11: Using the Microscope
- Page ID
- 123685
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\(\newcommand{\avec}{\mathbf a}\) \(\newcommand{\bvec}{\mathbf b}\) \(\newcommand{\cvec}{\mathbf c}\) \(\newcommand{\dvec}{\mathbf d}\) \(\newcommand{\dtil}{\widetilde{\mathbf d}}\) \(\newcommand{\evec}{\mathbf e}\) \(\newcommand{\fvec}{\mathbf f}\) \(\newcommand{\nvec}{\mathbf n}\) \(\newcommand{\pvec}{\mathbf p}\) \(\newcommand{\qvec}{\mathbf q}\) \(\newcommand{\svec}{\mathbf s}\) \(\newcommand{\tvec}{\mathbf t}\) \(\newcommand{\uvec}{\mathbf u}\) \(\newcommand{\vvec}{\mathbf v}\) \(\newcommand{\wvec}{\mathbf w}\) \(\newcommand{\xvec}{\mathbf x}\) \(\newcommand{\yvec}{\mathbf y}\) \(\newcommand{\zvec}{\mathbf z}\) \(\newcommand{\rvec}{\mathbf r}\) \(\newcommand{\mvec}{\mathbf m}\) \(\newcommand{\zerovec}{\mathbf 0}\) \(\newcommand{\onevec}{\mathbf 1}\) \(\newcommand{\real}{\mathbb R}\) \(\newcommand{\twovec}[2]{\left[\begin{array}{r}#1 \\ #2 \end{array}\right]}\) \(\newcommand{\ctwovec}[2]{\left[\begin{array}{c}#1 \\ #2 \end{array}\right]}\) \(\newcommand{\threevec}[3]{\left[\begin{array}{r}#1 \\ #2 \\ #3 \end{array}\right]}\) \(\newcommand{\cthreevec}[3]{\left[\begin{array}{c}#1 \\ #2 \\ #3 \end{array}\right]}\) \(\newcommand{\fourvec}[4]{\left[\begin{array}{r}#1 \\ #2 \\ #3 \\ #4 \end{array}\right]}\) \(\newcommand{\cfourvec}[4]{\left[\begin{array}{c}#1 \\ #2 \\ #3 \\ #4 \end{array}\right]}\) \(\newcommand{\fivevec}[5]{\left[\begin{array}{r}#1 \\ #2 \\ #3 \\ #4 \\ #5 \\ \end{array}\right]}\) \(\newcommand{\cfivevec}[5]{\left[\begin{array}{c}#1 \\ #2 \\ #3 \\ #4 \\ #5 \\ \end{array}\right]}\) \(\newcommand{\mattwo}[4]{\left[\begin{array}{rr}#1 \amp #2 \\ #3 \amp #4 \\ \end{array}\right]}\) \(\newcommand{\laspan}[1]{\text{Span}\{#1\}}\) \(\newcommand{\bcal}{\cal B}\) \(\newcommand{\ccal}{\cal C}\) \(\newcommand{\scal}{\cal S}\) \(\newcommand{\wcal}{\cal W}\) \(\newcommand{\ecal}{\cal E}\) \(\newcommand{\coords}[2]{\left\{#1\right\}_{#2}}\) \(\newcommand{\gray}[1]{\color{gray}{#1}}\) \(\newcommand{\lgray}[1]{\color{lightgray}{#1}}\) \(\newcommand{\rank}{\operatorname{rank}}\) \(\newcommand{\row}{\text{Row}}\) \(\newcommand{\col}{\text{Col}}\) \(\renewcommand{\row}{\text{Row}}\) \(\newcommand{\nul}{\text{Nul}}\) \(\newcommand{\var}{\text{Var}}\) \(\newcommand{\corr}{\text{corr}}\) \(\newcommand{\len}[1]{\left|#1\right|}\) \(\newcommand{\bbar}{\overline{\bvec}}\) \(\newcommand{\bhat}{\widehat{\bvec}}\) \(\newcommand{\bperp}{\bvec^\perp}\) \(\newcommand{\xhat}{\widehat{\xvec}}\) \(\newcommand{\vhat}{\widehat{\vvec}}\) \(\newcommand{\uhat}{\widehat{\uvec}}\) \(\newcommand{\what}{\widehat{\wvec}}\) \(\newcommand{\Sighat}{\widehat{\Sigma}}\) \(\newcommand{\lt}{<}\) \(\newcommand{\gt}{>}\) \(\newcommand{\amp}{&}\) \(\definecolor{fillinmathshade}{gray}{0.9}\)Content Skills
• Recognize and give the function of the parts of a compound microscope
Physical Skills:
• Properly care for a compound microscope
• Correctly use a compound microscope
We use a microscope to visualize objects that are too small to see with the naked eye. This is especially critical in Biology as all cells are quite small and require a microscope for visualization.
Parts of the Compound Light Microscope
Use Figure 1 to identify the various parts of your microscope. Read each description below, find them on your microscope and on the lab handout and manipulate the parts only where indicated. Before you start, make sure that the shortest objective is in the light path.
Figure 1. Parts of the Compound Microscope.
