Tom Hata

What&#8217;s inside of a Mexican jumping bean?

Inspired by Manu’s previous post regarding focus stacking (https://microcosmos.foldscope.com/?p=7830), I decided to try out the technique myself. Since I was testing a focus locking prototype of the Foldscope, it was very easy for me to progress through a series of focal lengths. For my first attempt, I imaged the remnants of a bryozoan colony that had grown on a blade of marine algae that I found on the beach. Apologies in advance, I forgot to take a macroscopic picture of the algae and bryozoans. Bryozoans are tiny colonial invertebrates, and these particular ones happen to be epiphytes that settle and grow on top of algal blades.




 <a alt="20151121_211414.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_10552/20151121_211414.jpg">
 <img alt="20151121_211414" class="alignnone wp-image-10555 gcloudimg" data-attachment-id="10555" data-comments-opened="1" data-image-meta='{"aperture":"1.8","credit":"","camera":"LG-H815","caption":"","created_timestamp":"1448140454","copyright":"","focal_length":"4.42","iso":"50","shutter_speed":"0.066666666666667","title":"","orientation":"1"}' data-image-title="20151121_211414" data-recalc-dims="1" height="614" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_10552/20151121_211414.jpg" width="800"/>
 </a>


 Above is an image of the bryozoan colony. Each animal in the colony lives in its own individual chamber, and it feeds by extending its tentacles into flow. Although the animals are long-gone, the chambers within the rigid exoskeleton remain. Since the exoskeleton was 3-dimensional in nature, I thought it would be the perfect candidate for focus stacking.


 <a alt="bryozoanFocusStack.png" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_10552/bryozoanFocusStack.png">
 <img alt="bryozoanFocusStack" class="alignnone wp-image-10554 gcloudimg" data-attachment-id="10554" data-comments-opened="1" data-image-meta='{"aperture":"0","credit":"","camera":"","caption":"","created_timestamp":"0","copyright":"","focal_length":"0","iso":"0","shutter_speed":"0","title":"","orientation":"0"}' data-image-title="bryozoanFocusStack" data-recalc-dims="1" height="582" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_10552/bryozoanFocusStack.png" width="800"/>
 </a>


 Here is the result of stacking 8 images together in Photoshop (see http://petapixel.com/2014/09/07/tutorial-easily-focus-stack-using-photoshop-feature-probably-didnt-know/). I must admit, the results are amazing. You can see a high level of detail across virtually the entire field of view. I am guessing that the tentacles would have emerged from the circular openings on the right side of each chamber.




 Seeinng these promising results,  I decided to take things a step further: imaging an entire organism that was larger than the Foldscope’s field of view by stitching multiple images together. Previous attempts at stitching multiple images together proved unfruitful, largely because only part of each image was in focus. I had hoped that focus stacking would alleviate this issue. For this experiment, I decided to use another marine animal: a barnacle that I had removed from its home (see https://microcosmos.foldscope.com/?p=6788). Below are a couple of images of the critter (no focus stacking yet).


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 <img alt="20151122_111221" class="alignnone wp-image-10557 gcloudimg" data-attachment-id="10557" data-comments-opened="1" data-image-meta='{"aperture":"1.8","credit":"","camera":"LG-H815","caption":"","created_timestamp":"1448190741","copyright":"","focal_length":"4.42","iso":"50","shutter_speed":"0.025","title":"","orientation":"6"}' data-image-title="20151122_111221" data-recalc-dims="1" height="450" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_10552/20151122_111221.jpg" width="800"/>
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 </a>


 As you can see, it is nowhere near fitting within the field of view. In fact, this particular barnacle ended up spanning about 8 fields of view. In total, I took about 30 pictures of the barnacle at various places of its body and at different focal lengths. I created a single focus stacked image for each particular field of view, then combined these new images by aligning them in photoshop (using the auto-align function). The result turned out even better than I expected!


