How to study cells - Microscopes, magnification and calibrating the eyepiece graticule
Summary
TLDRIn this educational video, Mr. Ik explores the world of microscopes, focusing on their types, magnification, and calibration. He explains the difference between optical and electron microscopes, highlighting the latter's superior resolution and magnification for detailed cellular observation. The tutorial covers the magnification formula, unit conversion, and the calibration process of the eyepiece graticule using a stage micrometer. This comprehensive guide is aimed at students seeking to understand microscopes for their A-Level biology studies.
Takeaways
- 🔬 The video covers the topic of microscopes, focusing on magnification calculation, unit conversion, and calibration of the eyepiece graticule.
- 🌟 Mr. Ik discusses three key types of microscopes: optical or light microscopes, and two types of electron microscopes - transmission and scanning electron microscopes.
- 🔍 Magnification in microscopes refers to the size increase of the image compared to the actual object size, while resolution is the minimum distance between two objects that can still be viewed as separate.
- 🌈 Optical microscopes have lower resolution and magnification but allow for color images and the use of living samples.
- 🖤 Electron microscopes offer higher resolution and magnification but produce black and white images and require samples to be in a vacuum, thus not suitable for living samples.
- 📏 The formula for calculating magnification is image size equals actual size times magnification, which can be rearranged to find either value.
- 📐 Unit conversion is necessary when measuring image sizes in millimeters and actual sizes in micrometers, with millimeters converted to micrometers by multiplying by 1000.
- 📏 The eyepiece graticule is a tool for measuring the size of objects under the microscope, but it requires calibration using a stage micrometer.
- 🔧 Calibration of the eyepiece graticule involves aligning it with a stage micrometer, counting divisions, and calculating the value per division at different magnifications.
- 🔬 The video concludes with a summary of key points, including the differences between microscope types, the use of the magnification formula, and the importance of calibrating the eyepiece graticule.
Q & A
What are the three key types of microscopes mentioned in the script?
-The three key types of microscopes mentioned are the optical or light microscope, and the electron microscopes, which include the transmission electron microscope and the scanning electron microscope.
What is the definition of magnification in the context of microscopes?
-Magnification refers to how many times larger the image that you look at is compared to the actual size of the object.
What is the definition of resolution or resolving power in microscopy?
-Resolution or resolving power is the minimum distance between two objects which they can still be viewed as separate, meaning they do not blur into looking like one single point.
Why do electron microscopes have a higher resolving power and magnification than optical microscopes?
-Electron microscopes have a higher resolving power and magnification because electrons have a much shorter wavelength than light, which allows for higher detail and magnification in the images they produce.
What is the difference between a transmission electron microscope and a scanning electron microscope?
-A transmission electron microscope passes a beam of electrons through a very thin specimen, creating a 2D image with varying shades of white and black based on electron absorption. A scanning electron microscope, on the other hand, scatters and reflects electrons off the surface of the specimen, creating a 3D image that provides details on texture and depth.
Why can't living samples be viewed with an electron microscope?
-Living samples cannot be viewed with an electron microscope because the electron beam would be absorbed by air, and the samples need to be in a vacuum for the electrons to reach and interact with the specimen to create an image.
What is the formula used to calculate magnification from a microscope image?
-The formula used to calculate magnification from a microscope image is Image Size (I) = Actual Size (A) × Magnification (M).
How do you convert millimeters to micrometers when measuring a microscope image?
-To convert millimeters to micrometers, you multiply the number of millimeters by a thousand, since one millimeter is equal to one thousand micrometers.
What is an eyepiece graticule and why is it necessary to calibrate it?
-An eyepiece graticule is a glass disk with a scale etched onto it, located within the eyepiece of a microscope. It is necessary to calibrate it because the value of each division on the graticule changes with different magnifications, and calibration ensures accurate measurements of the specimen.
How do you calibrate an eyepiece graticule using a stage micrometer?