- Light source. The compound microscope uses transmitted light to illuminate a transparent (or super thin) sample mounted on a glass slide. Find the light source/illuminator, the off/on switch, and the rheostat which can vary the light intensity so that it is not too bright.
- Condenser. A condenser is composed of a condenser lens and iris diaphragm to focus the light on the specimen so that it is equally and fully illuminated. The lever for the iris diaphragm of the condenser is used to open and close the condenser to increase or decrease the area illuminated and thereby light intensity.
- Stage. Your slide containing a sample is placed on a mechanical stage over a central hole. Position a slide on the stage by releasing the tension in the spring loaded moveable arm and using this slide clip to hold your slide in place. There are two knobs to the right or left of the stage: one to move the specimen forward or backward and one to move it side to side.
- Focusing knobs. The coarse focus knob is for use with the low power objectives ONLY whereas the fine focus knob is critical for focusing especially with higher power objectives.
- Objectives. Our microscopes have four different objectives 4 objectives: a 4x (scanning), 10x (low power), 40x (high dry) and 100x (oil immersion). Note that they are labeled and color coded for ease of use. The other number written on the objective is the numerical aperture (NA). The higher the numerical aperture, the greater the resolving power.
- Ocular. The magnifying lens that you look into is called the ocular. Our microscopes have two 10x oculars (and are therefore stereomicroscopes). Since each objective has a different magnifying power, the total magnification is calculated by multiplying the magnifying power of the ocular by that of the objective.
Importantly, the objectives are parfocal meaning that once one objective has been focus, you can rotate another one into place without losing the focus. You will likely need to very finely adjust the fine focus, but it specimen will be very close to being in focus. Also, the center of the field of view will stay the same as you move from one objective to the next.
These features will allow you to generally focus at the low power (4x) where focusing is MUCH easier and then move to a higher power and only have minimal adjustments.
Using the Microscope
Initial set up
- Carry your microscope to and from your lab bench by grasping the arm with your dominant hand and supporting the base with your other hand. Make sure to always keep the microscope upright and do not try to carry anything else at the same time.
- Remove the dust cover and, if needed, clean the lenses with a piece of lens paper. Never use anything else to clean your microscope as it could scratch and damage your microscope.
- Unwind and plug in power cord
- Turn on the light source – switch on right side of microscope
- Dial or Slider on right side of microscope will adjust the light intensity, depending on the sample and magnification used.
- Adjust the ocular focus:
- With your left eye closed and a slide on the stage (see below), focus your right eye on something on the slide using the normal focus knobs.
- Then close your right eye and adjust the focus of the left eye using the focus ring on the ocular lens.
- Adjust the spacing of the oculars so that you can look with both eyes simultaneously.
Viewing a Sample
- Select lowest power (4x) objective by turning the black ring just above where the objectives attach to the body of the microscope.
- Important: Never rotate by grabbing or putting pressure on an objective lens itself.
- Set slide on stage and hold in place with clip.
- Use the coarse and fine focus knobs to focus on the sample at lower magnification before rotating the collar (not touching the objectives themselves!) to switch to a higher objective.
- Refocus at the desired magnification, using the fine focus knobs.
- If you want to use the 100x oil objective, check with your instructor before using. Turn the collar so that the slide is centered between the 100x oil lens and the adjacent objective lens (neither one is clicked into place).
- Carefully put one small drop of immersion oil on the cover slip.
- Slowly rotate the collar so that the 100x objective clicks in place.
- The immersion oil should form a connection between the slide and the lens.
- Important: Use care never to get oil on any part of the microscope other than the 100x oil lens. Especially do not get oil on the other dry objectives – this can severely damage them.
- Adjust the level of illumination with the slider on the side of the microscope.
Tips For Focusing
- Always start by getting in focus at the lowest power objective (4x). Then increase objective strength one at a time, focusing before moving to a higher objective.
- If you have trouble finding the correct focal plane in which to view your sample, try focusing first on the edge of the cover slip.
- Then, without moving the focus knob, scroll the stage over so that the specimen is in the field of view.
- It now should be close to in focus, requiring only minor adjustment with the fine focus knob.
Putting the Microscope Away
- Clean the immersion oil off the 100x oil objective if used.
- Tear off a piece of lens paper and gently swipe it across the lens.
- Never use a Kimwipe or anything but lens paper on a lens; it could scratch the lens.
- Rotate the paper to a clean portion or get a fresh sheet and gently swipe again.
- Repeat until there is no visible oil coming off the lens onto the paper.
- Never “scrub” on the lens by pressing with your finger or rubbing back and forth – any tiny pieces of dirt can scratch the lens.
- If desired, clean the other lenses in the same way, with clean lens paper.
- They should not have any oil on them, but sometimes they will get a little dusty or dirty.
- Wipe up any oil that accidentally got on other parts of the microscope.
- The stage is most likely to become contaminated by stray smudges of oil.
- If necessary, moisten the corner of a Kimwipe with a little ethanol to help wipe up the oil. Remember, Kimwipes are never to be used on the lenses themselves.
- Turn the objectives so that either no objective points down toward the stage or the 4x objective points down (this helps to prevent damage to the objectives)
- Adjust the stage scroll knobs so that it doesn’t stick out.
- Wrap the cord around the holders.
- Carefully store it in the appropriate place in the microscope cabinet.