 <a alt="barnacle_stitched.png" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_10552/barnacle_stitched.png">
 <img alt="barnacle_stitched" class="alignnone wp-image-10560 gcloudimg" data-attachment-id="10560" data-comments-opened="1" data-image-meta='{"aperture":"0","credit":"","camera":"","caption":"","created_timestamp":"0","copyright":"","focal_length":"0","iso":"0","shutter_speed":"0","title":"","orientation":"0"}' data-image-title="barnacle_stitched" data-recalc-dims="1" height="611" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_10552/barnacle_stitched.png" width="800"/>
 </a>


 Though there are blurry regions, the overall image is very sharply in focus. This is my first time using the Foldscope to capture the entirety of an organism that is several millimeters long. Hopefully in the future I can find a freeware solution to create images such as these, perhaps through a program such as GIMP or ImageJ. I am very much open to ideas and suggestions!




 Here are a couple of bonus images using the same technique. First is a bee leg:


 <a alt="IMG_20151127_144300.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_10552/IMG_20151127_144300.jpg">
 <img alt="IMG_20151127_144300" class="alignnone wp-image-10559 gcloudimg" data-attachment-id="10559" data-comments-opened="1" data-image-meta='{"aperture":"0","credit":"","camera":"","caption":"","created_timestamp":"0","copyright":"","focal_length":"0","iso":"0","shutter_speed":"0","title":"","orientation":"0"}' data-image-title="IMG_20151127_144300" data-recalc-dims="1" height="757" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_10552/IMG_20151127_144300.jpg" width="800"/>
 </a>


 Second, here is the compound lens of the same bee’s eye:


 <a href="https://www.instagram.com/p/-mwe8cJsSz/">
 https://www.instagram.com/p/-mwe8cJsSz/
 </a>

Focus stacking and image stitching

This is a sample I’ve been holding on to for more than half a year, because until today I haven’t been able to image it to my satisfaction. Julia Pfeiffer Beach in Big Sur, CA is known for its amazing coastal features, like the Keyhole Arch.


 <img alt="20150621_154652" class="alignnone wp-image-13674 size-large gcloudimg" data-attachment-id="13674" data-comments-opened="1" data-image-meta='{"aperture":"1.8","credit":"","camera":"LG-H815","caption":"","created_timestamp":"1434901612","copyright":"","focal_length":"4.42","iso":"50","shutter_speed":"0.00052002080083203","title":"","orientation":"1"}' data-image-title="20150621_154652" data-recalc-dims="1" height="563" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13673/20150621_154652.jpg" width="1000"/>


 Unfortunately this particular feature doesn’t fit under a Foldscope. Another peculiar feature, much smaller in scale, is the abundant purple sand.


 <img alt="20150621_171656" class="alignnone wp-image-13675 size-large gcloudimg" data-attachment-id="13675" data-comments-opened="1" data-image-meta='{"aperture":"1.8","credit":"","camera":"LG-H815","caption":"","created_timestamp":"1434907016","copyright":"","focal_length":"4.42","iso":"50","shutter_speed":"0.00042016806722689","title":"","orientation":"1"}' data-image-title="20150621_171656" data-recalc-dims="1" height="563" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13673/20150621_171656.jpg" width="1000"/>


 The purple sand grains on the beach look to be created by erosion of the adjacent hills.


 <img alt="20160302_174450" class="alignnone wp-image-13676 size-large gcloudimg" data-attachment-id="13676" data-comments-opened="1" data-image-meta='{"aperture":"1.8","credit":"","camera":"LG-H815","caption":"","created_timestamp":"1456940690","copyright":"","focal_length":"4.42","iso":"50","shutter_speed":"0.016666666666667","title":"","orientation":"1"}' data-image-title="20160302_174450" data-recalc-dims="1" height="563" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13673/20160302_174450.jpg" width="1000"/>


 Excitedly, I loaded a sample of this sand on a paper slide…


 <img alt="20160302_172253" class="alignnone wp-image-13680 size-large gcloudimg" data-attachment-id="13680" data-comments-opened="1" data-image-meta='{"aperture":"1.8","credit":"","camera":"LG-H815","caption":"","created_timestamp":"1456939372","copyright":"","focal_length":"4.42","iso":"50","shutter_speed":"0.004","title":"","orientation":"6"}' data-image-title="20160302_172253" data-recalc-dims="1" height="563" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13673/20160302_172253.jpg" width="1000"/>


 and I was somewhat disappointed by the results. In bright field, the colors of the purple sand grains were completely obscured. This is where my initial exploration of the sample halted.


