Matthew Bull

Introduction:
 
 


 A great microscopist once reminded me that: “you can’t see 100% of the photons that your illumination does not provide” (as an insightful corollary to the famous Gretzky quote: “you miss 100% of the shots that you don’t take.”) Said another way,
 
 your illumination determines what you can and cannot see
 
 
 .
 
 For this reason, this post strives to start off as a practical resource for illumination by sharing one approach to making a second LED light source. My hope is to provide enough detail that newcomers can use this as a jumping off point and experienced microscopists can suggest improvements across the board.


 Solar (or room-light!) microscopy is very powerful, but sometimes it is just enough easier to document your amazing findings on a cell phone camera when you have a mobile light source. This is especially true when you want to gift your second microscope to someone you care about. The ideal outcome is that they are as hooked as you are so it helps if initially you can demonstrate the power of the tool relatively accessibly. One way to do this is by facilitating camera imaging by equipping them with the same tool you have in your kit — the battery powered LED.


 Another great aspect of having a LED is the window it opens to dark field type implementations which can sometimes help you pick out that extra feature of interest. See https://wp.me/p5uBMl-1PA


 In this post, I want to share a simple “tutorial” for making a second light source. I welcome comments from the community and hope that we — as a collective — can develop the ideal, DIY “second battery powered LED light source”.


 
 
 The parts you’ll need:
 
 


 For the following approach you need just a couple of components and tools


 <a alt="Picture11.png" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/Picture11.png">
 <img alt="Picture1" class="alignnone size-medium wp-image-13038 gcloudimg" data-attachment-id="13038" 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="Picture1" data-recalc-dims="1" height="300" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/Picture11.png" width="225"/>
 </a>

<ol>
 <li>
 foldscope back-mounted aperature (or cut your own from the scraps!)
 </li>
 <li>
 An LED (I used
 <a href="https://www.sparkfun.com/products/12621)">
 https://www.sparkfun.com/products/12621)
 </a>
 </li>
 <li>
 copper tape (
 <a alt="10561" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/10561">
 https://www.sparkfun.com/products/10561
 </a>
 )*
 </li>
 <li>
 a switch (
 <a alt="9609" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/9609">
 https://www.sparkfun.com/products/9609
 </a>
 )
 </li>
 <li>
 A battery and clamp (similar to:
 <a href="http://www.mouser.com/ProductDetail/Renata/SMTU2032-LFTR/?qs=sGAEpiMZZMvxTCYhU%252bW9mT9W%2ft5JZly7">
 http://www.mouser.com/ProductDetail/Renata/SMTU2032-LFTR/?qs=sGAEpiMZZMvxTCYhU%252bW9mT9W%2ft5JZly7
 </a>
 )*
 </li>
 <li>
 A soldering iron and solder*
 </li>
 <li>
 
 Optional: a dab of epoxy (to support the assembly against field use)
 
 </li>
 <li>
 
 Optional: a little double stick tape helped me to hold things in position
 
 </li>
</ol>

 (*Note: if you don’t have copper tape but do have some extra wire, I also had success with a end stripped wire and a dab of glue to hold them in place)


 (*Note2: We just had one lying around so I don’t know the precise supplier; anyone find something similar for a better price? Please share it if you do!)


 (*Note3: alternatives to soldering irons include:
 <a href="http://www.amazon.com/Bare-Conductive-Electric-Paint-10ml/dp/B00B888LQ8">
 http://www.amazon.com/Bare-Conductive-Electric-Paint-10ml/dp/B00B888LQ8
 </a>
 , but I have not personally tried this product)


 
 
 Assembly process:
 
 


 I recommend starting at the LED or the switch and ending with the orientation of a battery.


 My process started at the switch:

<figure aria-describedby="caption-attachment-13041" class="wp-caption alignnone" id="attachment_13041" style="width: 300px">
 <a alt="P_20160212_200022.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_200022.jpg">
 <img alt="Jpeg" class="size-medium wp-image-13041 gcloudimg" data-attachment-id="13041" data-comments-opened="1" data-image-meta='{"aperture":"2","credit":"","camera":"Z00AD","caption":"Jpeg","created_timestamp":"1455307222","copyright":"","focal_length":"3.78","iso":"180","shutter_speed":"0.0333","title":"Jpeg","orientation":"1"}' data-image-title="Jpeg" data-recalc-dims="1" height="225" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_200022.jpg" width="300"/>
 </a>
 <figcaption class="wp-caption-text" id="caption-attachment-13041">
 An image of my switch. Notice that the two pieces of copper tape are not in contact and that two of the three pins are soldered to the left and only one to the right.
 </figcaption>
</figure>

 Notice that for the switch, I put the copper tape down first being careful to make the two pieces close enough for the switch to reach both without them making contact on their own. Something that helped me was a double stick tape to hold the switch in place while I soldered it.