-To calibrate an eyepiece graticule, you place a stage micrometer on the microscope stage and align it with the eyepiece graticule. You then count how many divisions on the eyepiece graticule fit into one division on the stage micrometer, and use the known value of the stage micrometer's divisions to determine the value of each division on the eyepiece graticule at the current magnification.
Outlines
🔬 Introduction to Microscopes and Their Types
This paragraph introduces the topic of microscopes, focusing on their types and applications in cell biology. Mr. Ik begins by addressing the request for help with microscopes, particularly in magnification calculation, unit conversion, and calibration of the eyepiece graticule. The session is part of a broader study of cell methods. The three key types of microscopes are mentioned: the optical or light microscope and the electron microscopes, which include transmission and scanning types. The paragraph also covers the basic definitions of magnification and resolution, which are crucial for understanding the capabilities of microscopes. Optical microscopes are limited by the longer wavelength of light, resulting in lower resolution and magnification but allowing for color images and the use of living samples. In contrast, electron microscopes use electron beams with shorter wavelengths, providing higher resolution and magnification but only in black and white and requiring vacuum conditions, thus precluding the observation of living samples.
📊 Magnification Calculation and Unit Conversion
The second paragraph delves into the mathematical aspects of microscope usage, specifically the calculation of magnification and the conversion of units. The formula for image size in relation to actual size and magnification is introduced, and a refresher is provided on how to rearrange the formula to solve for magnification or actual size. The importance of unit conversion is emphasized, as measurements taken with a microscope are often in millimeters, which need to be converted to micrometers to match the scale of cellular structures. The process of converting millimeters to micrometers and vice versa is explained, and a worked example is provided to illustrate how to calculate the magnification of a micrograph image using a scale bar.
📏 Calibration of the Eyepiece Graticule
This paragraph explains the use and calibration of the eyepiece graticule, a tool for measuring the size of structures observed under a microscope. The eyepiece graticule is a glass disk with a scale etched onto it, used to measure objects by counting the number of graticule divisions they cover. However, because different objective lenses provide different magnifications, the graticule must be calibrated each time a new lens is used. Calibration involves using a stage micrometer, a glass slide with a known scale in micrometers. The process of aligning the graticule with the stage micrometer, counting the divisions, and calculating the value of each division at a given magnification is described. An example is given to demonstrate how to determine the actual size of a cell nucleus by counting the number of graticule divisions it covers and multiplying by the calibrated value per division.
🔍 Summary of Key Points on Microscopes
The final paragraph summarizes the key points covered in the session on microscopes. It highlights the differences between optical and electron microscopes, emphasizing the superior resolving power and magnification of electron microscopes, which allow for detailed observation of organelles. The paragraph reiterates the importance of the magnification formula and its application in determining the actual size of structures from microscope images. It also stresses the necessity of calibrating the eyepiece graticule for accurate size measurements at different magnifications. The session concludes with a reminder of the related videos available for further study and an invitation to subscribe for updates on new content.
Mindmap
Keywords
💡Microscope
💡Magnification
💡Resolution
💡Optical Microscope
💡Electron Microscope
💡Transmission Electron Microscope (TEM)
💡Scanning Electron Microscope (SEM)
💡Eyepiece Graticule
💡Stage Micrometer
💡Unit Conversion
Highlights
Introduction to microscopes and their importance in studying cells.
Explanation of magnification calculation and its significance in microscope usage.
Conversion of units between millimeters and micrometers for accurate measurements.
Calibration of the eyepiece graticule for precise cell measurements.
Types of microscopes: optical or light microscope, and electron microscopes (transmission and scanning).
Definition of magnification and resolution in the context of microscopes.
The role of wavelength in determining the resolution of optical and electron microscopes.
Advantages of optical microscopes: color images and the ability to use living samples.
Limitations of optical microscopes in resolving small organelles and internal structures.
Electron microscopes' requirement for vacuum and the impact on sample preparation.
Detailed explanation of transmission electron microscopes and their imaging process.