 <img alt="20160302_172141" class="alignnone wp-image-13679 size-large gcloudimg" data-attachment-id="13679" data-comments-opened="1" data-image-meta='{"aperture":"1.8","credit":"","camera":"LG-H815","caption":"","created_timestamp":"1456939301","copyright":"","focal_length":"4.42","iso":"50","shutter_speed":"4","title":"","orientation":"6"}' data-image-title="20160302_172141" data-recalc-dims="1" height="563" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13673/20160302_172141.jpg" width="1000"/>


 Many months later, I looked at the same sample under dark field (check out https://microcosmos.foldscope.com/?p=9060 for how to design a dark field aperture). The resultant image was indeed interesting, as you can begin determining the different colors of individual grains, but it still did not capture the deep purple I was seeing with my naked eye.


 Inspired by Marie’s post using an LED for side illumination ( see https://microcosmos.foldscope.com/?p=13486), I once again looked at my sample today.


 <img alt="20160302_172546" class="alignnone wp-image-13677 size-large gcloudimg" data-attachment-id="13677" data-comments-opened="1" data-image-meta='{"aperture":"1.8","credit":"","camera":"LG-H815","caption":"","created_timestamp":"1456939546","copyright":"","focal_length":"4.42","iso":"50","shutter_speed":"0.25","title":"","orientation":"6"}' data-image-title="20160302_172546" data-recalc-dims="1" height="563" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13673/20160302_172546.jpg" width="1000"/>


 Finally! The rich purple hues of the sand were clearly visible under the Foldscope. In retrospect, of course reflection microscopy was the solution to view the sample. It was especially apparent when I ran across opaque grains that would have appeared as dark shadows under transmission. I have made it a habit to look at my samples under these various viewing modes, as each mode seems to bring out a previously-unseen detail!


 DOI:
 <a href="https://dx.doi.org/10.15200/winn.145764.45105" target="_blank">
 10.15200/winn.145764.45105
 </a>
 provided by
 <a href="https://thewinnower.com">
 The Winnower
 </a>
 , a DIY scholarly publishing platform

Purple sand &#8211; Different ways to view color in your samples

<img alt="20160404_145526" class="alignnone size-large wp-image-14766 gcloudimg" data-attachment-id="14766" data-comments-opened="1" data-image-meta='{"aperture":"1.8","credit":"","camera":"LG-H815","caption":"","created_timestamp":"1459781726","copyright":"","focal_length":"4.42","iso":"50","shutter_speed":"0.016666666666667","title":"","orientation":"1"}' data-image-title="20160404_145526" data-recalc-dims="1" height="508" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_14765/20160404_145526.jpg" width="1000"/>


 Currently at Stanford, it’s literally raining caterpillars. Apparently, once every few years, there are population booms of Western Tussock Moth larvae
 <a href="http://bgm.stanford.edu/groups/grounds/ipm/tussock">
 http://bgm.stanford.edu/groups/grounds/ipm/tussock
 </a>
 . If you’re in the wrong spot, you’ll end up covered in these guys as they fall from the trees.

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 Of course, I couldn’t wait to put one of these guys under a Foldscope, so I caught one of them. As you can see, this little guy was perfectly content walking on the rim of a paper cup. Really, I left it there for about 2 hours and it did just that the whole time. As a Foldscope specimen, these guys are a bit large, so I had to think about how to tackle this challenge.