 I then soldered the LED in the center. At this point, I don’t need to think about the orientation of the LED leads (because I can flip the battery later), So I just focused on making sure I had the slight emitting side down (with leads up) and centered over the hole in the back foldscope mount.

<figure aria-describedby="caption-attachment-13042" class="wp-caption alignnone" id="attachment_13042" style="width: 300px">
 <a alt="P_20160212_200830.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_200830.jpg">
 <img alt="Jpeg" class="size-medium wp-image-13042 gcloudimg" data-attachment-id="13042" data-comments-opened="1" data-image-meta='{"aperture":"2","credit":"","camera":"Z00AD","caption":"Jpeg","created_timestamp":"1455307710","copyright":"","focal_length":"3.78","iso":"180","shutter_speed":"0.0166","title":"Jpeg","orientation":"6"}' data-image-title="Jpeg" data-recalc-dims="1" height="225" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_200830.jpg" width="300"/>
 </a>
 <figcaption class="wp-caption-text" id="caption-attachment-13042">
 The configuration post-LED soldering in the center and the addition of the second copper tape ‘L’. Notice that I had to add a bead of solder to make sure I had a reliable connection between copper tapes. Note that the adhesive of the copper tape is an effective insulator.
 </figcaption>
</figure>

 Next, I made certain that the switch was on and pushed my battery assembly down into place. When contact was made in the correct orientation, the LED illuminated. Following is an image of the illuminated LED, post soldering on of the battery. Notice that I pushed the battery as close the center as possible to make it a little less unwieldy on the back of the foldscope.


 <a alt="P_20160212_201148.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_201148.jpg">
 <img alt="Jpeg" class="size-medium wp-image-13043 gcloudimg" data-attachment-id="13043" data-comments-opened="1" data-image-meta='{"aperture":"2","credit":"","camera":"Z00AD","caption":"Jpeg","created_timestamp":"1455307907","copyright":"","focal_length":"3.78","iso":"150","shutter_speed":"0.0166","title":"Jpeg","orientation":"6"}' data-image-title="Jpeg" data-recalc-dims="1" height="225" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_201148.jpg" width="300"/>
 </a>


 The final assembly with LED illuminated.


 As a final step to help allow it to hold up longer, I added a couple of dabs of epoxy to the components (being careful not to glue my switch in the ‘off’ position!). I found this made the piece feel more put together, despite my ugly solder joints!

<figure aria-describedby="caption-attachment-13044" class="wp-caption alignnone" id="attachment_13044" style="width: 300px">
 <a alt="P_20160212_202046.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_202046.jpg">
 <img alt="Jpeg" class="size-medium wp-image-13044 gcloudimg" data-attachment-id="13044" data-comments-opened="1" data-image-meta='{"aperture":"2","credit":"","camera":"Z00AD","caption":"Jpeg","created_timestamp":"1455308445","copyright":"","focal_length":"3.78","iso":"190","shutter_speed":"0.0166","title":"Jpeg","orientation":"6"}' data-image-title="Jpeg" data-recalc-dims="1" height="225" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_202046.jpg" width="300"/>
 </a>
 <figcaption class="wp-caption-text" id="caption-attachment-13044">
 A little epoxy did the tick.
 </figcaption>
</figure>

 
 
 Now for the fun part: using the new light source.
 
 


 For those of you still with me, let’s see how the new source holds up. We’ll put it through its paces with emphasis on looking at the following couple of questions:

<ul>
 <li>
 Is this LED bright enough? Does it have nice white balance?
 </li>
 <li>
 Is this setup compatible with dark field?
 </li>
 <li>
 Are there any unexpected problems with the design?
 </li>
</ul>

 I imaged some old samples of Bat Star larvae because I now they are very feature rich. Let’s see if we can see the cilia along the edges of the organism (which you can see in darkfield with the included light source).