Scanning electron microscopes' ability to provide 3D images and texture details.
Mathematical formula for calculating image size and its rearrangement for different measurements.
Practical example of calculating magnification using a scale bar on a micrograph.
Step-by-step guide on calibrating the eyepiece graticule using a stage micrometer.
Summary of key points for understanding microscopes and their applications in biology.
Link to the next video on self-fractionation and ultracentrifugation for further learning.
Transcripts
hello and welcome to learn a level
biology for free with mr. ik today we're
gonna be going through microscopes and
this is one of the topics that I
frequently get requests in my tutoring
to have help with going through the
magnification calculation the conversion
of units and mainly the calibration of
the eyepiece grass cue so we'll be
covering all of that in today's session
so if you want get some paper at the
ready for some of the questions and make
notes as we go so this all falls under
topic 2 within the methods of studying
cells and once we've gone through the
cells it's knowing how scientists
discovered that these are the internal
structures our cells have and this has
been worked out through using
microscopes and self fractionation and
ultra Gatien now this video we're just
going to be focusing on the types of
microscopes magnification and
calibration of the eyepiece graticule
and but for the link to the next video
on self fractionation and ultra
centrifugation that'll be at the end of
this one so you can just click the link
to that one say microscopes this is
something that you did cover at GCSE and
there are three key types of microscopes
you have the optical or light microscope
and the electron microscopes but there's
two types of electron microscope you
have a transmission or a scanning
electron microscope now the first bit of
recap to have a go at is can you still
remember from GCSE the definition of
what magnification means and resolution
or resolving power so pause the video at
this point to see can you come up with a
definition for those to write the
definition for magnification is the
magnification of the microscope is
referring to how many times larger the
image that you look at is compared to
the actual size of the object the
resolution of a microscope is the
minimum distance between two objects
which they can still be viewed as
separates so by that we mean if you are
looking at your sample through the
eyepiece under the microscope the
minimum distance in which you can still
see two different parts of the cell has
been separate rather than then they have
just blurred into look like one single
point so the resolution or the resolving
power in an optical microscope is
determined by the wavelength of light
and in an electron microscope it's the
same idea it's the wavelength of the
beam of electrons so let's have a look
at these microscopes in a bit more
detail at this point I've just split it
into the two types generically so the
optical microscope will be the type of
microscope you're familiar with this is
what schools and colleges will have
access to and in this case you have a
beam of light being released by your
lamp or sometimes it's a mirror to
reflect a plugged in lamp and that light
then gets shown up and is condensed and
that is what creates the image now you
have a much lower resolution on light
microscopes compared to electron
microscopes because the light has a
longer wavelength and that is what
determines the resolution of microscopes
it also has a lower magnification in
comparison but you can get color images
so what you're viewing you will see
those real colors and you can use living
samples in contrast the electron
microscope whether it's scanning or
transmission for both of these this time
the source is going to be missing a beam
of electrons and those will be condensed
using electromagnets and that is what is
going to create the image electrons have
a much shorter wavelength so you get a
higher resolution or resolving power
higher magnification but you can only
get black and white images and the
samples that you've you have to be in a
vacuum so completely vacuum no air and
for that reason you can't view living
sample
so a bit more about the optical
microscope we've talked about in fact it
has a lower resolution because of that
longer wavelength what that means in
terms of its application is you can't
see the inside of organelles in detail
and some of the small organelles you
can't actually see at all so I've got an
example over here where we're looking at
mitosis in root tips and you can see
that there is an outer layer you can't
actually see the difference between the
cell membrane and the cell wall we know
that this liquid in the middle must be
the cytoplasm and stained purple are the
chromosomes so we can see where the
nucleus is but that is the level of
detail you can see with the
magnification and resolving power of an
optical microscope the electron
microscopes so going back to what I was
saying the specimen has to be in a
vacuum and the reason for that is the