 <img alt="20160404_172313" class="alignnone size-large wp-image-14768 gcloudimg" data-attachment-id="14768" data-comments-opened="1" data-image-meta='{"aperture":"1.8","credit":"","camera":"LG-H815","caption":"","created_timestamp":"1459790593","copyright":"","focal_length":"4.42","iso":"100","shutter_speed":"0.033333333333333","title":"","orientation":"1"}' data-image-title="20160404_172313" data-recalc-dims="1" height="563" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_14765/20160404_172313.jpg" width="1000"/>


 I wanted to image the critter live, so I made a well for it using a stack of paper slides and glass cover slips (e.g.,
 <a href="https://microcosmos.foldscope.com/?p=10184">
 https://microcosmos.foldscope.com/?p=10184
 </a>
 ). It was so large that it took a stack of 5 paper slides. The next problem to tackle was that the caterpillar was incredibly opaque, so lighting it with the light module would not be of much use. Instead, I used side illumination with an LED (see
 <a href="https://microcosmos.foldscope.com/?p=13486">
 https://microcosmos.foldscope.com/?p=13486
 </a>
 ) so that light on the critter’s body would be reflected toward the lens. The results were simultaneously amazing and somewhat terrifying.

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 A close-up look revealed hairs upon hairs upon hairs. Also, visible are its constantly-moving mouth parts. I was intrigued by the hairs, so I lightly put a piece of tape across the critter’s back to extract loose hairs.


 <img alt="20160407_150817" class="alignnone size-large wp-image-14769 gcloudimg" data-attachment-id="14769" data-comments-opened="1" data-image-meta='{"aperture":"1.8","credit":"","camera":"LG-H815","caption":"","created_timestamp":"1460041696","copyright":"","focal_length":"4.42","iso":"50","shutter_speed":"0.0081967213114754","title":"","orientation":"6"}' data-image-title="20160407_150817" data-recalc-dims="1" height="563" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_14765/20160407_150817.jpg" width="1000"/>


 Bright-field images using the light module revealed part of the picture. The structures came into view, but the distinct colors – the reds, whites, yellows, and blacks – weren’t distinguishable in this mode. Once again I used side illumination, and the story became much clearer.


 <img alt="20160407_152245" class="alignnone size-large wp-image-14770 gcloudimg" data-attachment-id="14770" data-comments-opened="1" data-image-meta='{"aperture":"1.8","credit":"","camera":"LG-H815","caption":"","created_timestamp":"1460042565","copyright":"","focal_length":"4.42","iso":"50","shutter_speed":"0.25","title":"","orientation":"6"}' data-image-title="20160407_152245" data-recalc-dims="1" height="563" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_14765/20160407_152245.jpg" width="1000"/>


 <img alt="20160407_151722" class="alignnone size-large wp-image-14771 gcloudimg" data-attachment-id="14771" data-comments-opened="1" data-image-meta='{"aperture":"1.8","credit":"","camera":"LG-H815","caption":"","created_timestamp":"1460042242","copyright":"","focal_length":"4.42","iso":"50","shutter_speed":"0.25","title":"","orientation":"6"}' data-image-title="20160407_151722" data-recalc-dims="1" height="563" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_14765/20160407_151722.jpg" width="1000"/>
 <img alt="20160407_152314" class="alignnone size-large wp-image-14772 gcloudimg" data-attachment-id="14772" data-comments-opened="1" data-image-meta='{"aperture":"1.8","credit":"","camera":"LG-H815","caption":"","created_timestamp":"1460042594","copyright":"","focal_length":"4.42","iso":"50","shutter_speed":"0.25","title":"","orientation":"6"}' data-image-title="20160407_152314" data-recalc-dims="1" height="563" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_14765/20160407_152314.jpg" width="1000"/>


 Not only did the true color of these hairs become observable, it became obvious that the different colored hairs were structurally distinct from each other. I wonder if these differences serve a purpose, and how they correlate to the locations of these hairs on the caterpillar’s body.


 The most rewarding part of this experiment was that the caterpillar was unharmed. I was able to remove it from the slide and send it on its merry way. Perhaps I should have read up on this particular caterpillar before setting it free, however. I didn’t realize that it’s actually a pest to the point that there are efforts to actively control their populations. I guess my first hint should have been how unnatural it is for it to be raining caterpillars.

