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Esplosivo
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I checked the fish right now. I could not see any clear luminous colonies, but on the part of the fish which was left outside the water I could see a
very faint light. I am not sure that these could be the bacteria. So what I will do is that tomorrow I will buy the necessary chemicals and try
innoculating a medium with all the bacteria present on the surface of the fish as mentioned previously.
If I manage to collect just a couple of luminescent bacteria a colony will form and from this colony I could make millions of others.
Btw, I tried taking a picture with my cheap camera but nothing was visible. The camera has no aperture settings so the ammount of light entering the
cam cannot be controled. Anyway, I will try enhancing the pic.
Edit: The growth of the bacteria will be postponed since I cannot obtain the yeast / beef extract till next month. Till then I will try acquiring two
different species of fish, and probably even a squid or octopus.
[Edited on 11-8-2004 by Esplosivo]
Theory guides, experiment decides.
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gorkem
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lightning bacteria
we want to produce lightining bacteria in the college environment but we fail.do you know how to do it?
You might want to read this very thread first. Chemoleo
[Edited on 21-12-2004 by chemoleo]
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vulture
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Read the thread first and DO NOT crossposts three times!!
One more stunt like this and your posting ability will be gone with the wind.
One shouldn't accept or resort to the mutilation of science to appease the mentally impaired.
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Madandcrazy
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Vulture you meant the crosspost to chemolos metabolic pathway topic ?
I don`t know the method isolating lightning substances from insects would walk ?
(wings of chafers or some tropic insects)
[Edited on 10-7-2005 by Madandcrazy]
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Ravenstein
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Help!
Hi all, I am from Singapore and and trying to do this ... I bought some octopus and and letting them rot in my fridgerator half submerged in sea
water. Its been nearly 18 hrs since i started. At first i did'nt read the things well and left my octopus outside for about 12 hrs and earlier
this morning, i placed them in my fridgerator after i realised what happened. Will this affect the photobacterium?
For the tryptone, i am in early high school and am not very sure how to obtain this, early in this thread someone said any protein extract should
work... is it true? and if not, what should i use.
For the medium can i use the LM and just dont add agarose gel?
How fast does photobacterium grow?
How long will it take to get about 100ml of the bacteria?
How do i grow the bacteria in such a way i can get alot of it quickly?
In singapore, anyone know where i can buy petri dishes and the rest of the ingridents?
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Bugger
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I've got a question:
I live in an inland state in a town where there's no fresh sea-fish available; everything's either frozen or already cut up and probably
washed and cleaned. Is there no other way I could get a sample of this glowing bacteria from nature then? Would aquarium water from a tank w/ live
rock contain the bacteria, for instance?
Thanks for any help!
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sparkgap
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OK, let's review what has been said thus far...
-P. phosphoreum is a *marine* organism.
-Freshly-caught marine animals *may* carry the microbe. Store-bought animals may have already been cleaned and as such may have little, if any,
microbe present.
-Yeast lysate and tryptone are possible protein sources. (There should be bakeries nearby that have yeast, FCOL).
-Use gelatin or agar as a medium. Shouldn't be difficult if you live in a tropical country... they're extensively used in a good number of
Oriental deserts.
sparky (~_~)
"What's UTFSE? I keep hearing about it, but I can't be arsed to search for the answer..."
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Xque
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You might want to look into Pseudomonia ssp. too. There's several species which are flourescent when lacking iron, such as P.
aeruginosa, P. flourescens , P. putida and P. syringiae. It seems Pseudomonia is pretty common.
I wouldn't mess too much with aeruginosa though, as it is a pathogen.
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nitro-genes
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That's funny We did exactly the same experiment when I was a student. This was
for a course in microbial ecology. We used "fresh fish" from the market nearby, but I've heard that fish from the market can be as old as two weeks
Maybe that was the reason that only one group actually got V. Phosphoreum. Our group wasn't able to spot any luminous colonies on the fish,
so obtaining the right fish is the hard part I think.
We incubated the fish at 4 deg. C to deminish competition with other microorganisms. (V. phosphoreum can grow quite fast at 4 deg. C.) And
looked for colonies twice a day...
Quite funny was that our professor told us that every now and then reports come in from terrified people that claim to have eaten "radiation
contaminated" seafood.  The bacterium however is of no harm IIRC.
I wonder what the reason is for it's dissapearance after the fish gets older. V. phosphoreum is facultative anaeroob and able to grow at a
wide range in temperature. Maybe because of competition with other faster growing microorganisms?