electron beam that is released those
electrons would be absorbed by the air
and never even reach the sample or
specimen to create the image so for that
reason has to be prepared as a vacuum
and that's why you can't view living
samples and you only get black and white
images so a bit more detail the
transmission electron microscope so
transmission means to pass through and
that is what is happening in a
transmission electron microscope that
beam of electrons some will be absorbed
by the specimen some will pass through
and that's why you get these different
shades of white and the black and the
darker is the more electrons have been
absorbed and that's how you get this
detailed image below so it's a 2d image
it's only black and white but you can
see this here is our chloroplasts inside
of a plant cell and you can see the
internal structures you can see these
thylakoid membranes you can see the
grana those stacks of the membranes as
well scanning electron microscope you
don't have to have these very very thin
samples like
with the transmission this time it's not
going to be transmitting so passing
through the specimen instead it will
scatter and reflect off the surface and
because of the different depths of the
specimen that will affect the scattering
of the electrons and that then creates
this 3d image so the scanning electron
microscope will give you details on the
image to do with the texture and the 3d
depths of either your cells or your
organelles so moving on then
magnification is one of the math skills
linked to microscopes and it's used with
optical microscope images so the formula
again straight from GCSE image size
equals actual size times magnification
now I've deliberately written it this
way rather then is the magnification
formula because I think this is the
easiest way to remember it I am so I
beam image a is the actual M is the
magnification and then once you can
remember I am you can rearrange the
formula to work out magnification or
actual size now that is a skill from
GCSE maths but if you can't remember
that I'll link up the top here just
click to see one of the GCSE videos I
have on rearranging the formula just to
recap to get your aid of a math skills
up scratch so the other thing that
you'll need to be able to do is one of
the math skills is converting units and
that's because your image size so this
is when you're going to be measuring
your microscope image which is also
called a micrograph you'll be using a
millimeter ruler so your image size will
be recorded in millimeters the actual
size of cells and organelles is much
smaller it's going to be micrometers and
in order to use this formula you have to
have both of those sizes in the same
unit so if you have recorded your image
size in millimeter then you'll need to
convert it into micrometers so you have
the same units
to go from millimeters to micrometers
you multiply however many millimeters
you have by a thousand so if you
measured two millimeters that means you
have two thousand micrometers all you
could do the conversion the other way
round if your actual size is in
micrometers you can convert your image
size into micrometers as well and in
which case if you had two thousand
micrometers to convert that back into
millimeters they'd be divided by a
thousand and you have two millimeters so
just to go through a worked example
we've got here it's a bit blurry but
I'll put in all the details you could be
asked to work out what is the
magnification of this micrograph image
and they've given you a scale bar and if
you've shown a scale bar what that means
it is the length of that bar is
representing an actual size of 50
micrometers on this image so our scale
bar is the actual size and that has 50
micrometers what you then need to do to
work out the magnification we noted need
to know the image size and you don't
need to measure any cells for this you
are measuring what is the image size of
that scale bar so you'd line up your
ruler and measure how many millimeters
long your scale bar is and in this
example it's 20 millimeters and I
measured it so now we know we'd be doing
our image size which is 20 millimeters
divided by our actual size of the scale
bar which is 50 micrometers but we need
to get those into the same units so I'm
going to convert 20 millimeters into the
micrometer units to match the scale bar
so I need to do 20 times a thousand so
that gives me 20,000 that is our image
size divided by our actual size is 50 so
our magnification of this image is 400
times magnification so finally the last
skill
is the use of an eyepiece graticule and
how you calibrate it so I'm just going
to show you this image here of the
microscope first of all just so you can
see where the eyepiece graticule is
located so this eyepiece graticule is a
glass disk which is within the eyepiece
and that glass disc has a scale
scratched or etched onto it and that is
so you can line it up on top of whatever
you're visualizing to see how many
divisions on your eyepiece graticule
does the nucleus cover for example and
that can then be used to measure the