It&#8217;s raining caterpillars!

Foldscope at the beach in Hawaii

<a alt="20141008_172344.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_6788/20141008_172344.jpg">
 <img alt="20141008_172344" class="alignnone wp-image-6792 gcloudimg" data-attachment-id="6792" data-comments-opened="1" data-image-meta='{"aperture":"0","credit":"","camera":"","caption":"","created_timestamp":"0","copyright":"","focal_length":"0","iso":"0","shutter_speed":"0","title":"","orientation":"0"}' data-image-title="20141008_172344" data-recalc-dims="1" height="300" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_6788/20141008_172344.jpg" width="533"/>
 </a>


 Harbors and rocky shorelines are often littered with barnacle tests (or shells), yet relatively few people have actually seen the animals that live inside of  these dome-shaped homes.  Here’s some high-speed footage that I took of a barnacle feeding in a wave.

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 <iframe allowfullscreen="true" class="youtube-player" height="563" src="https://www.youtube.com/embed/9GpDss0YvzQ?version=3&amp;rel=1&amp;fs=1&amp;autohide=2&amp;showsearch=0&amp;showinfo=1&amp;iv_load_policy=1&amp;wmode=transparent" style="border:0;" type="text/html" width="1000">
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</div>

 Barnacles are crustaceans, so they are related to animals such as crabs and lobsters.  They are in fact somewhat crab-like in appearance once you pull them out of their tests.  Unlike crabs and lobsters, barnacles have adapted to a sessile life by hanging upside down inside of their tests and extending their feeding legs (cirr) into the water to capture food particles.

<figure aria-describedby="caption-attachment-6791" class="wp-caption alignnone" id="attachment_6791" style="width: 533px">
 <a alt="20150602_141109.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_6788/20150602_141109.jpg">
 <img alt="Barnacle Cthamalus fissus" class="wp-image-6791 gcloudimg" data-attachment-id="6791" data-comments-opened="1" data-image-meta='{"aperture":"0","credit":"","camera":"","caption":"","created_timestamp":"0","copyright":"","focal_length":"0","iso":"0","shutter_speed":"0","title":"","orientation":"0"}' data-image-title="20150602_141109" data-recalc-dims="1" height="300" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_6788/20150602_141109.jpg" width="533"/>
 </a>
 <figcaption class="wp-caption-text" id="caption-attachment-6791">
 Barnacle Cthamalus fissus
 </figcaption>
</figure>
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 <a alt="20150602_142621-e1433294821264.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_6788/20150602_142621-e1433294821264.jpg">
 <img alt="Barnacle feeding legs, high magnification" class="wp-image-6790 gcloudimg" data-attachment-id="6790" data-comments-opened="1" data-image-meta='{"aperture":"0","credit":"","camera":"","caption":"","created_timestamp":"0","copyright":"","focal_length":"0","iso":"0","shutter_speed":"0","title":"","orientation":"0"}' data-image-title="20150602_142621" data-recalc-dims="1" height="444" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_6788/20150602_142621.jpg" width="532"/>
 </a>
 <figcaption class="wp-caption-text" id="caption-attachment-6790">
 Barnacle feeding legs, high magnification
 </figcaption>
</figure>

Barnacle feeding legs

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 </a>


 Above is the tree of life, so far, as chronicled by our amazing Foldscope community! Currently represented in our 800+ posts so far are over 150 species from over 75 orders. I hope you take a second to find the small space that we occupy on this tree and appreciate the vast diversity of microstructures and body plans that form the rest of this budding web. There were almost too many beautiful images to chose from, so represented in this collage is just a small fraction of community-created content. In case anyone is curious, there are just 2 species of bacteria identified in our posts and none yet from the kingdom of Archaea! The rest of the posts, of course, are Eukaryotes (some fungi, but mostly plants and animals).


 Now that we’re on our way to creating a rich database of observations, I have a couple of open challenges for the community:


 1. For those interested in a particular group of living things (e.g., ants, fungi, flowering plants) I challenge you to search for the posts containing these living things and identify them the best you can. Create your own ID guide using Foldscope pictures!


