil sample from?

Answer 33

You will collect your core halfway between the stem and the dripline.

Question 34

Do you store your core samples together, in a single bag, or in separate bags? Explain.

Answer 34

Once you collect your 3 core samples, you mix them thoroughly in your bag. Of course you do not mix core samples from different areas.

Question 35

How do you mark your collected sample?

Answer 35

It is important to put your name, sample name and date when sample was collected

Question 36

FoV Diameter?

Coverslip Area?

How many FoV in total?

Answer 36

FoV Diameter = Field Number (18mm) / Objective Magnification (40x)

= 18mm/40 = 0,45mm = 450 µm

Cover slip surface = 18mm x 18mm = 324mm²

FoV = 𝜋(0.5 * 450µm)²​ ​

= 0,159mm²

Number of FoV per cover slip = 324mm² / 0,159mm² = 2038

Question 37

Care procedure for microscope?

Answer 37

always put the cover on to protect the microscope from dust, to protect it from getting dust into gears and internal lenses, cover vulnerable surfaces when assembling/dissembling microscope

yearly regular clean from a professional

Question 38

Care procedure for slide/slip?

Answer 38

when done immediately pull the cover slip off, breath onto both and then wipe with a lense cleaning cloth, if still residues wash with water and drop of detergent

hold it into the light and make sure the slide and slip are clean (no fingerprints, residue, etc.)

Question 39

Care procedure for tubes/pipettes?

Answer 39

clean tubes thouroughly after each use with water and if necessary detergent, put upside down into a rack to dry them

after each use clean pipette in the “dirty water” cup first and then after take up water from the “clean water” and disperse it into the “dirty water” cup

leave the pippete filled with clean water in the clean water cup

Question 40

How to prepare a collected sample?

Answer 40

1. break up the sample with your fingers so there are no clumps left and until material is mixed and distributed evenly in the bag
2. use spatola and take material from different parts of the bag and fill test tube up until 1 ml mark
3. tap tube 10x to reduce air pockets
4. fill up the tube until the 5ml mark with clean unchlorinated water
5. now you have a 1:5 dilution

alsway be consistenst in the preparation of every single sample to be representative