size the actual size of the objects that
you're viewing under the microscope
rather than using the formula that we
saw on the previous slide however it's
not quite as straightforward as that
because as you're using your light
microscope you will be potentially
moving between these different objective
lenses and each lens is a different
magnification and what that means is the
divisions on this eyepiece graticule
scale will be worth different distances
depending on how magnified the image is
and that's why we have to calibrate the
eyepiece graticule each time we use it
at a new magnification so to calibrate
it you need to use a second scale which
is called a stage micrometer and this is
a glass slide looks quite like a glass
microscope sight and it's called a stage
micrometer because this is the scale
that you're place on the stage and it's
measuring distances in micrometers so
the scale on it now mine isn't quite to
scale but that scale that you have
scratched onto this piece of glass is
two millimeters long and each single
division is worth 10 micrometers now
I've only done every 10 divisions on
this that's all I could fit in on the
diagram so one division is worth 10
micro
meters so 10 of these divisions is 100
micrometers long so if we go through
step by step then how you would use the
stage micrometer and the eyepiece
graticule to calibrate the graticule
it's a step 1
you'd place your stage micrometer on the
stage look through your eyepiece and
this is what you should see you have the
eyepiece graticule scale and then line
that up directly next to your stage
micrometer scale so step 1 line them up
so they're next to each other as you can
see in this image step 2 you need to
count how many divisions on the eyepiece
graticule scale fit into one division on
your micrometer scale now you might find
that easier to work out how I'm doing in
this world example we can see here that
we have 20 divisions from the eyepiece
graticule scale fit into 10 divisions on
the stage micrometer scale so I've got
20 fitting into 10 or in other words 2
of the divisions from the eyepiece
graticule scale fit into 1 division of
the stage micrometer scale so we have a
ratio of 2 divisions to 1 so now we've
got that we can link it back to what we
said about stage micrometers on our
stage micrometer 1 division is always
worth 10 micrometers so you can use that
to then work out at this magnification
what is one division worth on the
eyepiece graticule so we said one
division on the micrometer scale is 10
micrometers we said two divisions fit
into one on the micrometer and if one
division is worth 10 but to fit in each
time is 10 divided by 2 and we know then
that on the eyepiece graticule one
division is worth 5 micrometers at the
current magnification so now you can
take out your stage micrometer
put in whatever slide you want to use to
measure the distances or size of some of
the cells organelles so I've gone back
to the slide that we saw earlier on and
in this case I'm going to measure the
distance or the length of the nucleus in
this example now you would have all of
the subdivisions I've just not shown it
in this image so where we've got 40 to
50 50 to 60 you would have an extra 10
divisions so you could see 41 42 43 and
so on so we've said that we've worked
out that our eyepiece graticule
at this magnification 1 division is
worth 5 micrometers so I'm now going to
measure to see how many divisions the
nucleus covers and I'm estimating that
is 13 divisions we've got 10 and there
may be 3 more so the nucleus is 13
divisions long 1 division is worth 15 so
multiply that by our 5 that's 1
divisions worth 5 and 13 divisions so
that means in total the nucleus actual
size this magnification we've worked out
it's 65 micrometers now it will be that
distance or length even at every
magnification the only thing that would
change is what one division on the
eyepiece graticule is worth at different
magnifications so that's it for
microscopes so just to go over again a
summary of some of the key points you
need to know you need to say the
differences between optical and electron
microscopes and the electron microscope
has a much higher resolving power and
magnification and that's why the
electron microscopes can be used to see
the details inside of organelles the
formula I am so image size equals actual
size times magnification can be used to
work out the actual size of a structure
or the magnification from your
microscope image and lastly the actual
size of structures can be
measured using the microscope if you
have an eyepiece graticule in the
eyepiece but you have to calibrate it
first with the stage micrometer it's
take into account the different
magnifications and that's it for
microscopes for a level bulging now
there are quite a lot of linking videos
to this so if you want to just go over
some of the content or the math skills
over here are linked to the rearranging
the formula as well as you've got your
eukaryotic cells and prokaryotic cell
structures and last if you aren't
subscribed already click the logo here
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