 First a collection of images in brighfield:

<figure aria-describedby="caption-attachment-13046" class="wp-caption aligncenter" id="attachment_13046" style="width: 300px">
 <a alt="P_20160212_224338.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_224338.jpg">
 <img alt="Jpeg" class="size-medium wp-image-13046 gcloudimg" data-attachment-id="13046" data-comments-opened="1" data-image-meta='{"aperture":"2","credit":"","camera":"Z00AD","caption":"Jpeg","created_timestamp":"1455317015","copyright":"","focal_length":"3.78","iso":"1450","shutter_speed":"0.0499","title":"Jpeg","orientation":"6"}' data-image-title="Jpeg" data-recalc-dims="1" height="225" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_224338.jpg" width="300"/>
 </a>
 <figcaption class="wp-caption-text" id="caption-attachment-13046">
 Full zoom out of the brightfield image of the larvae. Notice that I had a little trouble centering my magnetic coupler!
 </figcaption>
</figure>
<figure aria-describedby="caption-attachment-13047" class="wp-caption aligncenter" id="attachment_13047" style="width: 300px">
 <a alt="P_20160212_224607.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_224607.jpg">
 <img alt="Jpeg" class="size-medium wp-image-13047 gcloudimg" data-attachment-id="13047" data-comments-opened="1" data-image-meta='{"aperture":"2","credit":"","camera":"Z00AD","caption":"Jpeg","created_timestamp":"1455317166","copyright":"","focal_length":"3.78","iso":"370","shutter_speed":"0.0249","title":"Jpeg","orientation":"1"}' data-image-title="Jpeg" data-recalc-dims="1" height="225" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_224607.jpg" width="300"/>
 </a>
 <figcaption class="wp-caption-text" id="caption-attachment-13047">
 A look at the digestive tract of the larvae in brightfield.
 </figcaption>
</figure>
<figure aria-describedby="caption-attachment-13048" class="wp-caption aligncenter" id="attachment_13048" style="width: 300px">
 <a alt="P_20160212_224612.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_224612.jpg">
 <img alt="Jpeg" class="size-medium wp-image-13048 gcloudimg" data-attachment-id="13048" data-comments-opened="1" data-image-meta='{"aperture":"2","credit":"","camera":"Z00AD","caption":"Jpeg","created_timestamp":"1455317172","copyright":"","focal_length":"3.78","iso":"320","shutter_speed":"0.0249","title":"Jpeg","orientation":"1"}' data-image-title="Jpeg" data-recalc-dims="1" height="225" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_224612.jpg" width="300"/>
 </a>
 <figcaption class="wp-caption-text" id="caption-attachment-13048">
 Lower focal plane of ciliary band. Brightfield.
 </figcaption>
</figure>
<figure aria-describedby="caption-attachment-13049" class="wp-caption aligncenter" id="attachment_13049" style="width: 300px">
 <a alt="P_20160212_224615.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_224615.jpg">
 <img alt="Jpeg" class="size-medium wp-image-13049 gcloudimg" data-attachment-id="13049" data-comments-opened="1" data-image-meta='{"aperture":"2","credit":"","camera":"Z00AD","caption":"Jpeg","created_timestamp":"1455317175","copyright":"","focal_length":"3.78","iso":"320","shutter_speed":"0.0249","title":"Jpeg","orientation":"1"}' data-image-title="Jpeg" data-recalc-dims="1" height="225" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_224615.jpg" width="300"/>
 </a>
 <figcaption class="wp-caption-text" id="caption-attachment-13049">
 Upper focal plane of ciliary band, brightfield.
 </figcaption>
</figure>

 Now let’s move on to dark field implemented like the microcosmos dark field post. (
 <a alt="p5uBMl-1PA" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/p5uBMl-1PA">
 https://wp.me/p5uBMl-1PA
 </a>
 )

