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Jan Witkowski questions Alexis Carrel's experiment


How Alexis Carrel obtained his anomalous results
Leonard Hayflick Scientist
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At that time, trypsin was not used to prepare cultures. You took, in this case, a tiny piece of a beating chick heart from an embryo and a scrap of tissue no bigger than a pinhead. You then placed it on a glass slide or in a culture vessel. And indeed Carrel himself invented a vessel that bears his name, it's called a 'Carrel vessel'. It's no longer used. It's a drum-shaped glass container with a neck slanting upwards from the rim of the drum and it makes culturing a bit more easy than the vessels that were used previous to that invention. However, those are now obsolete.

The cultures then were prepared by putting one drop of the extract of a chick embryo... you would take a chick embryo, put it in the barrel of a syringe before it formed bones so it was all soft tissue, press the plunger of the syringe barrel... down the syringe barrel and, of course, that would macerate the tissue, you would collect it in a test tube half filled with the media that I described earlier. However, it didn't contain serum, it would just contain the salts that were necessary for – or that were thought necessary – for growth. That media... that part of the media is called 'balanced salt solution': sodium, potassium, calcium, etc.

And that mixture of macerated chick embryo and the media is then centrifuged – spun at a high rate of speed so that the heavy particles settle to the bottom – and then you remove what's called the 'supernatant fluid', which is the clear fluid on top of the packed garbage at the bottom of the test tube. And you use that fluid which contains, hopefully, all of the nutrients extracted from the chick embryo that a chick embryo cell would need. Perfect logic and, in fact, it does work.

So, one drop of that is put on top of this pinhead-sized piece of chick heart or any other tissue and then you add a drop of blood plasma. Blood plasma still contains the clotting elements and when you put that drop of chick plasma onto the drop of chick embryo extract, a clot forms, like a blood clot. However, there are no red blood cells there; there are very few left from the embryo and none in the plasma. So it's a clear clot and it acts as a glue gluing the scrap of tissue onto the surface of the slide and, simultaneously, providing the cells with all the nutrients they need. Put into the incubator and then what happens is rather complicated but for this story, what's important to know is that the cells begin to divide at the edge, around the rim of this explant, which is the technical name for it, an explant.

So you now have cells dividing, you can see under the microscope, coming out as a sheet of cells as they divide in the perimeter of this explant. After a week or so, the cells have divided to the point that they reach the perimeter of the clot and now you need to subculture. The way you do it, using this primitive technology, was to take sterile scalpels, cut the pie-shaped culture into quarters, remove each quarter and embed them each in a separate clot, as described, in four different vessels. And, of course, you can repeat that, as Carrel did, for some 35 or 40 years. And this was the way this culture was carried.

When this woman became knowledgeable in Carrel's laboratory about this culture and looked at it one day when it had just been... after a couple of days after transplantation, she called over her superior and said, 'Look here, what's going on? I see at the margin of this clot, a tiny island of cells separated by a millimetre or more from the main culture in the centre of the clot, and I know from my experience in this laboratory that cells do not migrate away from the centre explant and then build another mini explant or mini collection of cells one millimetre away with nothing between.'

'Oh', said the superior, her superior, 'don't worry about that, that's nothing. We don't worry about that.' And so this woman took the advice of her superior and after several months or years, she became more and more puzzled by this event that she observed many times and couldn't figure out how it happened. By this time it was 1929, stock market crash, people... 20 million people are out of work and after our young lady said for the umpteenth time, 'What's happening in these cultures', her superior said, 'If you want to maintain your job here, you will ignore what you are wanting to discuss.' Well, of course, everybody wanted to maintain their work and she did as directed.

And this is the story that she told me and she ended by saying, 'Your interpretation of Carrel's work is correct and what I had said during my lecture is the following.' After reading his papers... although this certainly wasn't proof, it was suggested only. Carrel showed in many of his papers – he published several papers on this culture, maybe several dozen – each time he fed the cultures, twice a week I recall, in the next day or two, there was a burst of mitotic activity. Now, you could easily argue that that's because they were fed fresh media and that encouraged them to replicate. The other interpretation is that somehow, the culture was being re-fed with new cells and that was my interpretation because the method of centrifugation in those days was so primitive that the probability of fresh cells still remaining in the fluid portion of the embryo extract was great, and that each time they did a sub-cultivation and put in one drop of embryo extract, they were in fact adding fresh cells. And that would explain the observation that this woman technician made in respect to islands of cells distant from the actual culture in the centre of the clot.

Leonard Hayflick (b. 1928), the recipient of several research prizes and awards, including the 1991 Sandoz Prize for Gerontological Research, is known for his research in cell biology, virus vaccine development, and mycoplasmology. He also has studied the ageing process for more than thirty years. Hayflick is known for discovering that human cells divide for a limited number of times in vitro (refuting the contention by Alexis Carrel that normal body cells are immortal), which is known as the Hayflick limit, as well as developing the first normal human diploid cell strains for studies on human ageing and for research use throughout the world. He also made the first oral polio vaccine produced in a continuously propogated cell strain - work which contributed to significant virus vaccine development.

Listeners: Christopher Sykes

Christopher Sykes is a London-based television producer and director who has made a number of documentary films for BBC TV, Channel 4 and PBS.

Tags: Carrel's vessel, Alexis Carrel

Duration: 8 minutes, 25 seconds

Date story recorded: July 2011

Date story went live: 08 August 2012