 2. Find an organism that’s not yet represented here and document it. We’d love to see how far the Foldscope tree of life can be filled out!






 In case anyone would like to know how to make a tree of life like this:


 1. I manually combed through ALL of the Foldscope posts so far, recording (when available) the common and scientific names of the specimens (and their phylogenetic information such as kingdom, phylum, class, order, etc.) in a google spreadsheet.


 2. I exported the column containing the ORDERS of each specimen as a .txt.


 3. I Uploaded this file to NCBI’s taxonomy browser (
 <a alt="wwwcmt.cgi" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_7643/wwwcmt.cgi">
 http://www.ncbi.nlm.nih.gov/Taxonomy/CommonTree/wwwcmt.cgi
 </a>
 ) to create a text version of the tree of life.


 4. I exported this data as a phylip tree (under the save as function), which exports the data in Newick tree format.


 5. I uploaded my data to the Interactive Tree of Life (iTOL
 <a href="http://itol.embl.de/">
 http://itol.embl.de/
 </a>
 ).


 6. I edited and exported this tree as an Adobe Illustrator file.


 7. Prettied it up and added the Foldscope images in Illustrator and Photoshop.


 For those in the community with expertise in generating phylogenetic trees, I would love tips! This trial-and-error method was a great learning experience to create a one-off tree, but we are always thinking of new features to add as the community grows. One potential idea I have in mind would be a dynamic Foldscope web of life accessible from the front page.




 -Tom

The Foldscope tree of life

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 While visiting a friend’s home, I noticed they had a pear tree. I instantly had a flashback to a botany lesson from an old biology course. Pears have a peculiar texture, one that is often described as “gritty”. This texture is caused by the stone cells present in the fruit’s flesh.




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 I sliced a thin section of the fruit and immediately found clusters of these stone cells throughout. Notice that there are faint but distinct “tunnels” running from the stone cells to the periphery.

The gritty truth about pears

I received a call out of the blue from a good friend of mine. He was driving up from Los Angeles to the bay area, because, apparently, one of his favorite tattoo artists was secretly working out of a parlor in San Jose for only 1 week! My friend was tipped off by some cryptic remark that this artist had made on instagram referring to this particular shop in San Jose. Apparently the artist, Koji Ichimaru, has such a huge following that he was booked solid within an hour of posting this tiny hint.


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 As was to be expected, the tattoo itself was amazing in detail and playful take on a classic design–a Dia de los Muertos candy skull wearing a samurai helmet. This made me wonder what pigmented skin would look like at a macroscopic level. Adapting a technique I learned from Manu, I was actually able to image the surface of my friend’s skin! Not only that, but I could pan across the entirety of the tattoo.


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 The video itself makes a great guessing game for anyone watching it without context… the first guess  I usually hear is “insect”. What struck me was the depth of pigmentation in the skin to the point that we cant immediately recognize it close up.

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 For now I’ll leave the exact technique I used a mystery (to be revealed in a future post, and hint, it does not involve shoving an entire human under a Foldscope), but I hope someone can take a guess and figure it out!

Tattooed skin &#8211; putting a human under a Foldscope?

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 Aiptasia are unusual beasts. To a naive eye, these palm-like anemone could easily be confused for plants. In fact, aquarists often refer to them as weeds, owing to their ability to rapidly grow and multiply in seawater aquaria. These creatures are most definitely animals. However, they do serve as a home for photosynthetic algae (dinoflagellates). In this symbiotic relationship, the algae provide energy (e.g., lipids and sugars) for the anemone in exchange for protection and nutrients. Within this sea of relatively large adults, I found a tiny, foldscopable polyp. almost invisible to the naked eye.

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 Aiptasia in a wax well
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 I mounted the tiny polyp on a glass slide, usinng a ring of wax to act as a well.

