This ice crystal was approximately 1 mm in size. The picture was made at 2:00pm and the temperature was 14 F. This type of a flake is called a stellar dendrite. Michael Peres

Michael Peres loves photographing the tiny details of our natural world. A professor of biomedical photographic communications at Rochester Institute of Technology, Peres specializes in capturing these intricate details by photographing with a microscope. Here, he explains how he does it.

I became interested in taking pictures of tiny things over 40 years ago while I was studying pre-med in college. I was exposed to this fascinating and invisible world while learning how to delineate muscle tissue from connective tissue using a light microscope. I was drawn into each new subject that I was asked to study and it still amazes me how things are organized. I love photographing snowflakes, flowers and other natural objects. I started sharing my work on Instagram in March 2014 and have been fascinated by the worldwide followers who are drawn to my images.

This photomicrograph is of the Anopheles mosquito. It is magnified approximately four times in this picture. The sample is called a whole mount and was a prepared slide that I purchased. The mosquito was approximately 2mm in size on the slide. Michael Peres

Finding a subject for microscopic photography

Finding a good subject starts with staying curious about the world. I find it is really important to be open-minded about potential samples. The other day, on a walk with my dog, he came home loaded with burrs. After removing as many as I could, I decided to examine one under the microscope. It was just a weed, but under the microscope it became elegant and complex.

Locating and handling small objects is a big part of the process’s success. Damaged samples or those with artifacts will have a different visual presentation than excellent samples and it’s important not to let the damage become the focus of the photography. Finding a sample without blemishes is the first priority for the type of pictures I want to make. That is not to say that the subjects of my photography are perfect—they are not.

Eastern White Pine
This photomicrograph is of a Pinus strobus or five needles. It was prepared in cross section section and reveals the various structures within the leaves including the vascular bundles shown in the center regions of the five needles and surrounding the green line-like cells. This image is magnified approximately five times. Michael Peres

There are two different things that I am thinking about when I prepare a sample to photograph—dissection and isolation, which of these takes precedent depends on the subject and its magnification. When photographing flowers, I typically use dissection scissors to remove petals to improve visibility of its structural elements. When I photograph aquatic organisms, I try to isolate them in a drop of water under a cover glass. Less is always more in this case. Each subject brings unique problems and require different methods to make the sample small enough, flat enough, or thin enough to photograph.

I also purchase prepared biological slides such as cross sections of plants or animal tissue—it is very difficult to prepare these types of slides without precision equipment. Wards Natural Science and Carolina Biological Supply Company sell thousands of subjects that are prepared for microscopic examination.

This picture is of malignate, a North American mineral and was photographed using polarized light revealing its birefringence. It was photographed from a geological slide that had been purchased which was .25mm thick. It is magnified approximately 25 times in this photomicrograph. Michael Peres

Gear needed to make a microscopic image

When shooting microscopic images my crucial pieces of gear are microscopes, a fiber optic light, a DSLR camera body, a macro lens, extension tubes, bellows and a tripod. When I’m photographing snowflakes, flowers or other subjects found in nature I’m often in my garage and I keep a ready supply of clean microscope slides and a piece of black velvet fabric for use as a background. I also have many needles, tiny brushes, and cotton tipped sticks that I use these for moving the samples and tidying up the area around them.

I typically use a compound light microscope for my photography. Light microscopes are quite common and easy to find. They can be expensive or relatively cheap, a student microscope might cost as little as $250 while a high-end research grade microscope might cost $200,000. A fairly good low-end research grade used microscope can be found for $5000.

This photomicrograph is of a squidlet, an immature squid from the species Loligo. It is magnified approximately eight times in this picture. This type of a sample is called a whole mount and was a prepared slide that I purchased. The immature squid on the slide was approximately 1mm. Michael Peres

Microscopes magnify subjects using two lenses. The first stage of magnification is produced by the objective lens and the second stage of magnification is the eyepiece. An objective lens has a focal length just like traditional photographic lenses that are used on a DSLR camera. Working distances are very small for this type of photography—a typical range of objective magnifications for a light microscope might include a 2x, 4x, 10x, 20x, 40, 60x or 100x. I pick my lens on the microscope based on the sample’s magnification requirements. Magnification influences the image’s depth of field, so a big thick sample (.5cm) benefit from low magnification, while very flat subjects need more magnification.

When using a light microscope it is possible to make pictures using a smartphone or a fixed lens camera and is a great starting point. With these cameras the lens should be placed at the eye point of the microscope, which can be located by holding a piece of paper approximately 1 cm from the eyepiece of the microscope. A very small dot of light will be visible on the paper, this is the eye point and where the lens should be directed. A small tripod can be helpful to hold the camera in place. Gaffer’s tape is often my best friend as a technical photographer because it lets me secure the phone of other elements of the system during my shoots.

A hepatica flower and fiber optic lights were used for the illumination. The photomicrograph includes the flowers stamen and pistil and is approximately three times magnified in this picture. Michael Peres

Although using a smartphone or fixed lens camera works, I like to use a Nikon D300s or a D800 with the lens removed and the body hung over the microscope’s eyepiece using a vertical copy stand or tripod. I also use extensions tubes or bellows on my camera to manage the ambient light—which can create flare and lower contrast. With a DSLR you need to align the camera’s sensor over the small dot of light from the eye point. To make a microscopic image I will align the sensor of my camera over the eyepiece at a distance where the circle of illumination produced by the microscope is large enough to cover the sensor without seeing the circle. Confirm that the light point has covered the sensor by checking the camera’s LCD screen. Be cautious when setting up though: it is very important to be aware of the distance between the eyepiece and the unprotected sensor.