<figure aria-describedby="caption-attachment-13045" class="wp-caption aligncenter" id="attachment_13045" style="width: 300px">
 <a alt="P_20160212_224325.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_224325.jpg">
 <img alt="Jpeg" class="size-medium wp-image-13045 gcloudimg" data-attachment-id="13045" data-comments-opened="1" data-image-meta='{"aperture":"2","credit":"","camera":"Z00AD","caption":"Jpeg","created_timestamp":"1455317004","copyright":"","focal_length":"3.78","iso":"400","shutter_speed":"0.0249","title":"Jpeg","orientation":"6"}' data-image-title="Jpeg" data-recalc-dims="1" height="225" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/P_20160212_224325.jpg" width="300"/>
 </a>
 <figcaption class="wp-caption-text" id="caption-attachment-13045">
 Full zoom out of the same organism as above. Notice the difference between this darkfield image and the brightfield one above. You can see the bands really well. And look how much that gut scatters.
 </figcaption>
</figure>

 We can also see individual cilia in darkfield with our DIY LED setup.


 <a alt="Picture2.png" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/Picture2.png">
 <img alt="Picture2" class="size-medium wp-image-13054 aligncenter gcloudimg" data-attachment-id="13054" 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="Picture2" data-recalc-dims="1" height="213" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_13036/Picture2.png" width="300"/>
 </a>


 
 
 Wishing all “micro-cosmonauts” happy exploration!

Practical Resource &#8211; making a DIY, LED light source

The ocean holds remarkable wonders from behemoth whales to iron-“breathing” bacteria ( See: http://www.sciencedaily.com/releases/2009/04/090416144512.htm).  You might even call some of them “alien” to emphasize just how different from our expectations some of them can be.


 Being granted the wonderful opportunity to spend some time at a local Marine Station, I set out — with foldscope in hand — to try to find and share one of these “aliens” with the microcosmos community.


 As micro-enthusiasts, we all know sometimes you needn’t look far — often peering into the micro is enough. Mindful of this approach, and inspired by some wonderful work at the ocean’s surface (any one interested in the ocean should look over this project! http://www.embl.de/tara-oceans/start/), I found some friends knowledgeable about plankton and swimming larvae.


 Watch this “alien” taken on a 140x foldsope (in a special “dark-field” mode which is detailed here : http://microcosmos.foldscope.com/2015/06/24/simple-dark-field-improve-your-imaging-contrast-with-another-classic-microscopy-technique/)

<div class="jetpack-video-wrapper">
 
 <iframe allowfullscreen="true" class="youtube-player" height="563" src="https://www.youtube.com/embed/C_DNJlT5Hh4?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">
 </iframe>
 
</div>

 Look at that beautiful “swallow”! Can you see the circulation of particles around the head and mouth? Lots of great structure to see in this short video. Enjoy.


 This is a great example of some of the astonishing things you can find in the ocean. Remarkably, these bilaterally symmetric larvae ultimately turn into an adult starfish (
 
 pateria miniata)
 
 with 5 appendages. (http://www.cokesmithphototravel.com/coastal-california.html)


 <a alt="37180188_scaled_416x374.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_6980/37180188_scaled_416x374.jpg">
 <img alt="37180188_scaled_416x374" class="alignnone wp-image-7158 gcloudimg" data-attachment-id="7158" 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="37180188_scaled_416x374" data-recalc-dims="1" height="270" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_6980/37180188_scaled_416x374.jpg" width="300"/>
 </a>




 Learn more about the metamorphosis by looking at http://oceandevbio2013.blogspot.com/2013/05/species-patiria-miniata-phylum.html.




 These “aliens” are just one of many in the ocean. Keep on exploring.

Microscopic &#8220;aliens&#8221; of the ocean!

Introduction and motivation:
 


 Dark- field microscopy is an essential microscope technique which allows you to see small features with high-contrast. This technique is named for its appearance — the background looks darker than the features you are imaging. This is in contrast with the technique of “bright field” in which the background is unscattered light straight from the light source (your foldscope is equipped to do “brightfield” upon arrival). Often, looking at the same sample in dark-field can give you a very different picture of the same sample (examples to come). Both techniques are essential for the explorers toolkit.


 
 Goal:
 


 Here, we instead present a simple and easily adjustable modification that should allow you to get simple dark field. This post is in part intended to encourage users to play around with the instrument and try to engineer better solutions and new imaging modes. The hope is this may be an accessible and rewarding starting point for those enthusiastic about improving the capabilities of their foldscope through simple (and reversible!) customization.


