Session 1 - Basics Flashcards
Anatomy of a microscope
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
Diopter Adjustment
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.
Headpiece
Connects the eyepieces to the objective lenses, also often here is the insertion to attach the viewing tube for the camera.
Body
Houses lamp and brightness control, “backbone” that connects the the head- and nosepiece all the way to the bottom of the microscope.
Condenser Adjustment
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).
Coarse Focus Knob
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)
Fine Focus Knob
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
Brightness slider
Adjusts brightness depending on the lenses you use and amount of illumination needed.
Illumination
LED lamp that acts as the source of light.
Stage Control
Two knobs to move stage in vertical and horizontal direction.
Condenser
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.
Iris Diaphragm
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.
Stage
This is where you position the sample slide with cover glass and hold it into place with the metal clip.
Objective lenses
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.
Nosepiece
Rotating part that holds the objective lenses, rotate until it clicks to switch to a different magnification lense.
Eyepiece
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.
What does it mean when we say, these microscopes are ‘compound light microscopes’?
They have more than one lens: ocular (eye pieces) and objective lenses as well as its own light source.
What does it mean when we say, the lenses are parfocal?
The lenses are all aligned and relatively little focusing needed as long as the 4X objective is focused correctly.
How should you store your microscope when you are done using it for the day?
Always place a dust cover on top. Even if you’re just on coffee break.
What cleaning procedure should you use to clean your slides and coverslips?
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
What is the function of the two jars ‘clean water’ and ‘dirty water’?
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.
How do you prevent cross contamination between samples?
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.
What equipment do you need to calibrate your pipette?
Volume method: dry test tube, pipette, clean water
How should you hold your pipette when counting drops that you are adding to the test tube?
Upright and without touching the side of the test tube.
Why is it important to hold the pipette upright and without touching the side?
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
How would you describe where the water level in the test tube should be to read the volume?
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.
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?
You need to divide the number by 2 in order to get the average drop in 1 ml
Why is the correct calibration of the pipette necessary?
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.
Why must you always take a sample from your test tube using a calibrated pipette?
In order to get the most accurate numbers in the sMApp.
What equipment is required to collect a field sample?
Apple corer, sealable plastic bag, permanent marker
How many core-samples do you need to take for a field sample?
A minimum of 3 when collecting around 1 plant or one core sampling area
Where in the soil do you collect your soil sample from?
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.
Where, relative to a plant, do you collect your soil sample from?
You will collect your core halfway between the stem and the dripline.
Do you store your core samples together, in a single bag, or in separate bags? Explain.
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.
How do you mark your collected sample?
It is important to put your name, sample name and date when sample was collected
FoV Diameter?
Coverslip Area?
How many FoV in total?
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
Care procedure for microscope?
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
Care procedure for slide/slip?
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.)
Care procedure for tubes/pipettes?
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
How to prepare a collected sample?
- break up the sample with your fingers so there are no clumps left and until material is mixed and distributed evenly in the bag
- use spatola and take material from different parts of the bag and fill test tube up until 1 ml mark
- tap tube 10x to reduce air pockets
- fill up the tube until the 5ml mark with clean unchlorinated water
- now you have a 1:5 dilution
alsway be consistenst in the preparation of every single sample to be representative