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 Symbiosis in action! I was shocked to make out individual dinoflagellates (the round balls) in the anemone’s tentacles. Not only that, many of these symbionts appeared to be rapidly traveling on internal highways. I wonder what the purpose of this rapid circulation is, and what sort of internal plumbing in the anemone would give rise to such interesting internal flows.


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Symbiosis!

Have any of you had this difficulty with the Foldscope? After scanning your sample for a few minutes, you find an awesome micro-structure you’ve never seen before. Excitedly, you hand your Foldscope to a friend so they can see it too, and they see… nothing. Not only has the sample gone out of focus, but the lens has moved quite a bit from the location you were just looking at. The Foldscope team has been very much aware of this hurdle, and we have been thinking of innumerable ways to tackle this issue.


 I am happy to report that the development of a  focus-locking model of Foldscope is well on its way. Meaning, once you find a cool microstructure and get it into focus, it’s set. You can hand the Foldscope to a friend and they will see the exact same thing you just saw (unless you have a living sample moving about, see the copepod below!).


 Here’s an image of the latest prototype at work, and a process I like to refer to as laptop microscopy:


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 I can’t reveal too much right now, as the design is subject to change, but I would like to leave some sample footage of focus locking in action.

Focus-locking Foldscopes (in the works!)

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 Thanks to an ongoing experiment, the lab is flooded with animals! The animal du jour is everyone’s favorite model organism, the nematode C. elegans.  The trouble with imaging C. elegans under the Foldscope is that they are (1) very small, (2) mostly transparent, and (3) highly mobile (Most of the time. See the video at the end for an example of not-so-elegans swimming). This makes tracking an individual on a glass slide a daunting task. One trick to increasing the contrast of transparent organisms is using dark-field microscopy. I used a method described in a previous post (https://microcosmos.foldscope.com/?p=7042) to achieve this. As this is my first pass at this technique, this post serves as evidence of my trials and tribulations. The image above distinctly shows the difference between an image taken in bright field (top) and dark field (bottom). It was much easier to locate C. elegans on the slide in dark field, as its features exhibited much greater contrast. The image below is… somewhere in the middle of the previous two, although it was an attempt at dark field. I’m guessing I needed to make ink dot on the tape much darker to obtain a stereotypical dark-field image, although the specimen’s features do contrast nicely. Tomorrow will be another day to refine this technique… more to come.


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 UPDATE: Turns out you can get much, much better dark field results with a paper aperture.


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 I cut this piece on the laser cutter, leaving the middle piece attached by 3 thin strips. I usually shy away from superlatives, but the resultant images absolutely blew my mind. Even after dozens upon dozens of Foldscoping, I haven’t even begun to scratch the surface of this tool’s potential! For videos of dark field in action, see below.


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C. elegans in bright field and dark field

In order to investigate the resolution of the Foldscope when paired to my particular phone (an LG G4), I seeded a slide with 45 micron and 6 micron polystrene beads. Below are images taken in dark field (top) and bright field (bottom). The 140x lens has a theoretical resolution of about 2 microns, so, expectedly, I was able to resolve the beads just fine. I noticed an odd effect present in the dark field image but absent in the bright field. Namely, the 45 micron beads were acting as microlenses in tthe dark field image. Curiously, this effect is absent in bright field. I wonder if I can create a microlens array out of these larger beads if I can form a mono-layer of them (if money were not an object… these beads are pretty expensive).


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 As a bonus, here are dark and bright field images of just the 6 micron beads in high magnification. The high mag lens was much more finicky, but we can see individual beads clearly.


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microbeads (which unexpectedly act like lenses)

For as long as I’ve owned a Foldscope, there’s been one animal that I’ve been trying to image in vain. Namely, the tardigrade (or water bear). These extreme animals (that resemble microscopic, 8-legged pigs) can live for years without food or water and can even survive outer space!