 
 Quick (and simple) optics
 


 The optics of dark-field are simple. The idea is to illuminate your sample such that only scattered light makes it into the objective. Practically, this means removing all of the rays of light (or spatial frequencies) which are collected by the objective lens. Forms of dark field can be achieved simply by illuminating the sample from an extreme angle. (To test how well this works, you might try adjusting the position of your LED relative to center of your objective lens).  In this post, we review a symmetric form of off-angle illumination using the included condenser lens (the ball lens with the bigger aperture), the light source module, a clear plastic (or glass) piece square, double-stick tape and a sharpie (or equivalent) marker.


 A sketch of the optics should look something light this.

<figure aria-describedby="caption-attachment-7043" class="wp-caption alignnone" id="attachment_7043" style="width: 500px">
 <a alt="Journal-22.png" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_7042/Journal-22.png">
 <img alt="Light travels from the bottom to the top (light source in yellow). The added component is the red box and represents an annular aperture between the light source and the condenser lens. When the central rays are completely blocked the background of the image should be dark (because yellow rays do not make it out of the objective lens). However, the scattered light -- signified by orange light -- can create new rays which couple into the objective. Therefore you only see scattered light from the sample." class="wp-image-7043 gcloudimg" data-attachment-id="7043" 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="Simple darkfield with condenser" data-recalc-dims="1" height="375" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_7042/Journal-22.png" width="500"/>
 </a>
 <figcaption class="wp-caption-text" id="caption-attachment-7043">
 Light travels from the bottom to the top (light source in yellow). The added component is the red box and represents an annular aperture between the light source and the condenser lens. When the central rays are completely blocked the background of the image should be dark (because yellow rays do not make it out of the objective lens). However, the scattered light — signified by orange light — can create new rays which couple into the objective. Therefore you only see scattered light from the sample.
 </figcaption>
</figure>

 Note in the quick sketch above, the unscattered light is yellow and the scattered light is orange-ish. Only scattered light is picked up by the objective and the rest is lost onto the aperture (the black plastic piece holding the lens).


 
 Methods and protocol:
 


 (1) Take your assembled scope for bright field (using the battery light module) and remove the back panel as follows. Make sure you have double stick tape and a condenser lens (should have come with the kit).


 <a alt="Photo-Jun-16-12-46-47-AM.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_7042/Photo-Jun-16-12-46-47-AM.jpg">
 <img alt="Photo Jun 16, 12 46 47 AM" class="alignnone size-medium wp-image-7044 gcloudimg" data-attachment-id="7044" 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="Photo Jun 16, 12 46 47 AM" data-recalc-dims="1" height="300" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_7042/Photo-Jun-16-12-46-47-AM.jpg" width="300"/>
 </a>


 (2) Acquire a piece of clear plastic (I started with a 170um glass coverslip, but transitioned to a cut-out of a transparency) or put multiple pieces of clear tape together to give a nice optically clear window.


 (3) tape the window onto the back of the condenser lens (opposite side of the smaller hole; the flat side). Double stick tape worked well for me.


 (4) use a sharpie to make a dark spot about the size of 2/3 of the visible ball lens. It helps to go over it a couple times to make it as optically opaque as possible, Mine looks something like this: Note that each time you go over the sample you add some of the ink but also more of the solvent so the final coloring should be to redistribute the wet ink as equally as possible. What worked for me was “dotting” in multiple locations instead of “smearing”.



<figure aria-describedby="caption-attachment-7149" class="wp-caption alignnone" id="attachment_7149" style="width: 300px">
 <a alt="2015-06-24-13.22.25.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_7042/2015-06-24-13.22.25.jpg">
 <img alt="2015-06-24 13.22.25" class="wp-image-7149 size-medium gcloudimg" data-attachment-id="7149" 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="2015-06-24 13.22.25" data-recalc-dims="1" height="300" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_7042/2015-06-24-13.22.25.jpg" width="300"/>
 </a>
 <figcaption class="wp-caption-text" id="caption-attachment-7149">
 Clear plastic piece with sharpie dark-out in center next to the condenser lens with double-stick tape across the top.
 </figcaption>
</figure>

 (5) Mount the condenser on the foldscope and without a sample hold the instrument up to light in “solar” mode.  You should see almost no light getting through. If you see light make your spot either darker or slightly bigger to block the rays making it through the optical system shown in the schematic above.


