Taking pictures of bugs with a scanning electron microscope is a way to encourage interest in microscopy among younger students and children.  (I really enjoy it myself.)  Getting the best image of our insect friends can be something of a challenge though.  In this blog I will describe some basic techniques for imaging bugs.

Think small

I like to choose small bugs, they may look like a spec to the naked eye but they tend to look more interesting when you image them in the SEM.  The insect in the above image has an entire body length of approximately 1.5 mm and a width of .5 mm.

Many people think that a big bug will be interesting to look at, but larger insects present a number of problems in the SEM.  First of all the entire insect may not fit in the field of view of the electron microscope and the part that can be imaged is not that interesting.  Secondly large bugs take a long time to dry out and they may become moldy before they are completely dry.  You can try placing them in a container with some desiccant to dry them.

Be prepared

I usually carry some small disposable centrifuge tubes with me in case I encounter an interesting insect or other arthropod.  The insect in the photo found its way into my car when I was at the top of Pike's Peak in Colorado (4302 m or 14,000 ft).  It is a type of springtail that is capable of leaping 30-40 cm into the air in spite of its small size.

Keep it clean

Some students like to bring in dead insects that they have found.  These will usually be covered with debris.  Even a fingerprint will deposit debris over an insect.  I find it usually works best if you find a bug alive and store it in a centrifuge tube until it has shuffled from the mortal coil.

Be careful

Usually they will be whole and relatively clean when you pour them out of the centrifuge tube.  You have to be extremely careful not to break any limbs or antennae off when preparing the insect.  At this point I will usually work with a stereo microscope.  I use a small amount of silver paint or colloidal graphite and deposit it on a specimen stub.  Then use a fine tipped tweezers to transfer the bug onto the specimen stub.  If you use too much silver paint it will tend to wick up onto the insect and cover up details.

Avoid charging

After they are mounted they usually have to be sputter coated with gold or carbon to image in the SEM to prevent charging.  They can still have problems with charging because they have a three dimensional shape and the sputtered gold will not be able to wrap around all of the shapes.  Charging occurs when electrons used to scan the specimen build up on the surface of a non-conducting specimen.  Eventually the charge the builds up can be enough to deflect the electrons which are used to form the sample image.

Using a backscattered electron detector can be used to eliminate much of the charging affect.  The following image is taken with a standard secondary electron detector.  Notice that the hairs are all much lighter than the body parts of the insect.  There is also a very bright spot on the left antenna.  Secondary electrons are much lower energy than backscattered electrons so they are more susceptible to charging. 

Following is the same image taken with backscattered electron imaging.

Notice that the contrast is much more subtle in this image, it has a more solid feel to it.

You may be able to do some cleaning of the insect.  I used a small tweezers with a hair in them to remove some debris from the mouth parts of this insect.

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Tags: Nanotechnology, SEM

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Publications by A. Paszternák:

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pH and CO2 Sensing by Curcumin-Coloured Cellophane Test Strip

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Directed Deposition of Nickel Nanoparticles Using Self-Assembled Organic Template,

Organometallic deposition of ultrasmooth nanoscale Ni film,

Zigzag-shaped nickel nanowires via organometallic template-free route

Surface analytical characterization of passive iron surface modified by alkyl-phosphonic acid layers

Atomic Force Microscopy Studies of Alkyl-Phosphonate SAMs on Mica

Amorphous iron formation due to low energy heavy ion implantation in evaporated 57Fe thin films

Surface modification of passive iron by alkylphosphonic acid layers

Formation and structure of alkylphosphonic acid layers on passive iron

Structure of the nonionic surfactant triethoxy monooctylether C8E3 adsorbed at the free water surface, as seen from surface tension measurements and Monte Carlo simulations