Oh, and an idea for people that need yeast extract...There are these nutritional yeast pills that are told to be rich in Vitamin B. I believe they are
mayde of pressed and dried cells of S. cerevisiae siccum 
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Esplosivo
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That's nice to hear. So you did manage to grow colonies on the fish. I have tried with two freshly caught fish (caught by a friend that very moment
and kept in sea water - fresher than that is impossible) but with no apparent success, except on the second time where I think there were visible
light emitting colonies which were very faint.
Could you please give us more detail regarding this experiment. You said you incubated the fish at 4degC - for how long? When did the first colonies
become visible? Did you incubate the fish in sea water? Do you have any idea of what species of fish did the bacteria actually grow on (the common
name is sufficient)? Thank you.
[Edited on 12-1-2006 by Esplosivo]
Theory guides, experiment decides.
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nitro-genes
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I'm sorry if I made the impression that luminous bacteria are specific for some species of fish... What I meant was that you need a fish with the
"right" fressnes. I/m saying "right" because I haven no clue why the colonies show up in some cases as in others it doesn't. Could be a lot of things,
but In our experiment we used plain cod. The group that had succes found colonies after less than two days @ 4 deg C already.
The light is only emmited during the growth fase, so you should be able to see the luminescence quite quickly. That is probably also the reason why
you must have very fresh fish.The bacteria will still be there, but not emmitting any light anymore!
Luminous bacteria are present in almost all seas at any depth and can be found in almost every marine organism. Of course there are more species of
luminous bacteria, so I don't know if this is the case for V. phosphoreum. But seen it's large temperature range in which it can grow, and
the commenly found niche it occupies, it is quite likely.
In the attachment is a pdf that is very simple, it contains almost every information you need, like media, growth conditions and background
information.
[Edited on 12-1-2006 by nitro-genes]
Attachment: Luminous bacteria.pdf (186kB)
This file has been downloaded 2294 times
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Vitus_Verdegast
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TCBS agar preparation
Thiosulphate Citrate Bile Salts Agar (TCBS)
(Oxoid designation is Cholera Medium TCBS)
Basic ingredients
Peptone 10,0 g
Yeast extract 5,0 g
Sodium citrate 10,0 g
Sodium thiosulfate.5 H2O (Na2S2O3•5 H2O) 10,0 g
Sodium chloride (NaCl) 10,0 g
Ox gall powder 5,0 g
Iron (III) citrate (FeC6H5O7) 1,0 g
Sodium cholate 3,0 g
Sucrose 20,0 g
Thymol Blue solution (0,2%) 20,0 ml
Bromothymol Blue solution (0,2%) 20,0 ml
Agar 15,0 g
Distilled water 1,0 L
pH 8,6 ± 0,2
Preparation
Mix all ingredients and apply heat until all goes into solution. DO NOT STERILIZE USING AUTOCLAVE. Allow to cool to 45-50°C and distribute.
Notes:
from http://www.disknet.com/indiana_biolab/b041.htm:
Peptones are the most widely used source of nitrogen in microbial media. Some are made by cooking milk or meat products in acid, but most are made by
incubating milk or meat with trypsin, pepsin, or other proteolytic enzymes to digest the protein to a mixture of amino acids, peptides, and
polypeptides. Many microbes, called proteolytic, can digest proteins, but most can't. The choice of peptone is sometimes of importance.
Some water-based inks contain ox gall (bile) as a dispersant. Pure ox gall can be obtained from art supply stores (eg. Winsor & Newton brand).
Sodium cholate, a bile salt, is used as an anionic detergent.
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kazaa81
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Maybe this is a little off-topic, but I would mention some dinoflagellates of the genre Noctiluca, that, when they're shaken, emits light.
Obviously, being monocellular organism of larger size, it would be more difficult to spot and grow as done with Photobacterium; it has been reported
of some species that can reach 2mm of diameter...that's very big for a monocellular organism!
bioluminescent dinoflagellates at luxgene.com
http://www.bioart.co.uk/lux/dino.html
"Noctiluca" on wikipedia
http://en.wikipedia.org/wiki/Noctiluca
Noctiluca scintillans information pages
http://www.marbot.gu.se/SSS/dinoflagellates/Noctiluca_scinti...
http://www.nmnh.si.edu/botany/projects/dinoflag/Taxa/Nscinti...
