Talmudic Question #53A

In recent years, the use of radioactive isotopes has diminished considerably in microbiological research. What has been lost in the process?

Commentary by Elio

Source.

This is a nostalgia trip. Molecular microbiologists of my generation relied on the use of radioactive isotopes for much of their research. Those days, the size of a department’s research effort could be measured by the number of scintillation counters it had at its disposal. Radioactivity was a convenient way of detecting and quantitating very small amounts of cell constituents. This has now been superseded by fluorescence and other non-radioactive methods. However, there are some things that radioactive isotopes do best. Thus, they are still in use. Here are two such uses.

1. Determining which molecules are made anew. Adding radioactive precursors of macromolecules to a growing culture will result in selective labeling of the newly synthesized molecules. If the labeled compound is added for a short time ("pulse labeling"), the radioactivity is found in the portion of the molecule most recently made. Among many other things, such methods were used to elucidate the direction of synthesis of proteins (from N- to C-terminus) and of nucleic acids (5'- to 3'-terminus). Moreover, with this strategy you could estimate the speed with which such macromolecules are synthesized.
2. Measuring the stability of a molecule. Adding a labeled precursor for some period of time, then replacing it with its non-radioactive counterpart (“pulse-chase”) lets us measure the stability of synthesized molecules. If the molecules are stable, they will retain the radioactive label; if unstable, the label will be lost. This method enabled us to determine the half-life of high-turnover molecules such as mRNA and certain regulatory proteins.

These days, when I mention radioactive isotopes, the response I get tends to focus on the dangers of working with them. Well, in the past we were not only careful but also aware of what constitutes danger. Mostly, we worked under safe conditions. Or so we thought. And I'm still around, many millicuries of P³² later.

Comments

[qetzal]

As a molecular biologist, I mostly miss the benefits of using 32P-end-labeling for detecting trace amounts of DNA in gels. Especially for things like gel shifts, there's nothing easier than 32P.

Well, nothing easier if you only consider the experiment itself. Once you factor in the regulatory, handling, & disposal requirements, things change.

In my limited experience, quantum dots are cool, but there are things you can do with 32P or other radionuclides that you can't possibly do with quantum dots.

As for lab safety, I think most molecular labs used small enough amounts that safety was not such a huge issue. Obviously, proper procedures are needed, but they're not really THAT hard to maintain. Safety for the folks who have to dispose of aggregated wastes from multiple labs may well be a bigger issue, though.

[Mark O. Martin]

Also, didn't Joshua Lederberg once suggest that we could look for "shadow life" (i.e., life that did not use nucleic acids) by enrichment in the presence of high levels of 32P?

Hmmm....

[David]

I want to know the answer to this question, too. I just started a job in a hospital research lab and had to go through radiation safety training, though I was informed I wouldn't be using radiation.

I know that one of my colleagues uses an irradiator to stop memory B cells from growing, but other than those kinds of cases, where is the need for radiation in today's microbiology/ID lab?

[Mark O. Martin]

It's certainly true that people of my "era" in biology became pretty paranoid (or cavalier) about radioactivity. It's good, I think, that hot isotopes are not used in the undergraduate classroom much these days.

Nanodots---also called quantum dots---may be used in the future in the way hot isotopes were used in the Good Old Days:

Frasco, MF and N Chaniotakis. (2009). "Bioconjugated quantum dots as fluorescent probes for bioanalytical applications." Anal. Bioanal. Chem. ---- this just came out on August 28th and is available electronically; PUBMED does not give a good record yet.

[Welkin]

Radioisotope use, in addition to its functional applications, always had unintended utility as a learning tool. Its use required students to become familiar with concepts like exponential decay and the poisson distribution, not to mention basic familiarity with the molecules themselves - which phosphate is attached where, which one carries the label, which linkage is involved? - and psychologically, the ominous chirping of the geiger counter uncovering unsuspected contamination of lab benches, shoes, floors, gloves, sharps, the doorknob on the lunchroom door, was always a humbling object lesson in laboratory safety.

Elio's response:
I hadn't thought of these extra benefits, but concur with you entirely. Understanding the metric (a popular term these days) of radioactivity was useful to me both in the lab and the classroom.

[AJ Cann]

Pollution? Landfill? Health risks?

[Mark O. Martin]

Well, I find that students have more trouble understanding that Hershey-Chase experiment. But maybe nanodots will come to the rescue.

[John S. Wilkins]

I can't help myself: alpha particles?