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In June 1935 Wendell Stanley, working at the Rockefeller Institute for Medical Research in Princeton, reported in the journal Science that he had turned tobacco mosaic virus into crystals. His short paper was titled "Isolation of a crystalline protein possessing the properties of tobacco-mosaic virus". Yet his crystals, dissolved and rubbed onto a leaf, still caused disease. That one result forced biologists to ask whether a virus is a living thing or a very large molecule.
How did Wendell Stanley crystallise tobacco mosaic virus?
When Stanley started in 1932, nobody knew whether viruses were made of protein, fat, carbohydrate or something else, he recalled in his 1946 Nobel lecture. He chose tobacco mosaic virus because infected plants gave plenty of infectious juice and the virus was unusually stable.
The digestive enzyme pepsin, which breaks down proteins, slowly destroyed the virus's power to infect. In 1934 he concluded that the virus "is a protein, or very closely associated with a protein, which may be hydrolyzed by pepsin". Tests with over one hundred other chemicals pointed the same way.
So he purified it the way chemists purify proteins. Here is the method as he described it in the lecture:
Infect young Turkish tobacco plants by rubbing a leaf with a virus-soaked gauze pad; cut and freeze them two or three weeks later.
Mince the frozen plants in a meat grinder, thaw, and press out the juice, keeping everything near 4 °C.
Adjust the juice to about pH 7 and filter it through a layer of celite.
Add ammonium sulphate to "salt out" the protein fraction that carries the virus. Redissolve it and precipitate again with less salt, which leaves inactive protein and pigment behind in the liquid. After two or three rounds the liquid runs almost colourless.
Lower the pH to about 4 so the virus protein settles out, then redissolve it at pH 7. This solution holds about 80 per cent of the virus in the starting material.
Add saturated ammonium sulphate until the solution turns slightly cloudy, then a little acetic acid to bring it to about pH 5. Tiny needle-shaped crystals form, giving the stirred liquid a satin-like sheen.
Why did virus crystals shock scientists?
For decades, viruses had been thought of as tiny living germs. In 1898 Beijerinck had called the tobacco mosaic agent a contagium vivum fluidum, a living infectious fluid. A crystal suggested the opposite: a pure substance with no life in it.
Stanley wrote that there was at first "great skepticism" that his crystals could really be the virus, "due chiefly to the old idea that viruses were living organisms". The tests in his Nobel lecture answered the doubters:
The crystalline material was about 500 times as infectious as the starting material.
One millilitre of solution holding only 10⁻⁹ grams of it usually caused infection.
The same crystals came from diseased tomato, petunia, spinach and phlox plants.
Heating to about 75 °C, or strong acid or alkali, wrecked the protein and the infectivity together.
So are viruses alive?
Viruses multiply and mutate inside living cells, he noted, but cannot grow on artificial media, and nobody had shown they had a metabolism of their own. In size they overlap with large protein molecules at one end and with accepted organisms at the other. His conclusion, in the lecture, was that "a sharp line dividing living from non-living things could not be drawn".
He also got one detail wrong. His 1935 paper called the crystals a protein. In December 1936 Pirie and his co-workers reported that nucleic acid could be isolated from the crystals, and Stanley reported the same a few days later. He doubted at first that the nucleic acid was a true part of the virus, but within months agreed the virus was a nucleoprotein whose nucleic acid could not be removed without losing activity. Electron microscope pictures later showed rods about 15 by 280 nanometres. For how those rods are built, readour story of Rosalind Franklin and tobacco mosaic virus.
Stanley shared half of the 1946 Nobel Prize in Chemistry with John Northrop "for their preparation of enzymes and virus proteins in a pure form".
How do you answer "are viruses living or non-living?" in an exam?
This question often sits next to the exceptions to cell theory. Use Stanley's experiment as evidence:
Non-living features: they can be crystallised like a chemical, and they cannot grow or multiply outside a living host cell.
Living features: inside a host cell they multiply, and they can mutate.
Conclusion: viruses sit on the border between living and non-living. Stanley's 1935 crystals of tobacco mosaic virus, which stayed infectious, are the classic evidence.
A model sentence you can adapt: "In 1935 Wendell Stanley crystallised tobacco mosaic virus, and the crystals remained infectious; this shows viruses behave like chemicals outside cells but like living things inside them."
If words like nucleoprotein or salting out are still hazy, the Fermi Concept Library gives each concept in biology and chemistry its own lesson, so you can fill the gap before you write the answer.