Noctiluca scintillans photo
http://micro.magnet.fsu.edu/featuredmicroscopist/vanegmond/i...
http://micro.magnet.fsu.edu/featuredmicroscopist/vanegmond/i...
http://members.fortunecity.com/anemaw/noctiluca.jpg
The Subcellular Origin of Bioluminescence in Noctiluca miliaris
http://www.jgp.org/cgi/content/abstract/50/5/1429
[Edited on 9-6-2006 by kazaa81]
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JohnWW
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What enzymes, and substrate, are involved in the chemoluminescence reactions? In the cases of fireflies and cave glow-worms, I understand it is
luciferase and luciferin.
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bereal511
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Hmmm, I wonder if anyone was successful in growing and maintaining a culture of the luminous bacteria so far.
I just spent the past week conducting the procedure to allow growth of bacteria on fish, with great success. The fish I caught off the pier, a
mackerel I believe, so it was as fresh as it was going to get. I immediately iced it to both kill and preserve the fish. I filled the container half
way (about 250 mL) with ocean water and added about 5 grams of salt to ensure that only the luminous bacteria would be growing on the fish and to also
make up for the addition of tap water from the ice. The container was left in another container which had ice water and was allowed to sit for 2
days.
I got a very strong glow in a completely dark room, and I just inoculated a 1000 mL solution of 20 grams sodium chloride, 5 grams of yeast extract
that I prepared using a 50 mL of 5 molar sodium chloride lysing solution, 10 grams beef boullion (with delicious beef extract, yeast extract, salt,
hydrolyzed proteins, and fatty acids), and 0.5 grams of magnesium sulfate. I hope by Thursday I can get some results up. I can't get a clear picture
of the glowing fish on my digital camera, but I kept it outside to see if it'll have an even stronger glow. Updates soon!
As an adolescent I aspired to lasting fame, I craved factual certainty, and I thirsted for a meaningful vision of human life -- so I became a
scientist. This is like becoming an archbishop so you can meet girls.
-- Matt Cartmill
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chemoleo
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Fantastic, that is good news!
Just make long exposures (>1 sec) and I'm sure you'll get good images, and do post with reference objects! Even cooler if you can grow it in some
sort of dish!
Well done!
Never Stop to Begin, and Never Begin to Stop...
Tolerance is good. But not with the intolerant! (Wilhelm Busch)
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bereal511
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I don't think my camera has a long exposure function. Which is unfortunate, because the bacteria was out-competed, so no more glow. I still have the
inoculated culture medium though, so I'm placing my bets on that. I just wish I had isolated it on an agar plate first, but I have no plates.
Hopefully the conditions will be good enough that the luminous bacteria aren't killed off by other bacteria. Otherwise I'll start over with a culture
plate.
As an adolescent I aspired to lasting fame, I craved factual certainty, and I thirsted for a meaningful vision of human life -- so I became a
scientist. This is like becoming an archbishop so you can meet girls.
-- Matt Cartmill
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Spargine
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Has anyone had success with fairly inland fresh fish? I am thinking about attempting this and I am wondering if anyone else has had success.
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chemoleo
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I don't think there are any phototrophic bacteria known that grow in fresh water...
Bereal - well next time make a glycerol stock, to conserve the bacteria! Shame you lost them 
With a glycerol stock, you just dip a sterile needle into the first successful culture, which is then dipped into to sterile (heated prior to this)
50/50 glycerol/water.
Then you should be able to restart your growth as many times you wish!
Never Stop to Begin, and Never Begin to Stop...
Tolerance is good. But not with the intolerant! (Wilhelm Busch)
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ChrisWhewell
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Quote: Originally posted by chemoleo  | I read something very interesting recently - I got a pdf but can't remember where I got it from. Anyway, here it goes:
| Quote: | PASSENGERS on board cruise liners are sometimes startled by an eerie phosphorescent glow emitted by seawater flush lavatories. The explanation lies in
luminous bacteria, which require oxygen for their light-generating reactions. Luminescent bacteria are present in all marine environments and develop
abundantly on the surface of decaying fish. Their presence can be a problem — most of
the research on marine bioluminescence has been funded by the US Navy, embarrassed by glowing trails behind surface vessels, and more recently by
bacterial interference
with the laser light used for submarine communications.
|
Upon searching some more:
The bacterium causing luminescence is Vibrio (Photobacterium) phosphoreum, it thrives between 20 and 30 deg C, and grows
well at saline conditions. The bacterium requires oxygen to produce light, and the reaction roughly works as a luciferase (enzyme) - FADH2
(flavo-cofactor) complex, where light is emitted once it reacts with O2 and RCH2OH to form RCOOH and H2O. Light is emitted in the bluish green range
(460–490 nm).