Chicken Embryo
A chick embryo that was between 18 and 24 hours old. The developing brain, spine and vertebrae are beginning to form and become visible within the embryo. The chick is magnified five times in this photomicrograph. Michael Peres

Lighting your microscopic subject

I primarily use the microscope’s built in light and supplement it with fiber-optic lighting. When I look at a sample for the first time, I envision what I want the image to look like and work to get there. I make many tiny adjustments to the light’s position, which makes a large difference in the results. Some of the subjects I am photographing might be one or two millimeters in size, or smaller. How much fill light is needed or what makes the light’s angle just right are the decisions that I make before shooting. My strategy is based largely on how the light is working with the sample itself. The three styles of lighting I use most regularly are Kohler, Darkfield and Polarized, read on for more info about each of them.

Bursaria Truncatella
This picture is of Bursaria truncatella. The organisms were alive and swimming. Bursaria are ciliated freshwater organisms and live all over the world. They feed principally on paramecium. The aquatic invertebrates are magnified approximately 25 times in this photomicrograph. Michael Peres

How I use Kohler lighting when shooting microscopic subjects

This photomicrograph is of Taraxacum officinale or common dandelion. It was prepared in longitudinal section and includes the seed cup and developing seeds. This image is magnified approximately 15 times. Michael Peres

When I photograph a prepared “thin-section,” I am trying to create a neutral and uniform back lighting called Kohler illumination. This style of lighting allows the microscopist to maximize the contrast and resolution in the image and the production of uniform and even lighting behind the sample. I continue to use it because it creates portrait-like treatments of the things I photograph.

How I use Darkfield lighting while shooting microscopic subjects

Developing Bone
This picture was made from a slide that included developing human bone. It was photographed using darkfield illumination. The photomicrograph shows maturing bone cells and spongy bone and is magnified approximately 75 times. Michael Peres

I will also use darkfield, which makes an object glow against a black background, creating an astronomical type look. Darkfield lighting comes from behind a transparent subject and is shined through the sample at oblique angles. This style of lighting produces a very dramatic look, the downside is that everything will be lit when using darkfield including the things you do not want to have lit—like dirt, scratches or air bubbles.

How I use Polarized lighting while shooting microscopic subjects

Prescription Pill
This picture is of the Merck medicine foradil that is prescribed for asthma. The photomicrograph was made using polarized light and reveals the crystals that were formed as the chemical evaporated and dried. For this photo, a solid pill was dissolved in hot water and then coated onto a slide to evaporate. The colors represent different components of the chemical. The picture is magnified approximately 50 times. Michael Peres

I use polarized light when I am photographing samples that exhibit birefringence, which is a technical term that describes how a sample may or may not “show” rainbow-like-colors when placed into a polarized light microscope. Samples that include hairs, fibers, chemicals, minerals, some insect wings, and many synthetic objects will look as though they are made of rainbows when examined using polarized light. Polarized light is used to reveal internal information in these samples that otherwise may not be visible.

Prescription Pill
This picture is of the Merck® medicine foradil that is prescribed for asthma. The photomicrograph was made using polarized light and reveals the crystals that were formed as the chemical evaporated and dried after a solid pill was dissolved in hot water. This photograph also contains the edge of the cover glass that I used in the preparation that is .15mm thick. The colors represent different components of the chemical. The picture is magnified approximately 15 times. Michael Peres

Focusing while shooting microscopic photographs

To focus the magnified image with my DSLR I remove the camera’s lens and project the image directly into the DSLR’s camera body. The microscope focus controls are then used to focus the image in the camera’s viewfinder. Making a crisp image can be challenging. The DSLR’s viewfinder does not pick up the same amount of fine detail that the microscope can create, so things often look a little rougher in the viewfinder then they end up looking in the RAW files. It takes some practice to be able to predict how the image will appear once it is recorded.

It is also possible to focus using live view. In a dimly lit room, this can be very helpful for observing critical image definition within a sample. If you are using a smartphone that has been carefully positioned over the eyepiece, you will still want to focus the image displayed on the phone’s display by manipulating the microscope’s focus knob.

Processing your microscopic images

One of the biggest challenges in this type of work is creating contrast and structural delineation of internal parts and I try and do the hard work during the shoots. When I am photographing, I work slowly to inch my way towards making an interesting result using lenses and light.

I am very cautious about too much image processing. In post-processing I am most interested in tone management, setting white or black points and removing irrelevant dirt. I shoot RAW, open the RAW file in Photoshop and then I preprocess the structural details that are present in the file, but often not very visible. I also perform a minor amount of clarifying or sharpening at this stage. After making minor tone changes in Photoshop, I will sharpen the file using the high band pass filter.

The ice crystal was approximately 1.5 mm in size. The temperature was 21 F. This type of snowflake is called a stellar dendrite. Michael Peres

I think it is important for the images to not be totally flawless. Nothing is perfect in life and I think if the images are too perfect they might seem computer generated. My goal for this work is to make scientific photographs that operate in non-scientific environments and enable people to learn something new about the world they live in. My pictures feature real things and life also has blemishes and areas that are out of focus.

Sunscreen Chemical
This picture is of the chemical oxybenzone an ingredient of sunscreen. The photomicrograph was made using polarized light and reveals the crystals that were formed as the chemical evaporated and dried. The colors represent different components of the chemical. The picture is magnified approximately 35 times. Michael Peres