Chloroplast movement in Elodea (a form of pond weed)

Cellular Turbulence. Rhys and I went with the family to the Big Bang Show at the NEC in Birmingham. On the Zeiss stand were a number of microscopes – and on one of them some Elodea showing chloroplast movement around the cell. This pond weed has particularly mobile chloroplasts and the site is amazing. This movement is referred to as cyclosis or cytoplasmic streaming.

See the photo and video below – wow! I wonder what the small things are, much smaller than chloroplasts? Organelles or parasitic protozoa or bacteria? Magnifications here using x32 and x63 objectives so bacteria would show up at this magnification.

Andy

Some facts about chloroplasts:

Chloroplasts are organelles, specialized compartments, in plant and algal cells. The main role of chloroplasts is to conduct photosynthesis, where the photosynthetic pigment chlorophyll captures the energy from sunlight and converts it and stores it in the energy-storage molecules ATP and NADPH while freeing oxygen from water. They then use the ATP and NADPH to make organic molecules from carbon dioxide in a process known as the Calvin cycle. Chloroplasts carry out a number of other functions, including fatty acid synthesis, much amino acid synthesis, and the immune response in plants. The number of chloroplasts per cell varies from one, in unicellular algae, up to 100 in plants like Arabidopsis and wheat (from https://en.wikipedia.org/wiki/Chloroplast)

The average chloroplast is about 3 µm (micrometers) in diameter. In one square millimeter of the surface of a leaf, there are about half a million chloroplasts (from www.answers.com/Q/What_is_the_size_of_a_chloroplast)

Chloroplasts in vascular plants range from being football to lens shaped and as shown in Figure 1, have a characteristic diameter of ≈4-6 microns (BNID 104982, 107012), with a mean volume of ≈20 μm3 (for corn seedling, BNID 106536). In algae they can also be cup-shaped, tubular or even form elaborate networks, paralleling the morphological diversity found in mitochondria. Though chloroplasts are many times larger than most bacteria, in their composition they can be much more homogenous, as required by their functional role which centers on carbon fixation. The interior of a chloroplast is made up of stacks of membranes, in some ways analogous to the membranes seen in the rod cells found in the visual systems of mammals. The many membranes that make up a chloroplast are fully packed with the apparatus of light capture, photosystems and related complexes.  The rest of the organelle is packed almost fully with one dominant protein species, namely, Rubisco, the protein serving to fix CO2 in the carbon fixation cycle. The catalysis of this carbon-fixation reaction is relatively slow thus necessitating such high protein abundances (from http://book.bionumbers.org/how-large-are-chloroplasts/)

Components of cells seen in photos and video:

   

Today’s photos and video:

If you want the wow factor go straight to the videos at bottom of page using x63 objective! I am quite excited by the views with the x63 objective below as this is first time I have used it so successfully with this microscope. The slide is turned upside down and Kohler illumination has been achieved using my “new” (second hand off ebay) NA 0.9 bright field condenser with Zeiss 475638 illuminator collimation tube (at least I think that is what it is for!).

The slide was prepared using free hand sections of Elodea leaf using razor blade put on slide with drop water and covered with cover slip. Edges of cover slip help firmly to slide by electrical insulating tape strips and then slide turned upside down and put on microscope stage (upside down as Zeiss IM microscope is an inverted microscope).

Photo of Elodea leaf section (cut free hand with razor blade) x32 objective, bright field, Zeiss IM microscope, showing cell walls and chloroplasts:

Photos of Elodea x63 objective bright field, now also shows small inclusions much smaller than chloroplasts – in later videos these are shown to move as well as chloroplasts:

Videos – first two videos are with x32 objective, bright field:

Videos – next videos used x63 objective, bright field:

Solar Image processing. Myths and Legends.

I have just read an (unattributed) article in April’s Sky at Night magazine on solar imaging, and I have to say, from my own experience I disagree with a lot of it!

It says

“— requires a monochrome high frame rate camera set-up” and “use of a colour camera is inefficient”

Who cares? There is plenty of light from the Sun, efficiency isn’t a problem!

It also says that Ha features change quite quickly. True. As do the atmospheric “cells” that cause image wobble. It suggests you take a 1000-1500 frame avi. The time this takes immediately cancels any advantage there might be from a high frame rate. When you stack all these, you get a blurred image. The only reason you would take so many frames is to reduce noise. Again there is plenty of light from the Sun, so this isn’t an issue.

It suggests that you might need a flat frame (possible) and that you take a defocussed 500-1000 frame avi to achieve this. Why? It is much easier and more accurate if you need a flat frame to simply blur an image you have already acquired.

My images use a £50 colour camera with a not particularly high frame rate. I find a good compromise is 200 frames. This takes around 7 seconds.

Click on “Solar” on the blog and judge for yourselves!