THen, at http://www.disknet.com/indiana_biolab/b203.htm it says:
It grows on glucose, mannose, fructose, or glycerol as a sole carbon source, and synthetic or real seawater as a growth medium.
ISOLATION :
Incubate fresh fish, squid, or octopus at 10-15 deg Celcius, half submerged in natural or synthetic seawater (I take it this shouldn't be sterile
seawater! This will stink, so do it outside!). Examine at half-day or daily intervals for glowing colonies of bacteria (in total darkness after your
eyes are dark adapted).
Streak these colonies on LM (Luminescence Medium). LM is synthetic or natural seawater containing per liter: 3 ml glycerol
(glycerine), 5 gram yeast extract, 5 g tryptone, 1 g CaCO3 (powdered limestone or chalk) and 10 g of agar (per litre). For streaking, sterile
toothpics can be used (i.e. soak them in ethanol and dry over flame, and use immediately after they cooled down)
I looked up agar, and it is a linear complex sugar made from beta-galactopyranose linked to 3,6-anhydro-L-galactopyranose. Basically, its just a
polysaccharide/carbohydrate, or complex sugar. I should think any other complex sugar would do, such as starch, alginate (a linear complex sugar made
from beta-D-mannuronic acid and alpha-L-guloronic acid) etc.
Regarding tryptone, this is a pancreatic digest of casein, a protein. I should think any protein extract should do, you will get that in Health
shops/Fitness shops etc.
Yeast extract is similarly easy to get, it is basically a whole cell lysate of yeat, and thus contains vitamins, protein, DNA/RNA, lipids, minerals
etc. Isn't Marmite a yeast extract?
Anyway, back to the isolation of Photobacterium phosphoreum!
So, once you have streaked luminous colonies onto the plates, you can either grow them at room temp. or at 4 deg C (fridge). With the latter, check
every day for growth by looking at the plates in the dark!
For growth in liquid medium, just leave out the agar (whcih is the gelling agent).
So I would first grow the bacteria on the fish, and meanwhile prepare covered plates with the growth medium. Alternateively, use a small bottle that
is sterile (Ethanol-rinsed) and poor in the hot growth medium. Let it cool down, and once at room temperature, add a bit of the luminous colonies from
the fish. Only very very little is needed! Even if you dont see anything you have picked up on the tooth pick, there still will be zillions of
bacteria on it!
Then grow the bacteria for a day or two, take out a small amount and bubble it with air! Luminescence will ensue! Apparently, luminescence can be so
bright that you can read a newspaper with it in the dark!
How cool is that!!
Anyway, apparently it is better to use fresh fish from a fish market next shore lines, simply because transport wasn't so long and the fish are fresh.
Water from a salt water aquarium gives varying, but usually poor results. If you live inland and happen to be on a seacoast, place a fresh squid in a
plastic bag with trapped air and place it in crushed ice for storage for the trip home. For better results place fish or part of a fish in a jar that
is pushed into crushed ice in a cooler. The purpose of the jar is to protect any growing colonies from smearing. By the time you get home glowing
colonies may be present. After one or two days at 4 to 18C, luminous colonies may be present. Examine the squid or fish daily. Incubation at 15 to 20C
may be the best temperature. Or use your home refrigerator which is about 4 C or above.
By the way, Photobacterium phosphoreum is the strongest light emitting bacterium known!
Whoever tries this gets the Mad Biochemist award!
PS An alternative, but similar growth medium is
30 g NaCl
1 g Glycerol
10 g Peptone (Bacto-Peptone)
3 g Beef Extract
(to obtain solid medium you should add 15 g of Bacto-Agar)
Dilute the ingredients in water (pH should be about 7,3)
Fill with H2O up till 1000 ml
PS2 At AEM Publications, I found a paper 'Isolation and Identification of Photobacterium phosphoreum from an Unexpected Niche: Migrating Salmon' by
Budsberg et al. they use 0.38 M NaCl, 0.02 M MgCl2 · 6H2O, 0.25 M MgSO4 · 7H2O, 8 mM KCl, 0.5% peptone, 0.3% yeast extract, 0.3% glycerol as a
growth medium. The first few are artificial seawater, which by itself is 0.4 M NaCl, 0.1 M MgSO4 · 7H2O, 0.02 M KCl, 0.02 M CaCl2 · 2H2O.
