Most of my childhood notes and cryo-memrobilia were lost when my house burned down in September, of last year. So, regrettably, I can’t consult my notes from those experiments. However, as best I recall, the mortality rate in yeast frozen in distilled water was ~90%. No special treatment was required beyond removing them from the incubating medium and resuspending them in distilled water prior to freezing. Viability was determined indirectly by adding the frozen-thawed yeast in water to culture medium in an Erlenmeyer flask connected to a water displacement set-up very much like this:
http://herbarium.usu.edu/fungi/funfacts/respiration.jpg
I later repeated this experiment with red cells (my own) which is much more sensitive and directly quantative of cell survival. You do, however, need a centrifuge and related equiupment to measure microhematocrit—things I could easily acquire back in the day (and in fact, still have).
If people did hands-on biology in the same way and to the same extent they do hands-onelectronics and programming, we’d all likely be either “immortal,” or dead, by now.
Here is an experiment I am currently struggling to tool to do which may serve as an example. Recently, a very simple way was discovered to induce apoptosis in a significant fraction of senescent cells in vivo in rodents, and in human cell culture cells, as well: http://onlinelibrary.wiley.com/doi/10.1111/acel.12344/pdf. This results in partial rejuvenation of the animals because senescent cells release myriad toxic cytokines, chemokines and other pro-inflammatory and probably telomere shortening species. While there is as yet no evidence that eliminating senescent cells—or reducing their number—will increase lifespan, there is ample evidence that it will greatly increase healthspan. This new class of drugs has been dubbed the “senolytics” by their discoverers, Zu and Tchkonia. The nice things about these two drugs is that they are both small molecules which are readily available, FDA approved/GRAS and have very low toxicity. One is the OTC nutrient quercetin, and the other is the relatively exotic molecularly targeted antineoplastic agent dasitinib, marketed under the name of Sprycell by Bristol-Meyers-Squibb.
In mice, one dose of these agents in combination was effective at reducing the senescent cell burden dramatically, with benefits lasting for 7 months. The cost of a dose of dasitinib for an adult human is about $400 - eminently affordable (the cost of the quercetin required is a few cents). So, what’s the problem? Well, if you are over 30, odds are that you have a significant burden of senesacent cells, and by the time you are 50, somewhere between 15 to 30% of your body mass may be senescent cells. In my days in ICU doing hemodialysis, I saw more than a few patients critically ill and in renal failure from something called “acute cell death syndrome” (ACDS) which most often resulted from chemotherapy given to lymphoma or leukemia patients too rapidly, resulting in a massive die-off of cancer cells. Large scale cell death is toxic and can be, and often is, lethal.
Animals treated with dasitinib+quercetin do not show signs of ACDS. However, careful monitoring of blood chemistrires during the treatment phase was not done and the animals so far studied were middle aged rodehts—not humans, and certainly not older, or elderly humans. Thus, additiional data are needed. In my opinion, dogs are ideal for such a study because they are available in abundance as old and very old (senile) animals, have large blood volumes which allow for harmless routine clinical laboratory evaluations, and have neurobehavioral faculties which are easily and reliably assessed by untrained humans. They also stand to benefit from the treatment if it does not prove lethal, or can be adjusted so that it is easily tolerated.
You have to “make” your own aged rodents and that takes years. And years are something many of us no longer have… Research begun now (or soon) will very likly yeild results that will be immediately clinically applicable to humans. Unfortunately, this research cannot practically be done anywhere in the West legally.
Actually, cryogenic vessels do not really fail, in the sense I think you mean, over time—with the notable exception of liquid helium and liquid hydrogen storage vessels. Liquid helium has bizzare effects of metal (in addition to quantum tunneling) causing high strength steel to embrittle over time. It is thoought that this occurs due to the presence of helium in solid solution in the metal subjected to loading, and being present at a temperature sufficiently low to form grain boundary cracks as a result of sliding along grain boundaries (which contain steps developed as a result of prior intragranular shear).
Hydrogen “embrittlement” is due to migration of lone hydrogen atoms into the metal where they re-combine in sub-micron sized voids in the metal matrix to form hydrogen molecules. In so doing, they create pressure from inside the cavity where they are located which can increase in vulnerable areas of the metal (e.g., where it has reduced ductility and tensile strength) to the point where the metal develops first micro-cracks and then a large, macro-fracture resulting in castastrophic failure.
Liquid nitrogen storage containers kept dry and free from liquid oxygen accumulation, and which remain stationary, can and do last “indefinitely.” They will require periodic re-hardening of the vacuum, but this is not due to structural failure, but rather is due to outgassing of materials from the reflective/convective barrier wrap and of hydrogen from hydrogen inclusions in the welds. If the units are not man-handled and are well cared for, there is essentially no work-hardening of the welds, or of the structural metal itself, and they may well last for many decades, or even centuries. If the nitrogen gas boil-off were used to create a dry nitrogen sheild around the exterior of the vessels, their lifespan would likely be in the range of many centuries. Work-hardening, hydrogen ingress into the metal from water condensed from the air and corrosion from atmospheric oxygen and water at the neck-tube are the three principal causes of structural cryogenic dewar failure. If the dewar is not moved about, and if water is eliminated from the environment, stainless steel dewars should last indefinitely. I’ve seen dewars in semen storage facilities that are 50 years old and have not yet required rehardening of their vacuum. Conversely, I’ve seen vessels in lab use and used to haul industrial gases fail after a few years, or even a few months of use. TLC is almost everything when it comes to liquid nitrogen dewars.
Probably the best example of how robust ultra-high pressure vessel engineering can be is to look to long range guns on battleships. These tubes are about 2″ in diameter shy of being big enough to hold an average human and can withstand pressures in the range Maxikov is talking about. These “vessels” also have a breech and operate under horrible conditions wih respect to heat and corrosion. And yet, failure is almost unheard of. When failure means the loss of a battle ship, failure is not an option; consider that one turret on a 20th century battleship, exclusive of the guns, cost ~$1.5 million, U.S. These guns were made with mid-1920′s technology and remained in service until the last decade of the past century. Then, there was the Paris-Geschütz (http://en.wikipedia.org/wiki/Paris_Gun) of Krupp, but that’s another story...