While still in Victor Meldrew  mode, in the same magazine there is a review of a new Skywatcher 20” goto dob for £5499. I am sure that this is a splendid scope, but following my earlier post it is worth remembering that it is only 1 stop faster than Rob’s new 14”! I am pretty sure Rob didn’t spend that amount on it! In fact, in the review there are pictures of M42 and the Trapezium. There is also a picture of M51 of recent discussion. They look nice, but I would invite you to compare the pics with these window-sill images with a scope costing £100 ish.

Moral – Just because something is in print does not necessarily mean it is correct. This is a hobby, it is whatever floats your boat. You can spend a fortune if that is what you want to do, but you don’t HAVE to!

Kohler illumination on Zeiss IM microscope using Carl Zeiss 0.9 NA Swing Flip Top Condenser, Zeiss 475638 extension tube and Zeiss condenser diaphragm

My Zeiss IM microscope came set up for phase contrast. Although phase contrast is an amazing technique, the condenser was limited in its ability to be used to set up Kohler illumination.

Kohler illumination is a method of illumination of microscopic objects in which the image of the light source is focused on the substage condenser diaphragm and the diaphragm of the light source is focused in the same plane with the object to be observed; maximizes both the brightness and uniformity of the illuminated field (http://medical-dictionary.thefreedictionary.com/Kohler+illumination)

To achieve Kohler illumination with the Zeiss IM or IM35 microscopes, I needed to obtain one of Zeiss’ bright light condensers. The manual showed the microscope in use with a flip top condenser so I purchased one of these from ebay together with the extension tube and condenser diaphragm shown in the manual – had to wait a bit until one was available.

Success! I can now focus the diaphragm edge in the same plane as the image of the slide, improving illumination and contrast to the maximum available for the microscope…..at least in theory – and seems to work today when I tried it.

Andy

Carl Zeiss 0.9 NA Swing Flip Top Condenser (below). With this arrangement, I am able to open the diaphragm up to the full field of view:

Condenser mounted on microscope – both with flip top lens in and out (below):

Image of condenser diaphragm edge shown against the background slide – I have closed down the diaphragm somewhat to show the edge (below):

I also tried mounting one of my other Zeiss condensers. This one has bright field, phase and dark field options. It achieved focus although the condenser had to be much closer to the slider, but, even when fully open, the diaphragm could not illuminate the whole field evenly (below):

Second light with 14 inch dob

Just heading to bed last night when I spotted the skies had cleared…

– Ken’s 31mm EP gave beautiful views! Was having too much fun to try the other one.

– Could definitely see the spiral arms on M51 but not M81. Very similar to the single unprocessed sub that Roger posted

– Spent lots of time on Leo galaxies

Skies were improving but it was past midnight… Damn you work!!

Rob

Observing report: 20/03/18 19:15

Clearing skies approaching sunset held the promise of Venus and Mercury, as well as a young crescent moon. 19:15 pm went out to front of house armed with 7×50 binoculars, Young crescent moon, with earthshine visible at about 30 deg in the WSW, Venus also visible with naked eye ,near enough W and low down at 5 deg , grazing the chimney pots and playing hide and seek through low cloud bank to the west. No sign of mercury visually, but with binoculars could be located a couple of degrees to right and above Venus, even when I knew where it was , still couldn’t discern it visually, 5 mins later cloud bank hid Venus visually, could still pick it out with binoculars and mercury was getting lost in a bank of higher wispy cloud.

Worth looking out for if clear at sunset.

 

Pete H

Some Musings on Aperture and DSOs

Rob’s recent post on his new scope led me to reflect (!) on my own experiences. My main interest is deep-sky objects, or “faint fuzzies”. I have a rule of thumb that says that to see any appreciable difference in object  brightness, you have to go up 2 stops in focal ratio (f-numbers). This translates to a doubling in aperture (doubling the aperture increases the light grasp by 4X). So when I upgraded from my old 4”, I went to 8”. Doing the same again would suggest a 16” would be needed. For me, a 16” would be unmanageable, so I went to a 12”. This is only 1 stop advantage to the 8”, and visual results were a bit disappointing. This more-or-less coincided with getting the PD camera, and even using this “live” as an electronic eyepiece gave views so superior to the 12” that the 12” virtually never got used.

So I wondered why this was, so here is a bit of basic physics.