[Edited on 7-2-2004 by chemoleo] |
Awesome. they ought to coat the toilets with a triboluminescent zinc layer
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12332123
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I know this thread is somewhat old, but I just wanted to recount an experiment we did in biology last year (14 yo). We inserted bacterial plasmids
engineered to contain antibiotic resistance and a gene from jellyfish which produces a UV fluorescent protein into some bacteria, then killed off the
bacteria which had not taken up the plasmid using an antibiotic solution (very clever IMHO). This required a very specific procedure involving cold
shock and I didn't get any colonisation but it worked for some other groups. I
remember my biology teacher saying that the required materials cost several hundred pounds.
[Edited on 15-4-2010 by 12332123]
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The WiZard is In
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The most common source of luminescence in da sea
dobe luminous alga e.g., Pyrocystis luna and Pyrocystis noctiluca.
This may take all the fun of self discovery out -
Wards sells a —
Fascinate Students with One of the Simplest Light-Producing Organisms Luminescent Bacteria Living Specimen Set
These glowing green and bluish-green examples of bioluminescence make a fascinating addition to microbiology studies. Cultures include: Azotobacter
vinelandii and Vibrio fischeri. The set comes with photobacteria medium, instructions, and the Working with Bacteria manual.
http://wardsci.com/product.asp?pn=IG0007238
If the back of whats left of my mind was --- Difco sold photobacteria medium, however, I find not it listed.
Run Photobactium medium through Google or same such.
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The WiZard is In
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Let me introduce a dirty word here — Book.
People on la Net spit and scream when someone suggests
a book and absolutely have sissy-fits if you suggest they
to the (an even dirtier word) Library.
I would mention in passing :—
E Newton Harvey's 692 page opus.
A History of Luminescence : From the Earliest Times Until 1900
American Philosophical Society
Philadelphia 1957
He discusses luminescence in all it variations, e.g.,
The Journal of industrial and Engineering Chemistry
2 [11] 478-9. November, 1910.
Tribo-Luminescence. [1]
The term “luminescence” has been applied to the light which is emitted by various substance at a temperature much lower than that which is
necessary to show visible incandescence, as illustrated for example in the light of the firefly, decaying wood, and many natural and artificial
mineral compounds.
The general phenomenon of luminescence has been divided into several classes by Dr. Sylvanus P. Thompson, as shown in the following tabulation which
is copied from his book entitled "Light Visible and Invisible," page 175 (1893).
Phenomenon. Substance in which it occurs.
1. Chemi-luminescence………... Phosphorus oxidizing in moist air;
decaying wood; decaying fish; glow-worm;
firefly; marine organisms, etc.
2. Photo-luminescence
(a) Transient: Fluorescence............Fluor-spar; uranium-glass; quinine;
scheelite; platino-cyanides of vari-ous
bases; eosin and many coal-tar products.
(b) Persistent: Phosphorescence…. Bologna-stone; Canton's phosphorus
and other sulphides of alkaline earths;
some diamonds; etc.
3. Thermo-luminescence ……………. Fluor-spar; scheelite.
4. Tribo-luminescence……………. Diamonds; sugar, quartz; uranyl
nitrate; pentadecyl-paratolylketone.
5. Electro-luminescence
(a) Effuvio-luminescence Many rarefied gases; many of the
fluorescent and phosphorescent bodies.
(b) Kathodo-luminescence Rubies; glass; diamonds; many gents
and minerals.
6. Crystallo-luminescence………… Arsenious acid.
7. Lyo-luminescence…………… Sub-chlorides of alkali metals.
8. X-luminescence…………….… Platinoeyanides; scheelite; etc.'
Light Visible and Invisible can be DL'd from Google.com/books.
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Sedit
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| Quote: Originally posted by The WiZard is In | Let me introduce a dirty word here — Book.
People on la Net spit and scream when someone suggests
a book and absolutely have sissy-fits if you suggest they
to the (an even dirtier word) Library.
|
Well, Welcome to Science madness where we will spit and scream if your ass don't read a book before speaking.
Knowledge is useless to useless people...
"I see a lot of patterns in our behavior as a nation that parallel a lot of other historical processes. The fall of Rome, the fall of Germany — the
fall of the ruling country, the people who think they can do whatever they want without anybody else's consent. I've seen this story
before."~Maynard James Keenan
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peach
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Not a bacteria, but here's Mycena luxaeterna from the mushroom kingdom doing it out in the wild.
And Mycena chlorophos.
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ScienceSquirrel
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13-3-2012 at 05:36 |
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