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- Hi, I'm Dr. Garcia at the Biotechnology program
at NC State University and today I'm going to be talking
to you about how to use our inverted microscopes.
I wanted to start with a little orientation of
the different parts of the microscope.
So, first of all, the light source for this microscope
is actually up here and passes through the sample
down this way and then the objectives
are actually under the stage
and we can switch the objectives simply by rotating
this little nose piece, it's called, down here.
We have two different kinds of objectives on our nose piece
indicated by the color.
So the silver objectives are optimized
for florescence microscopy
while the black objectives are optimized for
bright field and phase contrast microscopy.
Keep these in mind because it's really important
to use the correct objective when you're using
or when you're going to be taking pictures
and also when you're visualizing your cells.
In order to focus our samples, we're going to use
this knob right here.
It's actually two kobs where the larger one
focuses coarse adjustment while the smaller one
focuses the fine adjustment.
Obviously we need to turn on our microscope first of all.
So, in order to turn on the bright field
or the white light, the switch is right here,
this green switch,
and then there's also this dimmer knob
which can actually change the intensity
of the white light that's passing through the sample.
Next is the eye piece which is where you look through
the microscope in order to see your samples
and this eye piece is adjustable so that you can
make it comfortable for you.
So this is a very important knob for you guys
to know about because it switches the lights,
or the direction of the light between the eye piece
and the camera.
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Last thing to talk about is the stage.
Stage has a slide holder
if you are looking at cells that
have been mounted on slides.
Alternatively, you can take this piece out
if you're looking at a plate like we will be doing today.
Plate will fit within this little rectangle
and then we can use this knob over here
to move the plate in any direction.
Now that we've got our basics about how this microscope
is set up, we can now start to focus on our cells.
So I've got our plate already on the stage here.
What we're gonna do is we're gonna use bright field.
So we want to use the black for its objective,
we want to bring the objectives all the way down,
and we do that by turning the coarse adjustment knob
all the way towards the wall until it stops.
Once it's all the way towards the wall
and it can no longer go any further,
look through the eye piece
and then slowly turn the knob towards you
until the cells come into focus.
If you look through the eye piece
and you can't see any light actually coming through,
one of the things that you might wanna try doing
is changing the dimmer switch
or increasing the intensity of the white light
that's coming through.
You should be able to see the light passing through
the sample on the stage.
The other thing to try is to make sure that the knob
over here is set to bino which means that the light
is coming through to the eye piece
as opposed to going to the camera.
If you've already tried these things
and are still not able to see your cells,
or if you're having trouble focusing on your cells,
please ask your instructor.
Do not try manipulating any of the other knobs or sliders
because those have already been placed
in the correct position by your instructor.
Once you've been able to focus on your cells at 4X,
you can change to a higher magnification
by changing the objective.
Then use the fine adjustment to bring the cells into focus.
To visualize our cells using florescence,
we're going to need to turn off the bright field,
use the silver objectives,
and the light source at the back of the microscope.
In order to visualize grain fluorescent proteins,
we're going to need to use the cyan LED
for to see all the light source.
You can focus on your cells using the fine adjustment knob.
In order to visualize red florescence,
use the green LED in the back or the G setting.
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