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 This week, I was able to finally find my first tardigrades in the wild! Here’s how I did it:


 

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 I soaked a piece of moss in a petri dish with distilled water.
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 After about an hour, I scanned the water in the dish. There was so much microscopic life that I got distracted for a few hours, but eventually I did find several tardigrades. I was using a dissecting scope in dark field to scan through the water sample quickly. Hopefully I can find a way that won’t need this in the future.
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 Here is a tardigrade in bright field, approximately 100 microns in length.
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 Here is another in dark field. Notice that it actually does crawl along the substrate with its 8 legs. It’s hard for me to fathom walking at such a tiny scale.
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 These guys actually move pretty quickly, so it helped to pin the tardigrades to a small region of a slide by limiting the size of the droplet on the slide.


 




 
 I inadvertently made a tiny obstacle course for one of them, and you can see it fighting against surface tension on the air-water interface. I’m wondering if I can make a rudimentary calculation of force generation from this. Check out ~35 sec in, where the tardigrade makes himself a little port hole.
 

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Hunting for tardigrades (water bears)

In my ongoing quest to explore marine microfauna, I recently observed rotifers under a Foldscope. Like many motile marine invertebrates, rotifers use cilia to maneuver through the water column. They also use these cilia to feed by generating currents.
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 The anterior end of the animal is in the top right of the pictures, and there are 2 crowns of cilia on either side (this image in high mag). The cilia on these crowns beat in waves that generate current, allowing locomotion and feeding. Please excuse the psychedelic color changes in the first video, this was due to the light source oversaturating my camera phone’s sensor. I remedied this by using several layers of Scotch brand magic tape to diffuse the light.

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 High magnification was necessary to even see the cilia beating, as they appeared as a blur in low magnification. For scale, those little spheres of algae are approximatey 4 microns in diameter. I was curious to see whether I could observe the ciliary waves by filming it in high speed. Here’s a video of the cilia beating, filmed at approximately 130 fps and slowed down 8x. Although I wasn’t able to get the cilia entirely in focus, there are a number of curious things happening. First, the algae provide an excellent set of particles to visualize the flow patterns created by the cilia. Second, the rotifer would periodically stop feeding and would retract. I’m wondering if this is because it has caught a particle, or if there is some other reason for this behavior.

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 BONUS! I was also able to get my hands on a few anemone polyps (seen here in dark field and low mag). These polyps were so tiny that I was able to just barely fit them on a microscope slide, with 2 pieces of double-sided tape used as spacers.
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 My challenge was to see whether I could record an anemone capturing and feeding on a rotifer. After a few hours and a dozen trials, success was at hand. Below is an anemone with a rotifer trapped in its mouth (the round shape on the right end), about to be consumed.


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 In fact, I was able to capture the process of consumption over a 6-minute span.

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 By expanding and contracting, it looks like the anemone forces the rotifer down using a combination of cilia and peristaltic movement. At the end, you can even see the intact rotifer in the anemone’s gut! Here’s the same video, sped up 24x to get a better sense of the overall ingestion process.

A food web in a fluid droplet (algae, rotifers, and anemone)

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 Continuing our series of instruction translations, here are the Foldscope instructions in Swahili. Thank you to Ola Jahanpour for the translation.

Foldscope instructions translated in Swahili

Here’s a phenomenon that is perplexing me, but perhaps the answer is simple. The legs of small spiders are so thin that you can often see the “blood” or hemolymph flowing through them with the Foldscope. Though this is visible even in low magnification, here I took a closer look with the high magnification lens. For the first time, though, I was able to capture the twitching of the leg coupled with footage of blood flow. What puzzles me is this: spiders use hydraulic pressure to extend their legs, which is why their legs curl up when they die (the pressure suddenly decreases). For blood to be flowing, there must be a pressure gradient as well. How does a spider simultaneously manage both blood flow and coordinated movement? This may be a simple question for someone familiar with a spider’s body plan… but this has also inspired me to undertake a more comprehensive study mapping blood flow and body movement in a single spider. More to come!


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Tom Hata

Earth

Posts

What’s inside of a Mexican jumping bean?

Focus stacking and image stitching

Purple sand – Different ways to view color in your samples

It’s raining caterpillars!

Locations

Statistics

Type of Sample

Foldscope Lens Magnification