Firstly, the eyeball. This iris opens to about 7mm when fully adapted to the dark. Or maybe 5mm in an old fogey like me. Given the size of the eyeball this is around f/3. See http://www.faculty.virginia.edu/rwoclass/astr1230/human-eye.html

To get the brightest image from a scope, the magnification from the scope must produce an exit   pupil (the beam width leaving the eyepiece) pupil less than 5-7 mm. Otherwise some of the light leaving the scope doesn’t enter the eye and is wasted. This defines the minimum magnification you can use for a given aperture. For example with the 8” and a 5mm exit pupil, this minimum magnification is 200/5 = X40. (The SCT has a focal length of 2000mm, so this is an eyepiece of 50mm FL). A lower magnification than this is not harmful, it is just that you are then not making use of all the available aperture. 7X50 binoculars are known as “night glasses” for exactly this reason – their exit pupil is 7mm, making the best use of all night-time light. Therefore, going up in aperture does not necessarily make the view brighter, but rather allows you to us a higher magnification for the same brightness. This might be a huge advantage for small objects like planetary nebulae, but less so for extended objects. Bigger aperture also improves resolution, allowing you to split closer binaries, but this is usually not the critical issue for faint fuzzies. The other issues affecting brightness are the eye sensitivity (more of this later) and its “integration time”, or the time period over which the eye sums the image it sees. This is its “shutter speed” and there is some literature that suggests in the dark, this is about 0.2 seconds. Again. see http://www.faculty.virginia.edu/rwoclass/astr1230/human-eye.html

Now, from here, I have NEVER conclusively seen any galactic spiral arms visually, although sometimes I have persuaded myself I can. So when I first coupled up the PD camera to the 8”, and turned it on M51, this is what I saw, live, with no processing at all:

Bingo! Spiral arms!

So if we are now talking about imaging, rather than the eyeball, what comes into play for image brightness?

3 things.

  • Focal ratio (not aperture)
  • Integration time. The PD single image is 1/50 second. Its “senseup” parameter allows it to internally stack up to 1024 single images, giving an integration time of about 20 seconds, or about 100 times the eyeball.
  • If the CCD had the same sensitivity as the eye, the brightness of the CCD image would be the same as the eye if a senseup of 4 were used. An experiment is called for!. I attached a lens to the camera, set it to around f/3 (similar to the eye), then in a darkened room compared the image from it to that I could see with my eye for various senseups. Although this is a very crude experiment, I reckoned that about senseup=2 was about right-not miles away from the predicted value. So a senseup of 1024 suggests a sensitivity about 8 stops faster than the eyeball.

There is also another factor involved, and that is contrast. This is the brightness of the faint fuzzy compared to the sky “background”, and I have found that with our local skies, that is the controlling factor. You can improve this visually using filters (UHC or OIII for example). These tend to dim the whole view, but the right one can improve contrast. Light pollution filters used to be good in the days of low-pressure sodium street lighting but are not much use with LED lights. On the other hand, the camera has a “gamma” setting that allows you to “stretch” the contrast, live, or if you post-process, the sky is the limit, as they say! For example, stacked 11 of the basic frames of M51 (11264 frames in total), processed it with GIMP, and here is the result. It is flipped vertically to get the orientation right (See http://www.thornett.net/Rosliston/Astrophotography/DSO.pdf for the details). Stacking 11 frames with a senseup of 1024 gives another 3 ½ stops faster than the eyeball or 11 or so altogether.

Interesting as all this might be, let’s remember that this is a hobby – and you do whatever you enjoy!

As a final thought, Lord Rosse used a 72″ aperture reflector to first identify the spiral nature of M51!

(Slightly delayed) First Light report on 14 Inch Dob

The American humourist Will Rogers once observed that there are 3 kinds of men: Those who learn by reading, the few who learn by observation and the rest who have to pee on the electric fence for themselves. I’m joining the third category, as, against the advice of just about everyone I’ve discussed it with I’ve decided to get myself a large(ish) Dob.

Although in the last nine months or so, I’ve become interested in Astrophotography I also really enjoy visual observing, and especially hunting for objects. When I saw a 14 inch’er on Astrobuysell that was relatively portable there was only so long I could avoid temptation. Gotta have something to do whilst taking subs…

I’ve managed one short session between the clouds last week, which was a good reminder of the tribulations of getting to know a new scope but also a promise of fun to come. I could not find anywhere convenient to mount my Quickfinder- I put it too close to the eyepiece and managed at one point to head-butt it clean onto the grass. I also found that the 35mm Eyepiece that came with the scope gives truly horrible views (it may have a future career as a paperweight) and that the Azimuth adjustment is pretty sticky- especially near the zenith. All of these things are going to need some sorting. Attempts to observe M42 and M31 were both scuppered by banks of cloud rolling it at the wrong moment, but just as the frustration levels were rising I got M81 in the eyepiece and saw for the first time with my own eyes detail beyond the galaxy core. Next up was M51 and here I could see both cores quite clearly and some of the material that joins them. In five minutes I had swung from irritation to elation and with the clouds now rolling in I went for the Leo triplet, something I just haven’t been able to see from my location before. Just in time I found them- no detail, but the shapes quite easy to see even without averted vision. That was pretty much it, as the clouds rolled over and haven’t really parted since, but enough that I’m very excited about the next clear night…

PS- I’d like to apologise to everyone for invoking “The Curse of the New Scope” and ruining the weather for a few weeks.