 (5b) **Optional** Add a scattering piece of paper between the condenser and the lightsource. Toilet paper can work well here if you take a single layer of it. This can help improve the uniformity of your illumination but also contributes to the loss of light. If you have intensity to spare and nonuniform illumination, this addition might help.


 (6) Assemble your foldscope and put a sample in place. Look for strong contrast at edges in particular where you have a lot of scattering surface. Adjust the alignment of the spot, the condenser and the source until you see a nice dark blue background with a scattering body from the sample. Examples are following.


 All images are collected on an iphone 4s using the magnetic coupling.


 
 Results and some images of fruit fly
 
 
 embryos
 


 In what follows, I demonstrate the difference between the bright-field and dark-field imaging capabilities on fruit fly embryos. After sharing a sense for the differences we will consider a sample where dark-field gives dramatically different information that bright-field: that of  the live imaging of a near transparent starfish larvae.


 Fruit fly development is well studied. Here we look at late-stage embryos developing into the larval form. The goal being to see if we can resolve differences in the images.

<figure aria-describedby="caption-attachment-7154" class="wp-caption alignnone" id="attachment_7154" style="width: 365px">
 <a alt="DrosphilaEmbryo.jpg" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_7042/DrosphilaEmbryo.jpg">
 <img alt="Left: Dark-Field image Right: Bright-field image" class="wp-image-7154 gcloudimg" data-attachment-id="7154" 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="Fruit Fly Embryo" data-recalc-dims="1" height="262" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_7042/DrosphilaEmbryo.jpg" width="365"/>
 </a>
 <figcaption class="wp-caption-text" id="caption-attachment-7154">
 Left: Dark-Field image
 
 Right: Bright-field image
 </figcaption>
</figure>

 Along the left side, both images show the emergence of some ridging into what ultimately will define the segments of the fly’s body. There also appears to be some internal structure visible in the dark-field image. See if you can use this video (https://www.youtube.com/watch?v=FChS4KU5jDM) to identify how old these embryos are.


 
 Final example: Dark field of transparent larvae
 


 A clear situation in which dark-field should help is in the imaging of a transparent larvae. In the following video, we compare the imaging of a bat star larvae in both illumination methods. This demonstrates clearly a situation in which dark-field helps you understand better what is going on.

<div class="wp-video" style="width: 640px;">
 
 <video alt="Fruit Fly Embryo" class="wp-video-shortcode gcloudimg" controls="controls" height="360" id="video-7042-1" preload="metadata" width="640">
 <source alt="Combined-Stacks-desktop.m4v" class="gcloudimg" src="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_7042/Combined-Stacks-desktop.m4v" type="video/mp4"/>
 <a alt="Combined-Stacks-desktop.m4v" class="gcloudimg" href="https://storage.googleapis.com/microcosmosdelta.appspot.com/images/transfer_7042/Combined-Stacks-desktop.m4v">
 https://microcosmos.foldscope.com/wp-content/uploads/old/Combined-Stacks-desktop.m4v
 </a>
 </video>
</div>

 While this dark-field implementation does not yet match the performance of table-top scopes, it provides a valuable channel of information none-the-less. Notice the clear improvement in the ability to resolve the internal structures as well as the outline. We also see glimmers of the characteristic feature of dark-field: bright edges with large scatter.


 
 Ways to improve – show us your method
 


 There are many ways to improve on this implementation. You can begin by optimizing the size of your light block to reduce the collection of unscattered light. Also note that sharpie does not fully block the transmission and thus gives a bit of a blue background. Using an optically opaque paper should improve this.  Also we suffer from loss of too much light as the block size increases. One way to counter-act this may be to (a) increase brightness of the light-source, and (b) redirect instead of blocking the used light into a single parallel line of rays.


 Share your implementations with the community!


 
 Conclusion
 


 We have set out to demonstrate the addition of a single optical component to make a dark-field foldscope with the simplest possible modification. Many alternative methods for achieving the same imaging mode can be made each with their own advantages. This method is simple, easy to align and reasonably effective. Further it allows foldscope users to reversibly modify their components and is therefore safe for all users to try. We hope dark-field microscopy becomes a readily available tool and that future implementation can begin to match the performance of table-top microscopes.

Matthew Bull

Earth

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